diff --git "a/deduped/dedup_0775.jsonl" "b/deduped/dedup_0775.jsonl" new file mode 100644--- /dev/null +++ "b/deduped/dedup_0775.jsonl" @@ -0,0 +1,45 @@ +{"text": "P-glycoprotein expression was demonstrated in two human intestinal adenocarcinoma cell-lines by immunoprecipitation of a 170-180 kDa protein with monoclonal antibody JSB-1. Both HCT-8 and T84 formed functional epithelial cell layers of high transepithelial electrical resistance (greater than 700 omega.cm2) when grown on permeable matrices. These epithelial layers demonstrated vectorial secretion . These vectorial vinblastine secretions were sensitive to inhibition by verapamil. Passive transepithelial vinblastine permeation was limited by the presence of intercellular (tight) junctions, as demonstrated by the high transepithelial electrical resistance, and verapamil increased this passive vinblastine permeation concomitant with a reduction in the electrical resistance. Cellular vinblastine loading was significantly greater from the basal side, and this was also susceptible to inhibition by basal verapamil. The demonstration of vectorial transport of vinblastine in human intestinal colonic adenocarcinoma cell layers is direct evidence in favour of the hypothesis that the function of mdr1 in epithelial from the gastrointestinal tract is to promote detoxification by a process of epithelial secretion. This study also highlights that cellular vinblastine accumulation depends not only upon P-glycoprotein function, but also upon differential apparent membrane permeabilities and the presence of intercellular (tight) junctions that may restrict drug permeation and cellular accumulation to apical or basal membrane domains."} +{"text": "Patients with obesity, diabetes, and chronic kidney disease (CKD) are generally physically inactive, have a high mortality rate, and may benefit from an exercise program.2), and stage 2-4 CKD (estimated glomerular filtration rate [eGFR] 15-90 mL/min/1.73 m2 with persistent proteinuria). Subjects randomized to exercise underwent thrice weekly aerobic training for 6 followed by 18 weeks of supervised home exercise. The primary outcome variable was change in proteinuria.We performed a 24-week randomized controlled feasibility study comparing aerobic exercise plus optimal medical management to medical management alone in patients with type 2 diabetes, obesity . Caloric intake and body weight and composition also did not change with exercise training.Exercise training in obese diabetic patients with CKD is feasible and may have clinical benefits. A large-scale randomized controlled trial to determine the effects of exercise on renal functions, cardiovascular fitness, inflammation, and oxidative stress in diabetic patients with CKD is planned. Diabetes mellitus now affects approximately 250 million people worldwide, a figure expected to reach 400 million (approximately 7% of the adult population) by 2025 . This glRegular exercise with ,10 or wiThe present study seeks to investigate the hypothesis that increasing energy expenditure by aerobic exercise will have cardiovascular benefits, including decreased blood pressure, decreased heart rate, and increased exercise tolerance, and will result in a decrease in proteinuria in obese diabetic patients with CKD. We also wished to determine if exercise would result in weight loss and improved body composition, improved glucose and lipid control, and a decrease in inflammation.This was a 24-week randomized controlled pilot study. After baseline screening, subjects were randomized to an exercise or control group. Subjects randomized to exercise underwent thrice weekly training for 6 weeks in the research exercise laboratory at Hines VA Hospital, followed by 18 weeks of supervised home exercise. Both groups received standard of care medical treatment for diabetes and CKD, including diabetes education. The primary outcome variable was change in proteinuria. Exercise training parameters , hemoglobin, glycated hemoglobin, lipid profile, C-reactive protein (CRP), dietary caloric intake, and body weight and composition were secondary outcome variables. This research was carried out in compliance with the Helsinki Declaration and was approved by the Hines VA IRB. Written informed consent was obtained from each participant.2), and stage 2-4 CKD . They then underwent a symptom-limited, treadmill, exercise stress test. If the stress test was negative, the patient was continued in the study and asked to complete questionnaires, nutritional assessment, and laboratory tests. After all baseline data were obtained, if patients were still eligible, they were randomized using a 2 \u00d7 2 block randomization scheme to the exercise or control group. The above testing was repeated after 6 weeks (at the end of the training period) and 24 weeks (end of study).Individuals assigned to the exercise group received instruction in walking and proper walking shoe selection. An introductory session was completed to educate the patient on developing a walking program, familiarize him/her with the lab and practice walking on treadmill. Each exercise training session included 3 to 5 minutes of warm-up, range-of-motion exercises, interval training, cool-down, and post-exercise range-of-motion exercises. Each subject's training program was individualized and based on the results of the baseline exercise tests. Exercise intensities listed in Table To prevent exercise-induced hypoglycemia, patients were exercised one to two hours after a meal . A fingerstick blood glucose was performed prior to exercising; if the glucose was <110 mg/dL, 30 g of carbohydrate was administered prior to exercising.Control patients underwent the same testing battery but did not participate in any exercise training.Subjects exercised on a treadmill (Marquette Series 2000 treadmill) attached to an ECG monitor. Exercise began at 0% grade and a speed of 1.8 mph. Increases in percent grade occurred every 30 seconds, and, after the first six minutes, speed increased every three minutes. The following measurements were made during all exercise tests. Oxygen uptake was determined using the MedGraphics CPX/MAX/D\u2122 system for breath by breath analysis . Prior to each test, the analyzers were calibrated with reference gases and room air. Heart rate was derived from a standard ECG. A Marquette CASE (Cardiac Assessment System for Exercise Testing) system was used for continuous visual monitoring and recording ECG. A 12-lead ECG was taken every minute during all exercise tests. Blood pressure (BP) was determined by the auscultatory technique, using a sphygmomanometer, a pre-gauged adult cuff, and a stethoscope. The first and last Korotkoff sounds were recorded. BP was taken pre-exercise, every two minutes while the subject was exercising, and every minute post-exercise until the patient's BP approached baseline values.Subjects reported a food record at baseline and then were asked to keep 6 non-consecutive days of food records during the first 6 weeks and then every 3 weeks during weeks 7-24. Average daily intake in kilocalories was calculated using Food Processor SQL Edition . In addition, the Harris-Benedict equation (HBE) with appropriate adjustment for activity level was used to estimate energy requirements for each subject.Assessment of body fat/composition was measured by air displacement plethysmography (ADP) . After bLaboratory values measured included serum urea nitrogen and creatinine, urea and creatinine clearances , random urine albumin/creatinine and protein/creatinine ratios, 24-hour urine protein excretion, hemoglobin, glycated hemoglobin, lipid profile, and C-reactive protein (CRP). These tests were done at baseline, at the end of exercise training (6 weeks), and at study completion (24 weeks).We performed within-group and between-group comparisons using mixed-effect linear regression models that included indicators for group (control vs. intervention), time (in weeks), and an interaction term (group \u00d7 time). To address the possible correlation of an individual's measures over time, the models included a random effect. For analytic purposes, urine protein data were log transformed in order to normalize the data. A P-value of < 0.05 was taken to be significant.Twenty patients were recruited and signed the consent form. One subject did not qualify based on screening laboratory tests. Of the remaining 19 subjects, 6 were dropped from the study due to a positive stress test (2 patients), investigator decision (2 patients), patient wish (1 patient), and commencement of hemodialysis (1 patient). Thirteen subjects were then randomized, 7 to the exercise group and 6 to the control group. Two of the 6 control subjects were dropped subsequent to randomization and prior to any testing , leaving 4 subjects in the control group. All subjects were male; mean age was 66 (range 55-81).Data for the exercise and control group patients are given in Tables To our knowledge, this is the first randomized controlled trial that has examined the effects of exercise training in diabetic patients with CKD. However, there have been a few previous studies that have examined the effects of exercise on renal function in primarily non-diabetic CKD patients. Boyce et al. found, in a crossover-design trial, that four months of exercise training, via stationary cycling, in sixteen non-dialysis CKD subjects decreased blood pressure (systolic and diastolic), increased peak oxygen consumption, but had no effect on declining GFR . In anotDiabetic patients with CKD and overt albuminuria typically develop progressive kidney failure . TherefoWe also did not find any changes in glycated hemoglobin, lipid profile, or body weight or composition. Although all subjects did receive nutritional counseling, they did not specifically undergo caloric restriction or other forms of dietary modification. It is likely that a more prolonged and intensive exercise training program or an exercise program coupled with calorie restriction would have been required to result in weight loss and a decrease in fat mass. In this regard, long-term (1 year) aerobic coupled with resistance training has been reported to improve these parameters in type 2 diabetic patients without kidney disease and only mild obesity (BMI 30) . Renal fIn summary, we have demonstrated in a pilot study that exercise training in obese diabetic patients with CKD is feasible and may have clinical benefits. A larger-scale randomized controlled trial to determine the effects of exercise on renal functions, cardiovascular fitness, inflammation, and oxidative stress in diabetic patients with CKD is planned.The authors declare that they have no competing interests.DJL was responsible for the overall design and supervision of the study and manuscript preparation. IM helped to design the study and assisted in manuscript preparation. JPB and SQ enrolled patients and supervised exercise tests. CSJ was responsible for exercise training and data management. CC carried out the dietary studies. LCE served as the cardiology consultant. BMS did statistical analyses. EGC was responsible for administration of the research exercise laboratory and assisted in manuscript preparation. All authors read and approved the final manuscript."} +{"text": "This study aims to investigate the relationship between waist circumference and work-related injury in reference to the fourth Korea National Health and Nutrition Examination Survey.By analyzing data from the fourth Korea National Health and Nutrition Examination Survey conducted from 2007 to 2009, we estimated the rate of injury experience according to socioeconomic status, including occupational property, of 8,261 subjects. We performed logistic regression analysis with work-related injury experience rate as dependent variable and waist circumference as an independent variable, Odds ratios (OR) were calculated, which reflect the likelihood of work-related injury experience rate, and 95% confidence interval (95% CI) while controlling for relevant covariates with stratifying by sex, age, nature of injury, site of injury and occupational group.Among 797 persons who had injury experience over the past 1\u00a0year, 293 persons (36.8%) had work-related injury experience. After adjusting the confounding variables, the work-related injury was related to abnormal waist circumference . In subgroups, ORs were higher in men , professional, manager, and administrator . Higher rate of injuries were noted in back and waist , and transport accident had increased risk .Work-related injury rate differed depending on the waist circumference. The abdominal obesity was associated with higher risk of work-related injury. This study would be useful in selecting appropriate priorities for work-related injury management in Korea. Based on 2011 department of Korea statistics, generalized improvement in nutrient intake by Korean population has raised the rate of obesity. The rate was found to be higher in men 36.5%) than women (26.5%)6.5% than, and risInjury is disability of body caused by external factors, and injuries originate during work is defined as work-related injury. In KoreRecently, some claimed that obesity function as a protective factor in injury, but obeThere have been studies to identify obesity\u2019s relationship to injury in a work environment, and they reported that obese persons have about 26 to 107% more incidence of injury during working than people with normal weight,22-24. TIn this study, its purpose is to determine relationship between waist circumference and work-related injury. The authors want to apply outcomes found in this study to manage obesity for prevention of injury at work.The Korea National Health and Nutrition Examination Survey has been performed to identify Korean health and nutrition state since 1998. The fourth Korea National Health and Nutrition Examination Survey was conducted for 3\u00a0years from 2007 to 2009 with rolling survey sampling method. Each rolling sample was extracted from the survey, and it was designed to have independent and identical properties among samples. Year-round survey (50\u00a0weeks per year) was conducted, and 200 to 250 subjects were surveyed in a week. In each year, probability samples reflecting the whole country were extracted through stratified sampling in 3 stages. First stage involved stratification of 29 strata with a reference to 11 areas\u2019 population ratio by sex and age groups, and the samples were collected according to towns. Second stage extracted samples at district level, and final stage collected samples in family level. Number of participation to the survey was 23,632 (74.5%) among 31,705 national wide subjects. Final aIn this study, work-related injury was defined as an experience of injury for the past 1\u00a0year from the time of the fourth Korea National Health and Nutrition Examination Survey. Those who had answered yes to a question \"have you received a treatment in a hospital or emergency room due to injury or intoxication?\" were reviewed, and among them people who answered yes to \"during work\" in additional information section were finally defined as a work-related injury experiment.As demographic and socioeconomic characteristics, age, sex, waist circumference, alcohol intake, smoking state, exercise level, marital status, annual household income, and educational level were identified. Also, mechanism of injury and injury site were noted. We organized occupations into 6 groups according to major categorizations of the '6th Korean Standard Classification of Occupations\u2019; 'manager, professional, and administrator\u2019, 'clerk\u2019, 'service and sales worker\u2019, 'skilled agricultural, forestry, and fishery worker\u2019, 'craft, equipment, machine operating, and assembling worker\u2019, and 'elementary worker\u2019. AbdominIn this study, injury rates were calculated according to demographic and occupational characteristics of study subjects. Also, injury experience was set as a dependent variable, and abdominal obesity was used as an independent variable. Using the variables, logistic regression analysis was performed to calculate corrected odds ratio (OR) of work-related injury experience to population and 95% confidence interval (CI). Stratified analysis was carried out for age, sex, injury mechanism, injury site, and occupational groups. SAS 9.2 program was used for statistical analysis, and statistical significance was determined at 0.05 or below.Table\u00a0As listed in Table\u00a0Stratified multiple logistic regression analysis was made on age, sex, occupational groups, injury sites, and mechanism, and male with abnormal waist circumference had 1.42 times higher risk of injury during work (95% CI: 1.02\u2009~\u20091.98). For 'Manager, professional, and administrator\u2019 group, the risk of work-related injury was 2.41 times higher (95% CI: 1.10\u2009~\u20095.28) in people with abnormal waist circumference. The abnormal waist circumference group was 2.92 times higher risk of thoracic and lumbar spine injuries (95% CI: 1.49\u2009~\u20095.73). Motor vehicle accident also showed significant increase in risk by 1.60 times (95% CI: 1.60\u2009~\u20091.98) and 'Manager, professional, and administrator\u2019 groups showed significant relationship. Like previous studies that determined relationship between BMI and work-related injury, some suIn 2011, Canadian study on relationship between obesity and work-related injury noted that sprain, back and waist, lower extremity, and fall showed correlation to injury prevalence. In one This study showed that male with abdominal obesity had a higher injury risk than female. This supports previous studies which male with obesity had higher risks for motor vehicle accident and injury than female, and in turn the result reflects that obesity works differently to development of injury according to sex,36. MotoMeanwhile, not all occupational groups showed significant correlation between abnormal waist circumference and work-related injury. Only 'Manager, professional, and administrator\u2019 group showed significant increase of work-related injury risk by 2.41 times, and this suggests that injury is affected by waist circumference found in a specific occupational group. Like previous studies that discovered significant relationship between obesity and work-related injury in office workers, our resOne limitation in this study is that we were not able to remove recall bias, because the study is based on memory of subjects who answered the questionnaire about their experiences of work-related injury. In addition, thought a subject suffered from an injury, the subject may have been absent because he or she was admitted to a hospital during a visit by a surveyor for an interview. Other limitations include that occupational groups, injury mechanism, and site were not classified in detail and several variables are mixed together, limiting the recognition of injury\u2019s risk factors. Also, waist circumference was divided simply into normal and abnormal, and this simplification did not allow dose-reaction relationship. Further study that verifies distribution of injury risk in accordance with each waist circumference as continuous variable is required to determine clearer relationship. In addition, the study is insufficient to prove causal relationship between the circumference and injury because it was a cross-sectional study. Therefore, a prospective cohort study with a method that can verify risk factors in detail is needed for determining causal relationship between work-related injury and abdominal obesity, and with the study, it would be able to identify risk factors such as influence of chronic diseases, sleepiness and fatigue, and physical limitations that are suggested in previous studies.Despite of limitations present in this study, it again showed possible connection of abdominal obesity to work-related injury. This study is significant that waist circumference rather than BMI is presented as a predictive index to work-related injury. However, additional studies are needed to identify factors in waist circumference that affect work-related injury more clearly. With further refinement and learning about the risk factors, it would be possible to raise the need for obesity management to reduce occurrence of work-related injury in a corporation or a nation.The authors declare that they have no competing interests.SKK and SBK designed the research. SKK collected the data. SKK, HK and KL performed the statistical analysis. SKK, SSO, and SBK interpreted the data. SKK, JMS and JYS wrote the manuscript. All of the authors read and approved the final manuscript."} +{"text": "To the Editor: We report the case of a 27-year-old male Swiss tourist who spent 3.5 weeks vacationing in the vicinity of Tarapoto, a small city located in the rainforests of the Amazon Basin in northern Peru. An acute febrile illness developed in the man during the second week of his stay. Signs and symptoms of illness were chills, malaise, frontal headache, generalized myalgia, a self-limiting painful cervical and inguinal lymphadenopathy (lasting \u22481 week), slowly progressive and pronounced polyarthralgic pains of the peripheral joints, and a transient nonpruritic maculopapular rash (starting at the forearms \u22481 week after onset of fever and spreading to the trunk and later to the neck and face before fading after 3 days).The traveler sought care at the local hospital, where physicians diagnosed suspected dengue fever on the basis of the clinical signs and symptoms, and received symptomatic treatment with paracetamol. The fever and other signs subsided within 1 week, except the arthralgia, which did not improve. The polyarthritis initially was accompanied by swelling of the affected joints and showed a symmetric pattern, mainly affecting the small joints of the hands and feet as well as the wrists, ankles, and knees.Borrelia burgdorferi, Chlamydia trachomatis, Epstein-Barr virus, parvovirus B19, Salmonella Typhi, and S. Paratyphi, but none revealed a cause for the symptoms. Over almost 2 more months, the joint pains did not improve; thus, the patient was referred to a rheumatologic clinic and subsequently to the Swiss Tropical and Public Health Institute, Basel, Switzerland, for evaluation of a putative travel-related cause of the polyarthralgia.After returning home to Switzerland, the patient consulted his general practitioner because of persisting, incapacitating joint pains. The patient reported stiffness of the affected joints, mainly in the morning and after immobility. Physical examination of the affected joints did not reveal visible clinical signs of inflammation . Laboratory tests were performed for complete blood count, liver and kidney function, C-reactive protein, and serologic testing for dengue virus, chikungunya virus, Because of the patient\u2019s travel history, the course of the illness and clinical signs and symptoms experienced during the journey, and the evolution and characteristics of the persisting joint pains, we suspected an underlying Mayaro virus (MAYV) infection. Serologic testing for several alphaviruses were performed as described confirmeTogaviridae).Several viral infections can be accompanied by arthralgia. However, the most prominent and long-lasting polyarthritic symptoms occur in patients infected by alphaviruses that circulate among a wide variety of wild animals in relative mosquito vector\u2013specific and host-specific enzootic cycles; infection in humans (dead-end hosts) is almost exclusively incidental. Clinical cases and virus isolation have been reported only from northern South America, where MAYV circulates in an enzootic sylvatic cycle (similar to that for yellow fever) involving forest-dwelling MAYV was first isolated in Trinidad in 1954; since then, sporadic cases, clusters, outbreaks, and small epidemics of Mayaro fever have been reported from Brazil, Bolivia, Columbia, French Guiana, Guyana, Peru, Venezuela, and Surinam .Aedes aegypti mosquitoes (International travelers are rarely given a diagnosis of MAYV infection ("} +{"text": "Adelphocoris suturalis, which is one of the main cotton pests found in the 31 locations in China and Japan involved in the study. Results show that the species is highly differentiated between populations from central China and peripheral China regions. Analysis of molecular variance showed a high level of geographical differentiation at different hierarchical levels. Isolation-by-distance test showed no significant correlation between genetic distance and geographical distance among A. suturalis populations, which suggested gene flow is not restricted by distance. In seven peripheral populations, the high levels of genetic differentiation and the small Nem values implied that geographic barriers were more likely restrict gene flow. Neutrality tests and the Bayesian skyline plot suggested population expansion likely happened during the cooling transition between Last Interglacial and Last Glacial Maximum. All lines of evidence suggest that physical barriers, Pleistocene climatic oscillations and geographical heterogeneity have affected the population structure and distribution of this insect in China.Phylogeographic patterns of some extant plant and vertebrate species have been well studied; however, they are poorly understood in the majority of insects. The study documents analysis of mitochondrial and nuclear data from 419 individuals of Molecular phylogeographic studies provide valuable insights for exploring specific genetic structure, geographic modes, prevalent time scales, and demographic historySula leucogaster, S. sula). Morris-Pocock et al.et al.Alectoris chukar). Recent phylogeographic studies indicated that organisms were capable of responding to climate changes and adapting to different climatic conditionsPelophylax nigromaculataChilo suppressalisThe influence of physical barriers on genetic structure has been investigated in seabirds (Hemiptera: Miridae), is a widespread pest in East Asia . It is a major pest on cotton and several other crops in ChinaA. suturalis can attack a variety of plants (~115 species), primarily agricultural crops , pastures and weeds. It causes significant economic losses, and reduces yields and qualities of crops. In addition, this insect may easily reach critically important thresholds, switch host crops, or experience geographic spread because of its great tolerance to climate changeet al.1619The black striped plant bug, A. suturalis; (ii) examine the geographical pattern of haplotypes; (iii) infer the demographic history of A. suturalis. The genetic diversity was analyzed along with the levels of genetic variation within and among A. suturalis populations. Possible factors that affect the genetic variation were assessed and a reconstruction of the demographic history of A. suturalis populations developed.In this study, the geographical pattern of genetic variation for this species is assessed using three mitochondrial genes (mtDNA) and the nuclear internal transcribed spacer (rDNA ITS) region. The main goals of this study were to (i) analyze the genetic structure and phylogeography of Hd) within populations ranged from 0.619 to 1.000 with an average of 0.826 with nucleotide diversities (\u03c0) from 0.001 to 0.009.A final combined mitochondrial dataset included 2046\u2009bp of protein-coding regions . No insertions or deletions were detected among these fragments. Ninety-five haplotypes (including 70 unique haplotypes) and 107 polymorphic sites (including 58 parsimony sites and 49 singleton sites) were found among all samples . AverageFCT from K\u2009=\u20092 to 3, and a slight decrease of FCT values from K\u2009=\u20094 to 10. When K\u2009=\u20093, FCT was significant and up to the largest values ; K\u2009\u2265\u20094, the population structure disappeared with a single population assigned to a group (K\u2009=\u20093 was used as the population assignment scheme and predicted three groups corresponding well to three geographic regions. The first group (HJ) was the single population from Japan. The second group included 13 populations from Central China and North China Plain, one population (WZ) from Northwestern Plateau and one population (SY) from Northeastern China Plain. As most populations of the second group were at the relative central location of A. suturalis\u2019s distribution, corresponding to central China geographic regions simultaneously, this group was named the central China group (CC). The third group included the remaining 15 populations, of which 11 populations were from east coast, Northwestern China Plateau, Southwestern China Plateau, Northeastern China; four populations from North China Plain . Most populations located in the peripheral provinces of China, which were also at the peripheral location of pest distribution; therefore, this group was defined as the peripheral China group (PC).SAMOVA analysis showed a distinct increase in a group . TherefoFST value between PC and CC was FST\u2009=\u20090.380, P\u2009<\u20090.001; between HJ and PC was FST\u2009=\u20090.575, P\u2009<\u20090.001 and between HJ and CC was FST\u2009=\u20090.356, P\u2009<\u20090.001, respectively. These pairwise FST values suggested that the three defined groups were significantly differentiated. Some populations from the peripheral China group showed high FST values , indicating that a strong natural barrier to gene flow may exist in these populations and the two-level AMOVA analysis revealed the significant differentiation among populations (FST) and geographical distance among populations, and indicated that gene flow was not restricted by distance (Pairwise ulations . The thr<\u20090.001) . IBD tesdistance .Hd) were observed in dataset (ranging from 0.5824 to 1.0000 with an average of 0.857) , ITS2 (1096\u2009bp) and 28S (84\u2009bp)]. 130 haplotypes (including 109 unique haplotypes) and 125 polymorphic sites were found when gaps were treated as fifth state . 125 hapf 0.857) .FCT from K\u2009=\u20092 to 3, and FCT values continue to decrease from K\u2009=\u20094 to 10. When K\u2009>\u20092, the population structure disappeared with a single population assigned to a group .The genetic discontinuity analysis performed by BARRIER on the mtDNA dataset suggested that the hidden genetic barriers were correlated to genetic abruption. The five red lines for each population to adjacent and northern China regions , from northern China to northeastern China regions , and from Mt. Qinling regions to other regions .MIGRATE analyses indicated high levels of gene flow among populations, with the average value of 834.9 and the largest value of 1000.0. Asymmetrical gene flow was observed in 44 out of 930 pairwise comparisons. This was supported by non-overlapping 95% confidence intervals for the estimate of migration parameter , the Nem values were much smaller (<2 migrants per generation) in the above seven populations compared to other populations (\u03b8) observed in these seven populations implied the potential effect of a small size founder population showed much higher levels of gene flow than the rest of the studied populations. Interestingly, when translated ulations . The respulation .Based on mtDNA data, the BI tree of haplotypes formed two clades supported by 100% and 67%, respectively . Clade AA. suturalis experienced at least two population expansion events. The central China populations from Mt. Qinling had a relatively high proportion (71.76%) of shared haplotypes , whereas other populations had 60% shared haplotypes. This would suggest that populations from Mt. Qinling region might have established relatively early in these areas. Most Japanese haplotypes have the long-branch length in network suggesting that Japanese populations may have remained stable after colonization. That more haplotypes were found in central China than peripheral China suggested the existence of more relatively suitable habitats in central China.In the haplotype network, the ancient haplotypes generally should be located at the center of the network and have more widespread distributions, while the recent haplotypes should be at the tips of the network and be localized geographicallyA. suturalis experienced population expansion events.Similar to the results of mtDNA, the ITS network (gaps as \u201c5th\u201d state) presented two of the most frequent haplotypes H5 and H8 . H5 was A. suturalis populations were inferred using neutrality tests. When all samples were pooled as one group or defined into three respective groups , Fu\u2019s Fs, Fu and Li\u2019s D* and Fu and Li\u2019s F* were all significantly negative. However, the significantly negative values were not observed in the Japanese population of the mitochondrial clades ranged from 67700 to 359200 years before present. The beginning of rapid population growth of A. suturalis was around 50000 years ago, and then the demographic trend plateaued about 25000 years ago [i.e. population expansion occurred during the transition from Last Interglacial (LIG) to Last Glacial Maximum (LGM)] (m (LGM)] .FST values and AMOVA analysis showed significant genetic differentiation (FST) among three defined groups based on the analyses of mtDNA. However, the SAMOVA analyses showed a lack of genetic structure among A. suturalis populations using the ITS dataset. It is suggested that the cause of homogeneity in A. suturalis is the high level of gene flow. Compared to protein-coding mtDNA markers, non-coding ITS markers are expected to be more sensitive at detecting gene flow among interbreeding populationsPairwise FST values among groups support population differentiation in these populations in Beijing. A genotype differentiation study of Helicoverpa armigera, once a key cotton pest, indicated that existence of a very small number of native individuals in Beijing areaHigh genetic differentiation was observed between Beijing (BJ) and other populations. ITS and mtDNA data presented a geographic barrier in this area (Myricaria spp. (Tamaricaceae) showed that the region worked as a natural dispersal barrier, which promoted strong population structureHigh genetic differentiation between Changde (CD) and other populations was observed. Nuclear data show a barrier separating this population from central China populations . The intA. suturalis. Suihua is located in the Sanjiang Plain, the largest marshland area of China, and Tieling is located in the Liaohe River Plain in which flooding normally occurs that could influence or delay the development of A. suturalis. Haplotype diversity of TL and SH populations are significantly lower than other populations (P\u2009<\u20090.05), except for three eastern China populations . The ITS data showed a geographic barrier separating Suihua from northeastern China populations. The Songhua River might act as a physical barrier separating Suihua from northeastern populations. Songyuan located in Songnen Plain with superior resources is a suitable eco-geographic condition may allow A. suturalis to adapt to new habitats quickly under natural selection. Indeed, high genetic diversity further supports this hypothesis.Tieling (TL), Suihua (SH) and Songyuan (SY) are located in northeastern China which is a mega-diversity region, with complex topography in the latitude of 40-50\u00b0N, which includes the Northeast Plain and major mountain ranges, e.g. Mt. Changbai. High genetic differentiation was shown among Tieling, Suihua and other populations, which might be explained by local suboptimal ecological conditions for A. suturalis populations experienced population expansion, although a mutation rate of 0.0115 in recently diverged haplotypes at the population level is higher than the phylogenetic rates for some speciesA. suturalis, the historical population trend inferred by the Bayesian skyline plot seems to fit the climate trend relatively well during the Pleistocene periods. A. suturalis started expanding about 50000 years ago, which appeared during the Marine Isotope Stage 3 . In East Asian, this stage is a warm and humid period with temperatures about 2\u20135\u2009\u00b0C warmer than presentA. suturalis\u2019s physiological activity, but also provide suitable conditions for A. suturalis in relatively arid regions of Northwest China, increasing the number of habitats in Mainland China. In addition, the study of Jiang et al.Tsuga, Gramineae, Chenopodiaceae and Cyperaceae of the entire section. As Gramineae and Chenopodiaceae are the primary wild hosts of A. suturalis, the increase in population size of these species would distinctly improve the fecundity and survival capacity of the insect.Selective neutrality, subdivision or expansion can be distinguished by statistical tests of neutrality of mutations. The results of neutrality tests indicated A. suturalis, however, genetic analyses showed that the species\u2019 Ne was not reduced. A similar situation has been found in a rodent Microtus montanus, the population size of which decreased at times of climatic oscillations, whereas Ne remained stable throughout the past 2500 yearsNe can be highly inflated due to other factors that hinder species\u2019 dispersal among subpopulationset al.A. suturalis among expanded populations during the warmer interglacial period. However, the biological characteristics of the species, for example, high population growth rate, switching host crops, great tolerance to climate change, and strong dispersal capacity, might have resulted in a large effective population of this species through the LGM period.With regard to climate cooling during the LGM period, we expected that profound ecological upheavals would reduce the population size of A. suturalis is not known, but based on the genetic data presented here, it is possible to speculate the region of origin. Genetic data show that the coastline area facing the Yellow Sea, where the bug has been likely established for a longer period of time, to a very large mainland region that has been colonized in the recent years. Population expansion time and haplotype data support the idea that the insect is most likely originated in China\u2019s eastern coast region.The above analyses show that demographic expansion of the bug is concordant with the records of palaeogeography, palaeovegetation and palaeoclimate in the area. However, the geographic origin of A. suturalis. Beck & ReeseA. suturalis, such as Medicago sativa, and Melilotus albus. Furthermore, the planting area of the most important crop host of A. suturalis, the transgenic Bt (Bacillus thuringiensis) cotton, has obviously expanded from approximately 10000 ha to more than 3.7 million ha since 1997Malus spp. , Prunus spp. , Vitis vinifera, and Ziziphus jujube, and the extensive commercial cultivation of these trees in recent years may also contribute to the expansion of the insect territory.Regarding the recent population expansion into the present range, we suggest that the more recent expansion of primary hosts have provided a new opportunity for A. suturalis. One refugium was the eastern coast of China. Reconstructed LGM vegetation shows that the cool-temperate deciduous forests of today were the major vegetation type within the range of extant A. suturalisA. suturalis is the result of range expansion from the Southeastern China subregion during the last interglacial period. The fact that populations from Southeastern China subregion began to expand much earlier than other regions, along with the asymmetric gene flow from Southeastern China to other regions, and the relatively high haplotype diversity all imply that this region might have acted as the ice-age refugium. From this refugium, north- and northeastward expansion of A. suturalis along with the expansion of the deciduous forests might have occurred in these regions, which form the current distribution pattern. This distribution pattern of A. suturalis was consistent with the phylogeographical pattern observed in deciduous forests within these regions. For example, in the direction of north- and northeastward range expansion, our genetic data showed a substantial impoverishment of genetic diversity in some populations ; two northern China populations ; two populations from northeastern China (TL and SH).Based on the palaeoclimatic and palaeovegetational evidence as well as genetic data presented, we suggest that two independent refugia might have existed during the Pleistocene for A. suturalis in China. The Mt. Qinling region underwent complex changes in climate and vegetation distributions throughout the last ice age cyclesWe suggest that another refugium might be located at Mt. Qinling (at c. 34\u00b0N), in the central distribution region of A. suturalis between Japanese and Chinese fauna realms, we did not draw substantial conclusions from our statistical data, due to sampling only the population from Japan, and a lack of sampling populations and sufficiently available sequences from adjacent countries . More samples from adjacent countries need to be collected to study the phylogeography of the species in these countries.Regarding the phylogeography of A total of 419 individuals were collected from 30 locations in China and one location in northern Japan between 2011 and 2013 . The linTotal genomic DNA was extracted from single adult insect using the DNeasy Blood and Tissue Kit . The abdomen of samples was removed prior to DNA extraction. Fragments of three mtDNA genes and nuclear ITS region were used as molecular markers for this study.The PCR amplifications of mitochondrial genes were performed using specific designed primers, while the ITS region was amplified using primer pairs P1Purified PCR products were sequenced in both directions with the ABI 3730xl DNA Analyzer at Ruibo Biotechnology Co., Ltd . All sequences have been deposited in GenBank under accession numbers KM981463-KM981497 for COI, KM981498-KM981628 for ITS2 plus 28S and 5.8S, KM981629-KM981667 for ND5, and KM981668-KM981704 for CYTB.http://its2.bioapps.biozentrum.uni-wuerzburg.de/. The number of polymorphic sites (S), haplotype diversity (Hd), nucleotide diversity (\u03c0), average number of nucleotide differences (K) and the number of haplotypes (Ht) were calculated using DnaSP 5.0Sequences of ITS and mtDNA markers were aligned independently using ClustalW implemented in Mega 6.0K values ranging from 2 to 10 and the 100 independent simulated annealing processes. This method is based on a simulated annealing process that maximizes the proportion of total genetic variance due to differences between groups, and leads to the identification of genetic barriers between the groups . One study reported on the configurations K with one or more single population groups fails to produce the group structureFCT that did not contain any single population group could be used to identify the groups of populations.The SAMOVA 1.0 programFST values, and population genetic structure was analyzed based on the analysis of molecular variance (AMOVA) in Arlequin. Arlequin was also used to calculate the genetic distance (FST) matrix to test for the presence of Isolation-by-distance (IBD) in the dataset. Google Earth (http://earth.google.com) was used to estimate the linear geographic distance (km) between the sampling locations. The significance was tested with the Mantel test employing 1000 randomizations in IBDWS 3.23The genetic differentiation among populations and defined groups were further estimated based on pairwise \u03b8\u2009=\u2009xNe\u03bc, where x is the inheritance parameter; Ne is the effective population size; \u03bc is the mutation rate per site per generation) and mutation-scaled migration rate . The effective number of migrants of each population per generation (Nem) is \u03b8M. Heating chain was set: 1.0, 1.5, 3.0, and 6.0. Four independent runs of 20000000 generations were conducted to examine the consistency of output results with the first 10000 generations discarded.In order to identify the hidden genetic barriers resulting from microevolutionary processes , the program BARRIER 2.2A. lineolatus was used as an outgroup due to its close relationship with A. suturalis. ML analyses with 1000 bootstrap replicates were performed in PhyML. For MrBayes analyses, separate partitions were created for each gene in the mitochondrial dataset with the best-fit model, which was determined using jModelTest 2.1Two datasets of mitochondrial and nuclear haplotypes were analyzed under both Bayesian inference (BI) and maximum likelihood (ML) analysis using MrBayes 3.2A. suturalis, population expansion time (\u03c4) and neutrality analysis of Fu\u2019s FsF*, Fu and Li\u2019s D*To investigate demographic history of 6 generations with sampling every 10000 generations. The results of the calculation were checked in Tracer 1.4http://beast.bio.ed.ac.uk/TreeAnnotator/) was used to summarize sample information after discarding the first 10% of the trees.The Bayesian skyline plot, a method for estimating past population dynamics from molecular sequences, implemented in BEAST 1.6How to cite this article: Zhang, L. et al. Phylogeographic structure of cotton pest Adelphocoris suturalis (Hemiptera: Miridae): strong subdivision in China inferred from mtDNA and rDNA ITS markers. Sci. Rep.5, 14009; doi: 10.1038/srep14009 (2015)."} +{"text": "When the Ga content increased to 15.52 at %, the film\u2019s conductivity improved again. Furthermore, all films presented an average transmittance exceeding 80% in the visible region. Regarding the film\u2019s electrical stability, GZO thermally treated below 200 \u00b0C exhibited no significant deterioration in electrical properties, but such treatment over 200 \u00b0C greatly reduced the film\u2019s conductivity. In normal atmospheric conditions, the conductivity of GZO films remained very stable at ambient temperature for more than 240 days.In this work, Ga-doped ZnO (GZO) thin films were deposited via radio frequency sputtering at room temperature. The influence of the Ga content on the film\u2019s optoelectronic properties as well as the film\u2019s electrical stability were investigated. The results showed that the film\u2019s crystallinity degraded with increasing Ga content. The film\u2019s conductivity was first enhanced due to the replacement of Zn They can be used in fields such as solar cells, flat-panel displays, light emitting diodes, and smart windows ,2,3,4,5. domains ,7,8. How domains ,10. ZnO- domains ,14,15,16 domains ,19,20,212O3 targets. The influences of varying the sputtering power applied to the Ga2O3 target on the composition and optoelectronic properties of the films were investigated. The film\u2019s thermal and time stabilities are also discussed.In this work, Ga-doped ZnO (GZO) thin films were prepared via radio frequency (rf) sputtering on ZnO and Ga2O3 target (PGa2O3) on the variation in Ga content in the films was first investigated. The results are shown in PGa2O3, Ga content initially increased slightly (PGa2O3 < 20 W) before rising significantly as PGa2O3 rose above 20 W. The actual Ga content of the films with various PGa2O3 is given in the inset table. ZnO with 0.60 at % Ga thin film was obtained when PGa2O3 was fixed at 20 W. When PGa2O3 was above 30 W, Ga content increased to more than 4.29 at %. As reported by H. Gomez [The influence of the sputtering power applied to the GaH. Gomez and R. WH. Gomez , the sol3+ (0.062 nm) is smaller than that of Zn2+ (0.074 nm) [2+ cations replaced by Ga3+ cations was at a very low level, so the contraction in lattice parameters was not considerable. However, when the content of Ga3+ cations exceeded 4.29 at %, the ZnO diffraction peaks moved to lower diffraction angles. This movement became more pronounced with the rise in Ga content owing to redundant Ga atoms entering into the ZnO lattice and forming Ga interstitial (Gai) defects, which enlarged the lattice parameters of ZnO. Moreover, as the extrinsic defects of (GaZn) (the substitution of Zn by Ga), and (Gai) were introduced into the ZnO films, the films\u2019 crystallinity degraded as the Ga content increased.The X-ray diffraction patterns of the ZnO thin films with varying amounts of Ga are shown in .074 nm) , the amoPGa2O3 = 50 W) was found to be inferior to those with less gallium (PGa2O3 = 30 W). Some amorphous domains (highlighted by red dotted lines) embedded in the crystallized ZnO are visible in 1, but none can be found in 1. Additionally, in comparing the selected area electron diffraction (SAED) patterns of these two films, the conversion of the diffraction spots (2) to the diffraction rings (2) indicated an obvious reduction in grain size, which degraded the film\u2019s crystallinity.The degradation of the film\u2019s crystallinity with increasing Ga content was revealed by TEM observation . The cryon spots a2 to theon rings b2 indica2+ cations were effectively replaced by Ga3+ cations. However, this improvement of the film\u2019s conductivity disappeared when the Ga3+ content increased to a range of 4.29\u20139.27 at % (Zone 2), due mainly to the Ga content exceeding its solubility in ZnO and neutralized gallium atoms separating from the ZnO lattice. These neutralized species could not contribute to the generation of free electrons and enhanced the scattering effect of the carriers in ZnO films. Therefore, the film\u2019s conductivity declined in this range. Upon a further increase in Ga content to 15.52 at % (Zone 3), the film\u2019s conductivity improved because the neutralized gallium atoms gathered together and formed Ga clusters due to the high Ga doping concentration. Given that the work functions of gallium and intrinsic ZnO are about 4.2 eV and 4.5 eV, respectively [The film\u2019s conductivity as a function of Ga content in ZnO films is shown in ectively , ohmic c2+ cations. Free electrons were derived from this replacement. Thus, the carrier concentration increased with rising gallium content. In Zone 2, the carrier concentration decreased due to an increasing number of gallium atoms separating from the ZnO lattice and forming neutral defects, which did not contribute to the generation of free electrons. Furthermore, the intrinsic donor defects of oxygen vacancies could be occupied by Ga atoms, which reduced the carrier concentration. In Zone 3, gallium atoms gathered into clusters, and more electrons could easily transfer from gallium clusters to ZnO films. Thus, the carrier concentration increased. The variation in the carrier concentration in the films with increasing Ga concentration was analyzed via Hall measurement, and the results are shown in DE) of Ga3+ cations in ZnO was also investigated (DE is defined as the ratio of the carrier concentration to the atomic ratio of Ga/(Ga + Zn), where the free electrons in ZnO films are mainly considered to be created by Ga substitution. As shown in In order to understand the effective doping amount and the electrical activity of gallium in the ZnO films, the doping efficiency (\u03b7stigated . \u03b7DE is Zn) and (Gai), and Ga clusters increased, and the light scattering by these defects and clusters was enhanced. Hence, a slight decrease in the film\u2019s transmittance was observed in this region. With further increases in Ga content, gallium clusters gathered together, increasing in size while decreasing in number. The film\u2019s transmittance then improved. In addition, with increases in Ga content, blue shifting of the absorption edge was observed, corresponding to the expansion of the film\u2019s band gap. According to Vagard\u2019s law mentioned in [2O3 is about 4.9 eV larger than that of ZnO, the band gap of Znx1-GaxO films should increase linearly as the Ga doping amount increases.The film\u2019s transmittance is shown in ioned in , becauseVO) in the ZnO film were occupied by oxygen in oxygenated conditions. As reported by A.F. Kohan [VO defects, the carrier scattering by these defects was restricted, so the carrier mobility improved.The film\u2019s electrical thermal stability is shown in F. Kohan , oxygen The melting point of gallium is only around 29.7 \u00b0C , and theThe film\u2019s electrical time stability is given in 2O3 target varied from 0 to 80 W. The power densities applied on ZnO and Ga2O3 targets are given in Ga-doped ZnO (GZO) thin films with a thickness of about 100 nm were deposited on Corning 1737F glass substrates at room temperature via rf sputtering. The substrate holder rotation rate was about 10 rpm. Pure Ar was used as the working gas, and the working pressure was about 0.67 Pa. The sputtering power applied to the ZnO target was fixed at 80 W, while the sputtering power applied to the GaThe thickness of the films was measured with a surface profilometer . The composition of the films was detected with an electron probe X-ray microanalyzer . The phase structures of the films were analyzed with an X-ray diffractometer using the thin film mode with an incidence angle of 1\u00b0. Hall effect analysis with van der Pauw\u2019s configuration was employed to investigate the electrical properties of the films. Finally, the transmittance of the films was measured with a UV-Vis spectrophotometer .GZO films were deposited via rf sputtering. The optoelectronic properties of the films as a function of Ga content were investigated. It was found that, when the gallium content was near 4.29 at %, ZnO/Ga films possess optimal electrical properties. All such ZnO films present high transmittance in the visible region, regardless of gallium content. In addition, due to the occupation of oxygen vacancies by oxygen atoms and the migration of gallium atoms in ZnO film, the film\u2019s electrical properties are degraded under high temperatures in oxygenated conditions. However, this degradation was not observed in the time stability examination. This result suggests that ZnO/Ga films can withstand long-term loads at ambient temperatures but are not suitable for applications in high-temperature environments."} +{"text": "According to our calculation, average absorbance over 94% is achieved in the wavelength range between 400 and 2500\u00a0nm for the proposed absorber. The excellent performance of the absorber can be attributed to the localized surface plasmon resonance as well as the Fabry-Perot resonance among the metal-dielectric-metal layers. We compare the absorbing efficiency of tungsten nanosphere absorber with absorbers consisting of the other metal nanoparticles and conclude that iron can be an alternative material for tungsten in solar energy systems for its excellent absorbing performance and the similar optical properties as tungsten. Besides, a flat multilayer absorber is designed for comparison, and it is also proved to have a good absorbing performance for solar light.We propose and theoretically investigate an efficient solar light absorber based on a multilayer structure consisting of tungsten nanoparticle layers and SiO Solar energy systems have drawn more and more attentions in recent decades due to the excessive consumption of traditional energy sources and seriously deteriorating environment situation. In solar energy systems, solar energy can be converted to electricity or thermal energy for different usages with minor pollution to the environment. However, the present solar energy systems, like thermophotovoltaic (TPV) systems, solar steam generation systems, solar water heating systems, are inefficient in energy conversion, and efficiency approaching 20% in appropriate optical condition has been theoretically predicted in TPV systems , which iBesides, the materials of absorbers should be cheap enough, which can provide the possibility of a wide production. However, many reported absorbers use noble metals in their structure. Near-perfect absorption can often be achieved in these absorbers within the range of visible light, but their absorbing performance out of this region is terrible \u201313. As tIn this paper, we propose a broadband solar light absorber based on the design of nanoparticle-dielectric multilayers and the application of tungsten and iron in the structure. The paper is arranged as follows. First, we will introduce the 3-D absorber and show the simulation results. Then, we will illustrate the absorbing mechanism of the absorber and compare this structure with the flat MDM structure to get a deeper insight. Further, there will be a discussion between iron nanoparticle absorber and tungsten nanoparticle absorber for their performance when applied in this structure.2 layer. The diameter of the nanoparticles is 20\u00a0nm, and there is no gap between the neighboring nanoparticles. The dielectric layer in the most top of the structure is used for protecting the metal particles from being oxidized. A unit cell of single-layer NPA is plotted in Fig.\u00a0The basic structure of the metal nanoparticle absorber (NPA) is depicted in Fig.\u00a02) and metal (tungsten) are both adopted from the experiment data [y direction with the polarization along the x direction. Therefore, the simulation period P is the same as the diameter of the metal nanoparticle (20\u00a0nm). The minimum mesh size is set as 0.1\u00a0nm. Periodical boundary condition is adopted for single unit cell in Fig.\u00a0A\u2009=\u20091\u2009\u2212\u2009R\u2009\u2212\u2009T, where R is the reflection and T is the transmission. The thickness of the metal substrate is set as 300\u00a0nm, which is much larger than its typical skin depth to avoid transmitting light. Thus, there is nearly no transmittance in the overall frequency range, and the absorbance of the absorber can be calculated as A\u2009=\u20091\u2009\u2212\u2009R.As for simulation, we use 3-D finite-difference time-domain (FDTD) method. The corresponding software is Lumerical FDTD. The refractive indexes of dielectric [z\u2009=\u2009115\u00a0nm plane . To bettCompared with the single-MD-pair NPA, the absorbing performance is greatly improved in the longer wavelength range for the NPA structure with multiple MD pairs. To illustrate this phenomenon, we plot the spatial electric distribution of the eight-MD-pair NPA structure at Fig.\u00a02 is set as 100\u00a0nm, which is the same as the NPA structure. With thicker metal layers, the absorbance of the FMA structure turns to decrease. The absorbance over 90% is achieved for the wavelength range from 400 to 1500\u00a0nm when hd\u2009=\u200910\u00a0nm. However, when the metal layer thickness hd is set as 20\u00a0nm, which is the same as the metal layer thickness of the NPA structure, the absorbing efficiency of FMA drops obviously. This can be easily understood, because when the metal layers are getting thicker, the reflectance of the structure is more obvious and the absorbance reduces as a result. The selective absorption of FMA is better than NPA. When the wavelength is over 2500\u00a0nm, the absorption is below 20%. Although there are lots of MDM absorbers proposed for solar light absorbing [To get a deeper insight of the NPA structure, we calculate the absorbing performance of a similar absorber\u2014FMA (flat MDM absorber, plotted in Fig.\u00a0bsorbing , 27\u201332, bsorbing , 32 and For the MDM absorbers, their absorbing abilities for light are often based on the Fabry-Perot resonance , 6, 33. This also happens to the NPA structure. For absorbing spectrum in Fig.\u00a0The metal we choose for this absorber is tungsten. In our previous work , we haveLike tungsten NPA structure, the absorbing spectrum of iron NPA structure also has a redshift with the increase of silica layer thickness hh (plotted in Fig.\u00a0For the NPA structure, the fabrication of such uniform small particles may be difficult. Therefore, well robustness is required for the proposed structure. We calculated the absorbing performance of structures consisting of different shapes and sizes in Fig.\u00a0Besides, the damping constant of tungsten nanoparticle is often larger than the bulk tungsten due to surface scattering and grain boundary effects. According to the data in reference , we recaSo far, we have discussed the NPA structure and FMA structure and their absorbing performances and absorbing mechanism and the metals that can be applied in them to reach high absorption. However, the applications of these absorbers may be different. In TPV system, the well selective absorbing characteristics are often required to reduce thermal emission from the solar absorber. So, multilayer NPA structures whose absorbing performances are plotted in Fig.\u00a02 layers on the top of a metal substrate. With eight MD layers applied, the absorber can have an absorbance over 90% for most of the wavelength range from 400 to 2500\u00a0nm. The absorbing efficiency of this absorber exceeds the absorbing efficiency of many other solar light absorbers, which provide much possibility for the absorber to be applied in solar energy systems like solar steam generation, solar water heating, and waste water treating systems. Also, we compare the NPA absorber with FMA and found that the excellent absorbing performance of NPA absorber results from LSPR and Fabry-Peort resonance. We further compare the absorbing performance of several common metal nanoparticle absorbers under the same structure parameters. Results show that iron can be a promising candidate material for tungsten in solar absorber. All these simulation results help to the design novel solar light absorbing cells in solar energy systems, and the absorbers we proposed are promising to be applied in the real applications.In summary, we have proposed a highly efficient broadband absorber consisting of tungsten nanoparticle layers and SiO"} +{"text": "LV and RV cavity dimensions and standard indices of LV and RV systolic and diastolic function were similar between groups. NH&PI demonstrated reduced peak LV mid circumferential \u03b5 and early diastolic SR, as well as reduced global radial \u03b5. There was reduced basal rotation at 25\u201335% systole, reduced apical rotation at 25\u201340% and 60\u2013100% systole and reduced twist at 85\u201395% systole in NH&PI athletes. There were no differences between the two groups in RV wall mechanics. When compared to Caucasian controls, NH&PI rugby players have a greater LV mass, MWT and RWT with concomitant reductions in circumferential and twist mechanics. This data acts to prompt further research in NH&PI athletes.The aim of this exploratory study was to define the Athletes Heart (AH) phenotype in Native Hawaiian & Pacific Islander (NH&PI) Rugby Football League (RFL) athletes. Specifically, (1) to describe conventional echocardiographic indices of left ventricle (LV) and right ventricle (RV) structure and function in NH&PI RFL players and matched RFL Caucasian controls (CC) and (2) to demonstrate LV and RV mechanics in these populations. Ethnicity is a contributory factor to the phenotypical expression of the AH. There are no data describing the cardiac phenotype in NH&PI athletes. Twenty-one male elite NH&PI RFL athletes were evaluated using conventional echocardiography and myocardial speckle tracking, allowing the assessment of global longitudinal strain (\u03b5) and\u00a0strain rate (SR); and basal, mid and global radial and circumferential \u03b5 and SR. Basal and apical rotation and twist were also assessed. Results were compared with age-matched Caucasian counterparts . LV mass [42\u2009\u00b1\u20099 versus 37\u2009\u00b1\u20094\u00a0g/(m The \u2018Athletes Heart\u2019 (AH) describes cardiac structural and functional adaptations that occur in response to chronic exercise training . AlthougEchocardiography is regarded as an important tool in differential diagnosis within athletic populations. Novel techniques such as speckle tracking echocardiography facilitates the assessment of global and regional cardiac mechanics to determine strain (\u03b5), strain rate (SR) and twist data in the AH , 4. The Current clinical criteria to distinguish AH from inherited cardiomyopathies are largely derived from Caucasian athletes . More reRugby Football League (RFL) athletes provide an ideal model for assessment of the AH due to moderate dynamic and static components of the sport . The recThe aim of this exploratory study is to investigate the AH phenotype in NH&PI RFL athletes. Specifically, (1) to describe conventional echocardiographic indices of LV and RV structure and function in NH&PI RFL players and matched RFL Caucasian controls (CC) and (2) to demonstrate LV and RV mechanics in these populations.Twenty-one male RFL athletes of NH&PI ethnicity were recruited into this cross-sectional study. Results were compared to a group of age-matched male elite RFL athletes of Caucasian ethnicity . All participants were free of known cardiovascular disease and avoided alcohol and caffeine consumption 24\u00a0h prior to data collection, whilst refraining from training for at least 6\u00a0h prior to the examinations. Ethics approval was granted by the Ethics Committee of Liverpool John Moores University.Blood pressure (BP), body mass and height were recorded. Body surface area (BSA) was calculated as previously described . All parA standard echocardiogram was undertaken by a single experienced sonographer using a commercially available ultrasound system and a 1.5-4\u00a0MHz phased array transducer. All images were acquired in accordance with the American Society of Echocardiography (ASE) guidelines . Images Standard measurements were made in accordance with ASE guidelines . LV lineplax, RVOT1 and RVOT2). The RV inflow was measured from a modified apical four chamber orientation and included the base (RVD1), the mid-level (RVD2) and the length (RVD3). RV diastolic area (RVDa) and RV systolic area (RVSa) were measured and RV fractional area change was calculated (RVFAC). RV wall thickness (RVWT) was measured from a sub-costal approach. Tricuspid annular plane systolic excursion (TAPSE) was measured and TDI allowed for assessment of RV lateral S\u2032, E\u2032 and A\u2032.The RV outflow tract (RVOT) was measured at three locations . The ROI was adjusted to encompass the whole myocardium.LV longitudinal \u03b5 and SR were assessed using the apical four-chamber, two-chamber and three-chamber views, providing a global value based on the average of 18 segments. The parasternal short-axis view allowed the assessment of LV basal circumferential and radial \u03b5 and SR. Peak values were averaged from six myocardial segments assessed at the level of the mitral valve, whilst the level at the papillary muscle provided the same data at mid-level. The average global circumferential and radial \u03b5 and SR was calculated from basal and mid values and hence 12 myocardial segments.Basal rotation was measured using the circumference of the LV at the mitral valve level. A parasternal short-axis view at the level of the apex, defined as the level just above the point of systolic cavity obliteration, was used to assess apical rotation. LV twist was calculated as the net difference between apical and basal rotation.RV longitudinal \u03b5 and SR were assessed using a modified four-chamber view. RV global longitudinal \u03b5 and SR were averaged from three myocardial segments with the ROI restricted to the lateral wall only.Raw data from STE assessment was exported into a spreadsheet and processed using cubic spline interpolation. Temporal data across systole and diastole in 5% increments of absolute cardiac cycle were defined. Average \u03b5 and SR values across each 5% increment were used to create temporal curves and graphs.Study data were collected and managed using REDCap electronic data capture tools hosted at Liverpool John Moores University . All ech2), heart rate (57\u2009\u00b1\u200912 and 55\u2009\u00b1\u20099\u00a0bpm), cardiac output (5.26\u2009\u00b1\u20091.09 and 4.8\u2009\u00b1\u20090.66\u00a0L/min), systolic BP (131\u2009\u00b1\u200910 and 129\u2009\u00b1\u20098\u00a0mmHg), diastolic BP (70\u2009\u00b1\u20097 and 69\u2009\u00b1\u20099\u00a0mmHg), and training hours (23\u2009\u00b1\u200910 and 23\u2009\u00b1\u20097\u00a0h/week). NH&PI athletes had a significantly larger body mass than the CC (103\u2009\u00b1\u200911 and 96\u2009\u00b1\u20098\u00a0kg). There were no differences between groups for ECG criteria. Normal ECG findings included sinus bradycardia , sinus arrhythmia , 1st degree AV block (NH&PI 19% and CC 5%), partial RBBB , early repolarisation and isolated criteria for LVH [Both the NH&PI and CC were similar (p\u2009>\u20090.05) in height (1.83\u2009\u00b1\u20090.06 and 1.83\u2009\u00b1\u20090.05\u00a0m), BSA were present for mid circumferential strain in systole, with systolic twist producing a trend towards significance (p\u2009=\u20090.058). There were also significant interactions during diastole for global and mid circumferential, and mid radial SR, as well as apical twist.Individual systolic and diastolic 5% increment analysis demonstrated global circumferential \u03b5 (late diastole), mid circumferential \u03b5 (late systole through to early diastole and late diastole) and mid radial \u03b5 (early systole) as lower in NH&PI athletes see Fig.\u00a0. LikewisThe main findings of this exploratory study are (1) NH&PI athletes have larger LVM, MWT, RWT and RVWT compared to CC with no between-group differences in standard LV and RV conventional functional indices and (2) there is evidence of reduced peak and temporal LV mechanics but no difference in RV mechanics in NH&PI athletes compared to CC.In the current study the NH&PI demonstrated larger LV mass and thicker LV walls than CC with no significant differences in LV cavity dimensions. The combination of larger LV mass, thicker LV walls and similar LV cavity dimensions in NH&PI meant that although none of the athletes met the absolute criteria for concentric remodelling, they had values closer to the cut-off . A similRV structure in NH&PI athletes followed a similar pattern to the phenotype seen in the LV with increased RVWT but similar RV cavity dimensions. Zaidi et al. investigThis type of AH morphology may exacerbate the difficulty in distinguishing physiology from pathology in NH&PI athletes . That asAlthough systolic function as determined by conventional imaging was similar between ethnic groups, reduced circumferential and radial mechanics in systole and diastole were observed in NH&PI athletes. It is difficult to directly attribute these findings to the differences in LV geometry but reduced peak circumferential \u03b5 has previously been reported in body builders who exhibited increased LV mass and wall thickness compared to a marathon runner cohort . It may In addition to circumferential mechanics we also observed significantly reduced apical and basal rotation in early systole consistent with an extended pre-twist period in the NH&PI. A reduction in apical rotation throughout systole resulted in lower twist mechanics. It could be suggested that the alteration in early diastolic SR in the NH&PI could be a consequence of the reduced apical rotation observed. Apical twist is considered an important component of LV function to store additional potential energy that is released to increase early diastolic suction . This \u201crOur findings of similar RV mechanics in the presence of increased RVWT in NH&PI athletes is reassuring and highlights a similar response to the LV in this ethnic group. It is important to note that it is not possible to accurately assess RV circumferential function and therefore a comprehensive RV functional assessment may provide further insight.This study is limited by a small sample size and hence we deemed this an exploratory study. However, PostHoc analysis of the MWT demonstrated 100% statistical power, therefore demonstrating adequate statistical power with this sample size.It is also important to note that we selected our athletes for their sporting discipline, however, these findings cannot be generalizable to athletes of other sports, sex or age. Hence further work to address the multi-factorial nature of the AH alongside ethnic specific adaptation is required.It is important to note that longitudinal \u03b5 is the most commonly used indicator of LV mechanics in practice, with radial and circumferential \u03b5 deemed to carry limitations in terms of reproducibility in both physiological and pathological subjects.This study demonstrated that athletes of NH&PI ethnicity have greater LV and RV wall thickness and mass compared to their Caucasian counterparts. In addition, there is reduced LV circumferential and twist mechanics that may reflect the differences in LV geometry structural adaptation."} +{"text": "Pathophysiological concepts in delirium are not sufficient to define objective biomarkers suited to improve clinical approaches. Advances in neuroimaging have revalued electroencephalography (EEG) as a tool to assess oscillatory network activity in neuropsychiatric disease. Yet, research in the field is limited to small populations and largely confined to postoperative delirium, which impedes generalizability of findings and planning of prospective studies in other populations. This study aimed to assess effect sizes of connectivity measures in a large mixed population to demonstrate that there are measurable EEG differences between delirium and control patients.n\u2009=\u2009129 and 414, respectively). Assessors were not blinded for groups. Power spectra and connectivity estimates, using the weighted phase log index, of continuous EEG data were compared between conditions. Alterations of information flow through nodes of intrinsic connectivity networks were evaluated in source space using betweenness centrality. This was done frequency specific and network nodes were defined by the multimodal human cerebral cortex parcellation based on human connectome project data.This retrospective pilot study investigated EEG measures as biomarkers in delirium using a case-control design including patients diagnosed with delirium (DSM-5 criteria) and age-/gender-matched controls drawn from a database of 9980 patients \u2009=\u200970.3, p\u2009<\u2009.001; F\u2009=\u20092.69, p\u2009<\u2009.001; and F\u2009=\u20091.14, p\u2009=\u2009.007, respectively). Connectivity analyses revealed global alpha and regional beta band disconnectivity that was accompanied by theta band hyperconnectivity in delirious patients. Source and network analyses yielded that these changes are not specific to single intrinsic connectivity networks but affect multiple nodes of networks engaged in level of consciousness, attention, working memory, executive control, and salience detection. Effect sizes were medium to strong in this mixed population of delirious patients.Delirium and control patients exhibited distinct EEG power, connectivity, and network characteristics contains supplementary material, which is available to authorized users. Delirium describes an acute confusional state that affects about 10\u201370% of hospitalized patients with the risk increasing with patient age and disease severity . AlthougCurrent research indicates that electroencephalography (EEG) is a promising tool and suitable means to elaborate on the neurophysiological basis of delirium , 14. SevAbove-mentioned limitations render further investigations of EEG measures as biomarkers of cognitive dysfunction in delirium challenging. Importantly, effect sizes to be expected in mixed or other than cardiothoracic populations cannot be deduced. It is the objective of this study to estimate effect sizes of EEG connectivity measures in a mixed delirious population representative of patients treated in a large tertiary care hospital. This is a critical and mandatory step to estimate sample sizes and resources required for a planned prospective study on the utility of advanced EEG measures as therapeutic and prognostic biomarkers of delirium. For this purpose, we evaluated resting state EEG data obtained during the routine clinical work-up of delirious patients that were confirmed by validated clinical tools. Results were compared to age- and gender-matched controls. The retrospective study design allowed us to include a large sample size that accounts for potentially high variance inherent to the heterogeneity of delirium subtypes. Connectivity analyses included the exploratory investigation of oscillatory activity changes in established ICNs, which are discussed regarding their implications for the pathophysiology of delirium.Helsinki declaration. Data protection and ethics review committee approval were obtained from the Institutional Review Board of the Charit\u00e9 Universit\u00e4tsmedizin Berlin in line with regulations for retrospective studies. This study furthermore conforms with the STROBE Statement reporting standard for case-control studies ; P4\u2013F7, t(541)\u2009=\u20094.71, p\u2009=\u2009.04, Cohen\u2019s d\u2009=\u2009.4 [.37\u2013.43]; C4\u2013Cz, t(541)\u2009=\u20094.36, p\u2009=\u2009.05, Cohen\u2019s d\u2009=\u2009.37 [.33\u2013.41]). Decreased connectivity in the beta band was most prominent in parieto-occipital regions \u2009=\u2009\u2212\u20096.22, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009\u2212\u2009.53 [\u2212\u2009.57\u2013\u2212\u2009.49]; P3\u2013O1, t(541)\u2009=\u2009\u2212\u20094.65, p\u2009=\u2009.03, Cohen\u2019s d\u2009=\u2009\u2212\u2009.4 [\u2212\u2009.45\u2013\u2212\u2009.35]; P3\u2013C3, t(541)\u2009=\u2009\u2212\u20094.57, p\u2009=\u2009.03, Cohen\u2019s d\u2009=\u2009\u2212\u2009.39 [\u2212\u2009.44\u2013\u2212\u2009.34]). Strongest reductions in alpha connectivity were found between Fp2\u2013C4 and Fp2\u2013Cz (t(541)\u2009=\u2009\u2212\u200911.57, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009\u2212\u20091 [\u2212\u2009.1.06\u2013\u2212\u2009.94] and t(541)\u2009=\u2009\u2212\u200910.24, p\u2009<\u2009.001,, Cohen\u2019s d\u2009=\u2009\u2212\u2009.88 [\u2212\u2009.94\u2013\u2212\u2009.82], respectively).ANOVA also revealed a significant interaction of connectivity estimates for GROUP \u00d7 SENSOR \u00d7 FREQUENCY \u2009=\u20091.14, p\u2009=\u2009.007). This indicates that information flow between nodes of examined networks differs between delirious and control patients, and that this effect is further specified by oscillatory frequencies. A summary of findings is given in Table\u00a0t(541)\u2009=\u20094.26, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009.37 [.31\u2013.41]) while alpha \u2009=\u2009\u2212\u20093.73, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009\u2212\u2009.32 [\u2212\u2009.36\u2013\u2212\u200926]) and beta \u2009=\u2009\u2212\u20092.36, p\u2009=\u2009.01, Cohen\u2019s d\u2009=\u2009\u2212\u2009.2 [\u2212\u2009.22\u2013\u2212\u2009.18]) band centrality were decreased. Similar changes were found in the PCC where delta band centrality was increased \u2009=\u20092.99, p\u2009=\u2009.001, Cohen\u2019s d\u2009=\u2009.26 [.23\u2013.28]) in line with changes in the theta band \u2009=\u20095.47, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009.47 [.36\u2013.55]) while faster frequencies revealed only decreased centrality in the alpha band \u2009=\u2009\u2212\u20093.95, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009\u2212\u2009.34 [\u2212\u2009.39\u2013\u2212\u2009.27]).There was a significant interaction of GROUP \u00d7 REGION \u00d7 FREQUENCY for the ANOVA of betweenness centrality estimates \u2009=\u20095.52, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u200947 [.38\u2013.54]) while enhanced delta band information flow was found in parcel MIP of the superior parietal cortex (t(541)\u2009=\u20091.86, p\u2009=\u2009.03, Cohen\u2019s d\u2009=\u2009.16 [.15\u2013.17]). The DLPFC, in line with changes in the DMN, showed increased centrality in the theta band \u2009=\u20094.23, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009.37 [.31\u2013.41]) while faster frequencies in the alpha \u2009=\u2009\u2212\u20093.73, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009\u2212.32 [\u2212\u2009.36\u2013\u2212\u2009.26]) and beta \u2009=\u2009\u2212\u20092.36, p\u2009=\u2009.01, Cohen\u2019s d\u2009=\u2009\u2212\u2009.2 [\u2212\u2009.22\u2013\u2212\u2009.18]) band revealed decreased centrality.Centrality parameters of the executive control network (ECN) showed alterations throughout frequency bands that are summarized in Fig.\u00a0t(541)\u2009=\u20091.66, p\u2009=\u2009.05, Cohen\u2019s d\u2009=\u2009.14 [.14\u2013.15]) while centrality in the theta band was enhanced in two parcels with maximum changes in parcel 24 (t(541)\u2009=\u20093.7, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009.32 [.28\u2013.35]). Changes in the DLPFC were in line with changes in the DMN and ECN confined to the theta \u2009=\u20094.26, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009.37 [.31\u2013.41]), alpha \u2009=\u2009\u2212\u20093.44, p\u2009<\u2009.001, Cohen\u2019s d\u2009=\u2009\u2212\u2009.3 [\u2212\u2009.33\u2013\u2212\u2009.25]), and beta band \u2009=\u2009\u2212\u20092.36, p\u2009=\u2009.01, Cohen\u2019s d\u2009=\u2009\u2212\u2009.2 [\u2212\u2009.22\u2013\u2212\u2009.18]) while delta band centrality was unchanged between conditions. A summary of findings is given in Fig.\u00a0Only two regions of the salience network revealed changes in centrality. The anterior cingulate cortex/medial prefrontal cortex (ACC/MPFC) was exclusively altered regarding slow frequencies. Delta band centrality was only increased in parcel p32pr , posterior cingulate, and parietal and mesial temporal cortices . A fine-The salience network is regarded a system that integrates multisensory information with visceral and autonomic states to identify homeostatically relevant inputs and actions , 81. In The executive control network constitutes, next to the salience network, the second ICN within the task-activation network . It operWe cannot rule out a selection bias that is immanent to retrospective sample collection procedures. Although substantial efforts were made to include only EEG data from patients that had a confirmed diagnosis of delirium, it is possible that the fluctuating course of delirium severity may have caused routine EEGs not to be recorded when delirium was most severe. Another limitation is that the discussion of behavioral effects is not based on explicit data obtained from this study\u2019s population but on typical features in delirious patients. We yet made substantial efforts to include only data of patients that were clearly diagnosed with an episode of delirium in line with current diagnostic criteria. Patients included in this study thus presented by definition with impaired attention, perception, and cognitive disturbances including memory, executive, and orientation domains.Delirium is the most common neuropsychiatric condition in hospitals. Yet it is a grossly underdiagnosed condition and there are no objective biomarkers to guide its clinical management , 90. ThiThis pilot study provides comprehensive evidence that EEG biomarkers are promising tools to advance research and enhance care in delirium. Altered homeostasis of oscillatory brain activity is a key finding and functional networks are critically disrupted in delirium, which may be central to clinical features. Source analyses revealed that multiple nodes of intrinsic connectivity networks tasked with cognitive functions such as working memory, salience detection, sustained attention, and executive control are affected during delirium. Effect sizes were generally medium to strong indicating that EEG-based connectivity and network analyses are viable means to elaborate on the pathophysiology of delirium. Results will be used for planning a prospective observational study investigating identified biomarker candidates regarding their therapeutic and prognostic significance.Additional file 1:Search terms used for the retrospective identification of delirious patients. List of key words that were entered to the EEG documentation system in order to screen for delirious patients. (DOCX 12 kb)Additional file 2:Flow chart of the patient selection procedure. Flow chart of the patient selection procedure in line with suggestions made by the STROBE guidelines for reporting of case-control studies. (DOCX 33 kb)Additional file 3Power differences between groups. Complete table of frequency band specific pairwise comparisons for power differences between groups. (PDF 21 kb)"} +{"text": "Conventional analyses using UV absorption spectroscopy or atomic absorption spectrometry (AAS) need to dissolve the silica-based metal chelate sorbent as sample pretreatment. In the first step, those methods were validated on the basis of an ideal homogenous NiSO4-solution and unveiled that UV with an intermediate precision of 2.6% relative standard deviation (RSD) had an advantage over AAS with an intermediate precision of 6.5% RSD. Therefore, UV analysis was chosen as reference method for the newly established NIRS model which has the advantage of being able to measure the material directly in diffuse reflection mode. Partial least squares regression (PLSR) analysis was used as multivariate data analysis tool for quantification. The best PLSR result obtained was: coefficient of determination (R2) = 0.88, factor = 2, root mean square error of prediction (RMSEP) = 22 \u00b5mol/g or 7.5% RSDPLSR. Validation of the Ni2+-capacity using UV absorption spectroscopy resulted in an intermediate precision of \u00b118 \u00b5mol/g or 5.0% RSD. Therefore, NIRS provides a fast alternative analysis method without the need of sample preparation.The present paper reports a new method for the quantification of the Ni Thereforesidues .Typically, the immobilized metal ion content regarding the capacity of the IMAC material is determined by UV absorption spectroscopy or atomic absorption spectrometry (AAS). For that, the analysis is in need of sample preparation to release the metal ions in the first step and to quantify them in the second step ,12.\u22121 (800\u20132500 nm) is used [\u22121), which is also known as short-wave NIR (SWNIR) or Herschel region, bands emerge not only due to overtones and combination modes but also due to electronic transitions. In region II (8500\u20135500 cm\u22121), bands due to first overtones of stretching vibrations and combination modes can be observed. In region III (5500\u20134000 cm\u22121), bands due to combination modes arise [2+-capacity of IMAC materials, which are based on the synthesis postulated by Anspach [Near-infrared spectroscopy (NIRS) is known to be a fast analytical technique which operates noninvasively and usually needs no sample preparation. This vibrational spectroscopic technique is based on the interaction of light with matter, where light in the wavenumber region from 12,500 to 4000 cm is used . The neaes arise . In the es arise . The comes arise . The moses arise . Broadlyes arise . In the Anspach , was est2+-capacity was not reproducible. The minimum difference by using the same silica gels as starting materials was 50 \u00b5mol/g and went up to a maximum difference of 138 \u00b5mol/g of the Ni2+-capacity. Therefore, 32 IMAC materials differing in Ni2+-capacity were synthesized by reapplying 12 silica gels. The usage of different pore and particle size ensured that a variation of these physical parameters which strongly affect the NIR spectra was taken into account. Correlation analysis of the Ni2+-capacity and pore size gave a Pearson correlation coefficient [2+-capacity of all the IMAC materials was in the range from 156 to 444 \u00b5mol/g. Details of all 32 IMAC materials are given in The repeated syntheses of the IMAC materials showed that the resulting Nifficient of \u22120.6 2+-capacity of IMAC materials: UV absorption spectroscopy and AAS [2), limits of detection (LOD) and limits of quantification (LOQ) for the calibrations were 0.9949 \u2264 R2 \u2264 0.9998, 0.00028 \u2264 LOD \u2264 0.0014 mol/L and 0.0010 \u2264 LOQ \u2264 0.0047 mol/L for the UV method and 0.9821 \u2264 R2 \u2264 0.9972, 5.6 \u00d7 10\u22126 \u2264 LOD \u2264 1.4 \u00d7 10\u22125 mol/L and 1.9 \u00d7 10\u22125 \u2264 LOQ \u2264 4.7 \u00d7 10\u22125 mol/L for the AAS method, respectively. UV measurements resulted in a repeatability of 1.9% relative standard deviation (RSD) and intermediate precision of 2.6% RSD. AAS measurements resulted in a repeatability of 6.2% RSD and intermediate precision of 6.5% RSD. The results unveiled that the UV absorption spectroscopy method performed better than the AAS one and therefore was better suited for the determination of the Ni2+-capacity of the IMAC materials. The raw data and regression parameters for all nickel solution measurements and calibrations are given in Two techniques are typically used for the quantification of the Ni and AAS ,12. The 2+-capacity reference analysis for creating the PLSR model was evaluated. Therefore, six IMAC materials were synthesized for the determination of the repeatability and the intermediate precision. The R2, LOD and LOQ of all calibrations were 0.9977 \u2264 R2 \u2264 0.9998, 0.00028 \u2264 LOD \u2264 0.0010 mol/L and 0.00092 \u2264 LOQ \u2264 0.0032 mol/L, respectively. The Ni2+-capacity reference analysis gave a repeatability of 4.2% RSD or \u00b115 \u00b5mol/g and an intermediate precision of 5.0% RSD or \u00b118 \u00b5mol/g. In contrast to the UV method validation of the standard solutions which had an intermediate precision of 2.6% RSD [\u22121 can be assigned to silanol groups hydrogen bonded to water molecules (literature: 7225 cm\u22121) [\u22121 is due to water molecules hydrogen bonded to silanol groups and water molecules hydrogen bonded to other water molecules (literature: 7121 cm\u22121) [\u22121 (literature: ca. 6900 cm\u22121 [\u22121 [\u22121 in region from 6000 to 5600 cm\u22121 can be assigned to overtones of CH stretching vibrations [\u22121) and a band at 5268 cm \u22121 due to the water molecules adsorbed onto hydrogen bonded silanol groups (literature: 5270 cm\u22121) [\u22121, can be assigned to the combination of the asymmetric stretch and bending of the water molecule (literature 5150 cm\u22121) [\u22121 in the raw spectra or seen at 4568 and 4524 cm\u22121 in the second derivative represent the combination band of OH stretch with one of the SiO2 fundamentals ,23,24,2525 cm\u22121) . The ban21 cm\u22121) . The bro900 cm\u22121 and 6861cm\u22121 [\u22121 ) is due cm\u22121 [\u22121 . The banbrations . Analysi70 cm\u22121) ,23. The \u22121 and 6120\u20134008 cm\u22121. \u22121 was kept out since the very broad band that can be found in this region in the raw spectra regarding the Ni2+-capacity represents the intermediate precision of the NIRS method . The UV absorption spectroscopy method used as Ni2+-capacity reference analysis achieved an intermediate precision of \u00b118 \u00b5mol/g (5.0% RSD). Therefore, the NIRS model performed comparable. Raw spectra, reference data and the established PLSR models are provided in a Microsoft Excel File and The Unscrambler X File in the The best PLSR model was obtained by applying the following spectral pre-treatment: second derivative . Wavenumber regions used for the model were: 7500\u20136700 cm spectra gives no spectra . This me2, \u226599%) was purchased from Fluka . Nickel(II) sulfate hexahydrate , sodium hydroxide pellets and sodium chloride were purchased from Merck . Hydrochloric acid was ordered from Carl Roth . Deionized water was purified by Milli-Q\u00ae Reference water purification system and was specified by an electrical resistivity of 18.2 M\u03a9cm.3-Glycidyloxypropyl)trimethoxysilane and iminodiacetic acid were obtained from Sigma-Aldrich . Ethylenediaminetetraacetic acid disodium salt dihydrate ; compressed air.The following 12 silica gels differing in pore and particle size were gained: Carl Roth , 35 \u00c5 (40\u201363 \u00b5m), 60 \u00c5 (20\u201345 \u00b5m), 60 \u00c5 (30\u2013200 \u00b5m), 60 \u00c5 (35\u201370 \u00b5m), 60 \u00c5 (40\u201363 \u00b5m), 60 \u00c5 (200\u2013500 \u00b5m), 150 \u00c5 (35\u201370 \u00b5m), 150 \u00c5 (70\u2013200 \u00b5m), 1000 \u00c5 (35\u201370 \u00b5m), and 250 \u00c5 (40\u201363 \u00b5m); Fluka , 60 \u00c5 (63\u2013200 \u00b5m); and Sigma-Aldrich , 100 \u00c5 (63\u2013200 \u00b5m).2O. Then, 3 g silica gel were added to the solution and stirred for 30 min at room temperature. Next, 2.3 mL 6 M HCl (14 mmol) were used to neutralize the solution. Afterwards, the silica gel was filtered and washed several times with H2O. The particles were dried at 45 \u00b0C under vacuum (300 mbar).First, 0.56 g (14 mmol) NaOH were dissolved into 28 mL H2O was prepared at 0 \u00b0C. Thirty-five grams (144 mmol) of 3-glycidyloxypropyl)trimethoxysilane (GLYMO) were slowly added during 30 min. The mixture was allowed to warm up to room temperature and was stirred for 4 h. Afterwards, the temperature was raised to 65 \u00b0C and the reaction mixture was stirred for 18 h [A solution of 29.4 g NaOH and 19.6 g (144 mmol) iminodiacetic acid (IDA) in 210 mL H2O and 2 g of activated silica gel were added. The suspension was refluxed at 100 \u00b0C for 3 h. The mixture was allowed to cool down to room temperature. The particles were filtered, washed several times with H2O and dried at 45 \u00b0C under vacuum (300 mbar) [Nine milliliters of 6 M HCl were added to 24 mL of the GLYMO-IDA solution to adjust the pH-value to 3.5. The solution was diluted with 50 mL H00 mbar) .2O, 0.5 M NaCl solution and H2O successively. The obtained turquoise IMAC material was dried at 45 \u00b0C under vacuum (300 mbar). The structure [Forty milliliters of 0.1 M nickelsulfate solution were added to 1 g of the silica-GLYMO-IDA material. The suspension was stirred for 2 h at room temperature. To remove unbound nickel ions, the particles were filtered and washed with Htructure is shownFirst, 1.5 mL 0.05 M EDTA solution were added to 100 mg of the IMAC material and mixed for 10 min at 1600 rpm using a Thermomixer C . After centrifugation for 5 min at 13,200 rpm or 15,000 rcf using a Centrifuge 5418 , the supernatant was removed and the procedure was repeated. The supernatants were combined and filled up to 5 mL using a volumetric flask. The solution was transferred into a cuvette and the absorption was measured in triplicate at \u03bb = 380 nm using a Jenway Genova Plus Life Science Spectrophotometer . External calibration was done by dissolving nickel sulfate hexahydrate in 0.05 M EDTA solution.A Perkin Elmer AAnalyst 800 equipped with the burner system and a Nickel (Ni) Lumina Hollow Cathode Lamp Series N3050152 was used for AAS. The system was controlled with the software WinLab32 for AA . Parameters of the used method: Flame-Oxidant, Air; Oxidant Flow, 17.0 L/min; Acetylene Flow, 2.0 L/min; Spectrometer-Element, Ni; Lamp Current, 30 mA; Wavelength, 232.0 nm; Slit width, 0.2 H nm; Signal-Type, AA; Measurement, time average; Read Parameters-Time, 3.0 s; Delay Time, 0 s; Replicates, 3.The solutions used for UV absorption spectroscopy were diluted with 0.05 M EDTA to a nickel concentration of approximately 0.14 mM or 8 \u00b5g/mL using a 50 mL volumetric flask to be in the range of the calibration (2\u201315 \u00b5g/mL). Measurements were done in triplicate. External calibration was done by dissolving nickel sulfate hexahydrate in 0.05 M EDTA solution.\u22121 accumulating 64 scans with a spectral resolution of 8 cm\u22121 and a digital resolution of 4 cm\u22121. Sample measurements were done in triplicate and averaged to one representative spectrum for each sample.Measurements of the NIR spectra of all 32 samples were done using the NIRFlex N-500 FT-NIR spectrometer equipped with the Solids Cell N500-001 and the Vial Add-on N510-002, which sequentially scans 6 samples. The operating software was NIR Ware 1.4.3010 . Spectra were recorded in diffuse reflection in the wavenumber region from 10,000 to 4000 cmThe precision of the UV absorption spectroscopy and AAS methods were evaluated by determination of the repeatability and intermediate precision referred to ICH guidelines ,28. Ther2+-capacity determination was done by using UV absorption spectroscopy which showed a better performance than AAS , regression coefficients, the number of factors and the relative standard deviation (RSD) calculated as follows: % RSDPLSR = \u00d7 100%.MVA was carried out by using the software The Unscrambler X Version: 10.5 . Since the measured spectra were recorded in reflectance, all spectra had to be transformed to absorbance units to ensure linear relation between spectral and reference data. Furthermore, spectra were pretreated by multiplicative scatter correction (MSC) or stand2+-capacity of the synthetized IMAC materials. This method was selected as reference analysis for the establishment of a PLSR model using NIRS in hyphenation with MVA. The NIRS model performed almost as well as the UV reference analysis and better than the AAS method. Therefore, the analysis using NIRS represents an attractive alternative for the determination of the Ni2+-capacity of IMAC materials as it operates noninvasively, fast and needs no sample preparation.UV absorption spectroscopy was unveiled to be superior to AAS for the quantification of the Ni"} +{"text": "Rhodoccocus and Pseudonocardia as the most abundant in terms of the absolute number of sequences. Streptomycetes presented the highest diversity , with most of the OTUs unlinked to the culturable Streptomyces strains that were previously isolated from the same deposits. Furthermore, 43% of the OTUs were shared between the three studied collection points, while 34% were exclusive to one deposit, indicating that distinct speleothems host their own population, despite their nearby localization. This important spatial diversity suggests that prospecting within different moonmilk deposits should result in the isolation of unique and novel Actinobacteria. These speleothems also host a wide range of non-streptomycetes antibiotic-producing genera, and should therefore be subjected to methodologies for isolating rare Actinobacteria.Moonmilk are cave carbonate deposits that host a rich microbiome, including antibiotic-producing Actinobacteria, making these speleothems appealing for bioprospecting. Here, we investigated the taxonomic profile of the actinobacterial community of three moonmilk deposits of the cave \u201cGrotte des Collemboles\u201d via high-throughput sequencing of 16S rRNA amplicons. Actinobacteria was the most common phylum after Proteobacteria, ranging from 9% to 23% of the total bacterial population. Next to actinobacterial operational taxonomic units (OTUs) attributed to uncultured organisms at the genus level (~44%), we identified 47 actinobacterial genera with Molecular approaches evaluating microbial communities in caves have revealed a level of diversity greater than initially expected . Microor3 deposition [3 precipitation, or even directly precipitating carbonate minerals [Among cave speleothems, moonmilk draws a particular scientific attention due to its distinctive crystal morphology. The origins of various moonmilk crystalline habits, including monocrystalline rods, polycrystalline chains, and nanofibers, are tentatively attributed to the moonmilk indigenous microbial population . Among aposition ,20,21. Aminerals ,21. Membminerals ,14,18,19minerals ,23, volcminerals ,25,26, aminerals . The brominerals . Streptomyces. Such a dominance of streptomycetes was rather unexpected, according to other moonmilk microbial diversity studies performed through culture-dependent [Rhodococcus, Pseudonocardia, Propionibacterium, Nocardia, Amycolatopsis, Saccharothrix, Geodermatophilus, Mycobacterium, Aeromicrobium, Kribella, Nocardioides, Actinomycetospora, Nonomuraea, Euzebya, Rubrobacter, and Arthrobacter, in addition to Streptomyces. Nonetheless, the diversity of the moonmilk actinobacterial microbiome still remains largely unknown and, beyond evaluating \u201cwhat and how much have we missed in our culture-dependent bioprospecting approach\u201d [to what extent are moonmilk-dwelling Actinobacteria different between the moonmilk deposits within a single cave, or in different caves?As recently reported, moonmilk Actinobacteria represent novel microorganisms, which is a discovery that opens great avenues for the bioprospecting of novel drugs ,10,18. Rependent ,13,18 anependent , denaturependent ,16,17, aependent . The actIn this work, we carried out a comparative (HTS) of 16S small subunit (SSU) rRNA gene from DNA extracted from spatially separated moonmilk deposits within the same cave, \u201cGrotte des Collemboles\u201d (Springtails\u2019 Cave) in Comblain-au-Pont, Belgium , in ordeLibraries spanning the V4\u2013V6 variable regions of the 16S rRNA gene using universal bacterial primers were used to assess the proportion of Actinobacteria in comparison to the whole bacterial community of three moonmilk deposits of the cave \u201cGrotte des Collemboles\u201d . The obsBacterial OTUs were grouped into 21 phyla and 18 candidate phyla . ActinobEvaluation of the actinobacterial profile was performed with libraries spanning the V6\u2013V7 variable regions of 16S rRNA gene, and using modified Actinobacteria-specific primers . The speThe diversity indices for Actinobacteria showed the same trends as the ones observed for the whole Bacteria domain, i.e., evenness and diversity were the highest in COL4, followed by COL3, and COL1 . PhylotyA taxonomic assignment of actinobacterial OTUs revealed the presence of two major classes\u2014Acidimicrobiia and Actinobacteria, next to the low-abundant Thermoleophilia class a. AcidimRhodococcus (38.37%), while uncultured and unclassified Actinobacteria were the most abundant in COL3 and COL4 were exclusively found in a single sampling site accounting together for 90% of the sequences obtained via our HTS approach . Out of Nocardia .Rhodococcus, and accounting for 38% of the total population in this speleothem was also prevailing in COL3, while Pseudonocardia was found to be the most abundant in COL4 , which largely dominated the actinobacterial community in COL3 (29%), while only representing 0.3% of the actinobacterial microbiome in COL1 . Streptomyces moonmilk-dwelling community we managed to isolate in our previous bioprospection work [Streptomyces OTUs retrieved from the HTS approach with the sequences of the 31 previously isolated Streptomyces phylotypes (MM strains), which were trimmed to the corresponding V6\u2013V7 variable regions of HTS amplicons. Streptomyces phylotypes (MM strains) and OTUs. The identity threshold for clustering sequences in the same branch of the tree was fixed to 97%, i.e., the same threshold as the one used to define OTUs in our HTS approach (see methods for details). As deduced from the generated phylogenetic tree, the 31 isolated Streptomyces strains matched with only five of the 19 Streptomyces OTUs, suggesting that the isolated strains represent a minor fraction of the Streptomyces species dwelling in the moonmilk deposits of the studied cave. Expectedly, Streptomyces species that are associated with the most abundant Streptomyces OTUs, e.g., OTU15, OTU21, OTU30, and OTU99 clustered together with OTU21 [Pseudonocardia, Amycolatopsis and Saccharothrix (not identified in our study) (Pseudonocardiaceae) [Propionibacterium (Propionibacteriaceae) [Streptomyces (Streptomycetaceae) [Arthrobacter (Micrococcaceae) [Mycobacterium (Mycobacteriaceae) [Nocardioides, Aeromicrobium, and Kribbella (Nocardioidaceae) [Geodermatophilus (Geodermatophilaceae) [This new study uncovered a surprisingly diverse Actinobacteria taxonomic profile that demonstrates the limitations of our previous cultivation-based screening, in which only the deposits . Here, adiaceae) ,18, Pseudiaceae) ,14,19, Petaceae) ,12,18,19ccaceae) , MycobacStreptomyces are reported as the most prolific \u201cantibiotic makers\u201d, advances in the cultivation and characterization of rare Actinobacteria revealed similarly promising capabilities for the production of bioactive natural compounds [Streptomyces, other members of Actinobacteria with valuable secondary metabolism were detected at a high proportion, such as Pseudonocardia, Amycolatopsis, Streptosporangium, Nocardia, Nocardioides, and Rhodococcus. Such findings clearly prompt to apply appropriate selective cultivation methods to isolate rare Actinobacteria from moonmilk deposits.Extreme environmental niches have recently become the main targets for intense bioprospecting, as they are expected to host diverse yet-unknown microorganisms, which could offer unexplored chemical diversity. While ompounds ,35,36. TMoreover, particular importance should be also focused on Acidimicrobiia, which constituted an important part of the community in the studied deposits. Members of this class are a recently identified taxonomic unit that is Streptomyces isolated in our previous study and the Streptomyces OTUs identified in this work , located in Comblain-au-Pont , Belgium . Moonmilw/v), and the dsDNA concentration was evaluated by Qubit fluorometer . The metagenetic approach applied in this work was performed on DNA extracted from three moonmilk deposits originating from the \u201cGrotte des Collemboles\u201d. Environmental genomic DNA isolation was carried out from 200 mg of the freeze-dried moonmilk samples COL1, COL3, and COL4 , using tThe 16S rRNA gene amplicon libraries were generated using bacterial (S-D-Bact-0517-a-S-17/S-D-Bact-1061-a-A-17 spanning V4\u2013V6 region ) and act16S rRNA amplicon analysis was based for both Bacteria and Actinobacteria on forward reads only, owing to the poor quality of reverse reads. Quality trimming was carried out using CLC Genomic Workbench . USEARCH was applNormalized OTU abundance data was used to calculate \u03b1-diversity and \u03b2-diversity estimators using MOTHUR . CommuniStreptomyces were combined to 31 sequences (16S rRNA region V6\u2013V7) from previously isolated Streptomyces phylotypes (MM strains) and dereplicated with the UCLUST algorithm [Corynebacterium diphtheriae JCM-1310 as an outgroup. A multiple sequence alignment was built with MUSCLE [4 model. Due to the limited amount of phylogenetic signal (short sequences from very related organisms), the resolution of the tree was low .The 19 OTUs identified as lgorithm using anh MUSCLE (defaulth MUSCLE under a Streptomyces strains of our collection isolated from the moonmilk deposits of the cave \u2018Grotte des Collemboles\u2019 [tips and tricks\u2019 to isolate other Streptomyces and representatives of other antibiotic-producing Actinobacteria that are present in different proportions in each moonmilk deposit. The results of our adapted protocols for the isolation of rare Actinobacteria are presented in the article \u2018Isolation, Characterization, and Antibacterial Activity of Hard-to-Culture Actinobacteria from Cave Moonmilk Deposits\u2019, which is published in the same special issue [Before the advent of metagenomics, bioprospecting was carried out blindly, with poor knowledge on the real potential of an ecological niche mined for novel organisms, enzymes, or bioactive compounds. The results of the metagenetic study presented here confirmed that different moonmilk deposits host their own indigenous microbial population, and thus each individual speleothem can be a source of a great biodiversity. Consequently, the observed important differences in the spatial diversity of Actinobacteria imply that bioprospecting within different moonmilk deposits\u2014from different caves or within the same cave\u2014could result in the isolation of unique and novel natural compound producers. Our study also revealed how many and which actinobacterial genera have been missed in our first attempt to isolate antibiotic producers. We now know that the emboles\u2019 are justal issue ."} +{"text": "ADP-ribosylation is a ubiquitous post-translational addition of either monomers or polymers of ADP-ribose to target proteins by ADP-ribosyltransferases, usually by interferon-inducible diphtheria toxin-like enzymes known as PARPs. While several PARPs have known antiviral activities, these activities are mostly independent of ADP-ribosylation. Consequently, less is known about the antiviral effects of ADP-ribosylation. Several viral families, including Coronaviridae, Togaviridae, and Hepeviridae, encode for macrodomain proteins that bind to and hydrolyze ADP-ribose from proteins and are critical for optimal replication and virulence. These results suggest that macrodomains counter cellular ADP-ribosylation, but whether PARPs or, alternatively, other ADP-ribosyltransferases cause this modification is not clear. Here we show that pan-PARP inhibition enhanced replication and inhibited interferon production in primary macrophages infected with macrodomain-mutant but not wild-type coronavirus. Specifically, knockdown of two abundantly expressed PARPs, PARP12 and PARP14, led to increased replication of mutant but did not significantly affect wild-type virus. PARP14 was also important for the induction of interferon in mouse and human cells, indicating a critical role for this PARP in the regulation of innate immunity. In summary, these data demonstrate that the macrodomain is required to prevent PARP-mediated inhibition of coronavirus replication and enhancement of interferon production. in vivo. Here, using primary macrophages and mice infected with a pathogenic murine coronavirus, we identify PARPs, specifically PARP12 and PARP14, as host cell ADP-ribosylating enzymes important for the attenuation of these mutant viruses and confirm their importance using inhibitors and siRNAs. These data demonstrate a broad strategy of virus-host interactions and indicate that the macrodomain may be a useful target for antiviral therapy.ADP-ribosylation, an understudied post-translational modification, facilitates the host response to virus infection. Several viruses, including all members of the coronavirus family, encode a macrodomain to reverse ADP-ribosylation and combat this immune response. As such, viruses with mutations in the macrodomain are highly attenuated and cause minimal disease This process is catalyzed by intracellular poly(ADP-ribose) polymerases (PARPs) also known as diphtheria toxin-like ADP-ribosyltransferases (ARTDs), although extracellular cholera toxin-like ADP-ribosyltransferases (ARTCs) and some sirtuins also catalyze ADP-ribosylation . Wh. Wh-/- a/- cells , it sign/- cells . In gene-/- in contrast to PARP14+/- cells showed no increase in IFN expression following infection with N1347A virus replication. We engineered six PARP14 knockout (KO) clones of A549 cells and a pool of PARP14 KO cells in normal human dermal fibroblasts (NHDFs) using lentiCRISPR/CAS9-v2 Figs & S7A. Tin vivo [Here we show that PARPs, specifically PARP12 and PARP14, are required to inhibit the replication of a macrodomain-mutant CoV and that PARP14 is also required for optimal IFN expression. Previous reports have shown that CoVs were unable to cause disease in the absence of viral macrodomain ADP-ribosylhydrolase activity and that this attenuation was associated with reduced viral loads and changes in pro-inflammatory cytokine expression in vivo , 47, 50.in vivo .PARPs have both antiviral and immunomodulatory roles . Here weStudies with chikungunya virus and Sindbis virus demonstrate that the alphavirus macrodomain also counters cellular ADP-ribosylation as mutants with decreased ADP-ribose binding and enzymatic activity had mild to severe replication defects , 45, 73.-/- BMDMs as directed by the Guide for the Care and Use of Laboratory Animals (Protocol #6071795). Anesthesia or euthanasia were accomplished using isoflurane and ketamine/xylazine or ketamine/xylazine, respectively.-/-, IFNAR-/-, PARP14+/-and PARP14-/- C57BL/6 mice were differentiated by incubating cells with 10% L929 cell supernatants and 10% FBS in Roswell Park Memorial Institute (RPMI) media for seven days. Cells were washed and replaced with fresh media every day after the 4th day.Delayed brain tumor (DBT) (propagated in Perlman laboratory since 1983), normal human dermal fibroblasts (NHDF) (ATCC), A549 (ATCC), Vero , BHK-21 and HeLa cells expressing the MHV receptor carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) (HeLa-MHVR) were grown in Dulbecco\u2019s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 100 U/ml penicillin and 100 \u03bcg/ml streptomycin. Bone marrow-derived macrophages (BMDMs) sourced from WT, MAVSpcDNA3-GFP was previously described , pcDNA3-Following differentiation, BMDMs were treated with the indicated compounds for 24 hours. Cell viability was assessed using a Vybrant MTT Cell Proliferation Assay (Thermo Fisher Scientific) following manufacturer\u2019s instructions.-/- mice were purchased from Jackson Laboratories, and MAVS-/- mice were obtained as a generous gift from Dr. Michael Gale . These mice were bred and maintained in the animal care facility at the University of Iowa. PARP14-/- mice were a generous gift from Dr. Mark Boothby and were bred and maintained in the animal care facility at Harvard Medical School.Pathogen-free C57BL/6 WT and IFNARIA-GFPrevN1347) and N1347A (rJIA-GFP-N1347A) MHV were previously described [7 17Cl-1 cells at an MOI of 0.1 plaque-forming units (PFU)/cell and collecting both the cells and supernatant at 20 hpi. The cells were freeze-thawed, and debris was removed prior to collecting virus stocks. Virus stocks were quantified by plaque assay on Hela-MHVR cells. BMDM cells were infected with MHV at an MOI of 0.1 PFU/cell with a 45\u201360 min adsorption phase. Infected cells were then incubated and collected at the indicated timepoints. Recombinant eGFP-expressing mutant VSV (rM51R-M-EGFP) was obtained from Dr. Douglas Lyles, Wake Forest School of Medicine, Winston-Salem, NC [4 PFU of virus in 12 \u03bcL DMEM. Mice were either monitored for weight loss or were sacrificed at 4\u20136 days post infection (dpi) to harvest the brain tissue. Brain tissues were homogenized, and leukocytes were isolated as previously described [Recombinant WT as per manufacturer\u2019s instructions. For surface staining, brain leukocytes were treated with Fc block and then incubated with specific mAbs or isotype controls. Monoclonal antibodies used for these studies included CD45-PECy7/FITC and CD11b-e450 . Cells were analyzed using a FACS Verse flow cytometer (BD Biosciences). All flow cytometry data were analyzed using FlowJo software .5 DBT cells were transfected with 0.5 \u03bcg total plasmid expressing GFP, PARP12, or WT or CM PARP14 using PolyJet In Vitro Transfection Reagent (SignaGen Laboratories) as per manufacturer\u2019s protocol. Media was replaced 8 hours after transfection, and cells were incubated for 16 hours before collection.5\u00d710For siRNA knockdown, DsiRNA oligonucleotides were purchased from Integrated DNA Technologies (IDT). Sequences are listed in PARP14 KO A549 and NHDF cells and IFNAR KO A549 cells were generated using the lentiCRISPR/CAS9-v2 system. For bot5 cells/well). 16 hours later, cells were mock transfected (wild-type cells) or transfected with 2 \u03bcl Lipofectamine 2000 reagent (Invitrogen) and 500 ng poly(I:C) in 200 \u03bcl of DMEM in duplicate as per manufacturer\u2019s protocol (for recipient WT and IFNAR KO A549 cells). To reduce the effect of the remaining poly(I:C) in the conditioned media, cell media were replaced with fresh media 2 h post transfection. At 6 h post transfection, cell media was collected and centrifuged to remove cell debris, and supernatants were collected as conditioned media. Recipient WT or IFNAR KO A549 cells were then incubated with one of the duplicate samples of conditioned media for 2 h prior to infection with eGFP-VSV (rM51R-M-EGFP) at an MOI of 1 PFU/cell. VSV titers in the supernatants were determined by counting plaques on Vero cells.WT or PARP14 KO A549 or NHDF cells were seeded in 12-well plates for 4 h prior to infection with eGFP-VSV (rM51R-M-EGFP) at an MOI of 1 PFU/cell. At 16 hpi, infected cells were monitored under an AMG-EVOS FL Digital Inverted Fluorescence Microscope (software version 15913). GFP fluorescence microscopy was performed with a light cube of GFP (Ex 470 nm/Em 525 nm) under a 4X objective lens , and images of six different fields were taken for each condition. The quantification of GFP-VSV-infected cells was performed using ImageJ software (NIH) to count GFP (green) points. The means \u00b1 SEM of percentages of infected cells from six different fields (three each from two different replicate wells of a 12-well-plate) are shown.CT =CT (gene of interest)\u2014CT (HPRT). All results are shown as a ratio to HPRT calculated as -2CT\u0394.RNA was isolated from cells using Trizol (Thermo Fisher Scientific) via phase separation or Direct-Zol column purification (Zymo Research) as per manufacturer\u2019s instructions. cDNA was prepared using MMLV-reverse transcriptase as per manufacturer\u2019s instructions (Thermo Fisher Scientific). Quantitative PCR (qPCR) was performed on a QuantStudio3 real-time PCR system using PowerUp SYBR Green Master Mix (Thermo Fisher Scientific). qPCR primers are listed in Total cells were lysed in sample buffer containing SDS, \u03b2-mercaptoethanol, protease/phosphatase inhibitor cocktails (Roche), PMSF, and universal nuclease (Thermo Fisher Scientific). Proteins were resolved on an SDS polyacrylamide gel, transferred to a polyvinylidene difluoride (PVDF) membrane, hybridized with a primary antibody, reacted with an infrared (IR) dye-conjugated secondary antibody, visualized using a LI-COR Odyssey Imager, and analyzed using Image Studio software (LI-COR). Primary antibodies used for immunoblotting include anti-FLAG monoclonal antibody ; anti-PARP14 polyclonal antibodies ; anti-PAR monoclonal antibody ; rabbit anti-MHV polyclonal antibody ; and antSupernatants from infected cells were collected at 12 hpi, and protein levels of IFN\u03b1 and IFN\u03b2 were determined using the Luminex Protein Assay (Thermo Fisher Scientific) according to the manufacturer\u2019s instructions.BMDMs plated on glass cover slips were infected with GFP-expressing MHV. At 14 hpi, cells were fixed with 4% paraformaldehyde, and coverslips were transferred to a glass slide. Vectashield Antifade Mounting Media with DAPI (Vector Laboratories) was applied, and a second coverslip was overlaid. Slides were visualized on an Olympus IX-81 inverted fluorescence microscope (Olympus), and images were analyzed using SlideBook software (Meyers Instruments).An unpaired two-tailed Student\u2019s t-test was used to assess differences in mean values between groups, and graphs are expressed as mean \u00b1 SEM. MHV titers are presented as geometric mean \u00b1 SEM. The n value represents the number of biologic replicates for each figure. The n for WT and N1347A virus-infected samples were the same unless otherwise indicated. Significant p values are denoted with *p\u22640.05, ** p\u22640.01, *** p\u22640.001.S1 Fig(A) Mice were infected as described in Methods with WT and N1347A MHV. Brain tissues were collected at 4 dpi, and CD11b+ cells were purified as described in Methods. Flow cytometry was used to assess purification efficacy. The data in (A) are from one representative experiment of three independent experiments; n = 4. CD11b+ cells were purified, and RNA was isolated and analyzed for viral genomic RNA (gRNA) at indicated time points (B) or for PARP mRNA at 4 dpi (C). The data in show one representative experiment of two independent experiments; n = 4 for WT and N1347A except N1347A at day 5 where n = 3. For na\u00efve samples in (C), n = 2. Numbers above bars represent fold difference between WT and N1347A.(TIF)Click here for additional data file.S2 Fig-/-, or IFNAR-/- C57BL/6 mice were infected as described in Methods and monitored for weight loss and survival over a 12-day period. WT, n = 5; MAVS-/-, n = 9 for WT and n = 11 for N1347A; IFNAR-/-, n = 3 for WT and n = 7 for N1347A.WT, MAVS(TIF)Click here for additional data file.S3 Fig BMDMs were incubated with PARP inhibitors 3-AB (5 mM), XAV-939 (10 \u03bcM), or vehicle (0.25% DMSO) (A) or with PARP14-specific inhibitor compound 8K (5 \u03bcM) or vehicle (B). At 24 hours, cell viability was measured using an MTT assay as described in Methods. The data in show one experiment representative of two independent experiments; n = 4. (C) DBT cells were treated with or vehicle (0.25% DMSO), 3-AB (5 mM), XAV-939 (10 \u03bcM), or 8K (5 \u03bcM). After 18 h, cell lysates were collected and immunoblotted for poly(ADP-ribose) (PAR) or for actin. The data in (C) show one experiment representative of at least two independent experiments.(TIF)Click here for additional data file.S4 FigBMDMs were transfected with control siRNA (siCtl) or PARP-specific siRNA as described in Methods. Approximately 28 hours later, cells were infected with WT or N1347A MHV and collected at 18\u201322 hpi. RNA levels were determined by RT-qPCR with primers specific for each transcript and normalized to HPRT. The level of PARP mRNA in siRNA-treated cells was then normalized to expression in control siRNA-treated cells. The data show the combined results of two to five experiments; n = 9 for siPARP7, n = 6 for siPARP9, n = 6 for siPARP10, n = 12 for siPARP11, n = 15 for siPARP12.2, n = 9 for siPARP12.3, n = 12 for siPARP14.1, n = 12 for siPARP14.2.(TIF)Click here for additional data file.S5 FigBMDMs were infected with WT or N1347A MHV and collected at 12 hpi. Lysates were analyzed by immunoblotting with the indicated antibodies using a LI-COR Odyssey Imager. The data show the results of one experiment representative of two independent experiments.(TIF)Click here for additional data file.S6 Fig(A) DBT cells were transfected with indicated plasmids and collected 24 hours after transfection. Lysates were analyzed by immunoblotting with the indicated antibodies using a LI-COR Odyssey Imager. The data are the results of one experiment representative of two independent experiments. FLAG/PARP12 was utilized as a positive control for the anti-FLAG blot. (B) DBT cells were transfected with plasmid expressing GFP, PARP14, or a PARP14 catalytic mutant (CM). Cells were collected at 24 hours after transfection, and PARP14 mRNA levels were determined by RT-qPCR and normalized to HPRT. The data in (B) show one experiment representative of two independent experiments; n = 3.(TIF)Click here for additional data file.S7 Fig(A) A pool of CRISPR/Cas9-gRNA-mediated PARP14 KO NHDF cells or of gRNA-NC-transduced control NHDF cells were collected and analyzed by immunoblotting for PARP14 protein. A549 cells were treated with conditioned media for 2 h, VSV replication was analyzed by fluorescence microscopy at 16 hpi (B), and titers were determined by plaque assay on Vero cells (C). Titers were normalized to those in mock-treated wild-type cells. p-value markers in (C) represent comparisons of poly(I:C) (pI:C)-treated PARP14-KO cell conditioned media to poly(I:C)-treated WT cell conditioned media used to pretreat WT or IFNAR KO A549 cells. The data in (B) show a single experiment representative of three independent experiments, and the data in (C) show the combined results of three independent experiments; n = 3. (D) WT or IFNAR KO A549 cells were pre-treated for 4 hours with varying amounts of IFN\u03b1 and infected with eGFP-VSV (rM51R-M-EGFP) at an MOI of 1 PFU/cell. Quantification of eGFP was performed directly using fluorescence microscopy and ImageJ software. Shown is virus titer in IFN-treated cells relative to titer in mock-treated cells as determined by percent eGFP-positive cells. The data in (D) show one experiment representative of two independent experiments; n = 6.(TIFF)Click here for additional data file.S1 TableSequences of small interfering RNAs used to knockdown gene expression are listed.(TIFF)Click here for additional data file.S2 TableGuide RNA sequences used to develop lentiCRISPR/CAS9-v2-mediated knockout pools and clones of cells are listed.(TIFF)Click here for additional data file.S3 TablePrimer sequences used to quantify transcription of specific genes are listed.(TIFF)Click here for additional data file."} +{"text": "Extracellular vesicles (EVs), including exosomes, are membranous particles released by cells into the extracellular space. They are involved in cell differentiation, tissue homeostasis, and organ remodelling in virtually all tissues, including the central nervous system (CNS). They are secreted by a range of cell types and via blood reaching other cells whose functioning they can modify because they transport and deliver active molecules, such as proteins of various types and functions, lipids, DNA, and miRNAs. Since they are relatively easy to isolate, exosomes can be characterized, and their composition elucidated and manipulated by bioengineering techniques. Consequently, exosomes appear as promising theranostics elements, applicable to accurately diagnosing pathological conditions, and assessing prognosis and response to treatment in a variety of disorders. Likewise, the characteristics and manageability of exosomes make them potential candidates for delivering selected molecules, e.g., therapeutic drugs, to specific target tissues. All these possible applications are pertinent to research in neurophysiology, as well as to the study of neurological disorders, including CNS tumors, and autoimmune and neurodegenerative diseases. In this brief review, we discuss what is known about the role and potential future applications of exosomes in the nervous system and its diseases, focusing on cell\u2013cell communication in physiology and pathology. Eukaryotic cells in multicellular organisms need to communicate with each other in order to maintain tissue homeostasis and to respond to pathogens in the extracellular milieu. Generally, cells exchange information through direct cell\u2013cell contact or by secretion of soluble factors . MechaniIt was initially thought that exosomes could be a mechanism for shedding the cytoplasm in maturing sheep reticulocytes . Later, Exosomes are released into the extracellular space after the merging of late endosomes with the cell membrane. Previously, early endosomes become part of multivesicular bodies (MVBs), which undergo a maturation process characterized by a gradual change in protein composition of the vesicles . During this maturation process, the vesicles that have accumulated in the MVBs can follow three different pathways: 1) merge with the lysosomes, which leads to the degradation of their protein cargo ; (2) constitute a temporary storage compartment; and (3) blend with the plasma membrane, releasing exosomes. MVBs merge with the plasma membrane, resulting in exocytosis of the vesicles contained in them so that the vesicles\u2019 membrane maintains the same topological orientation as the plasma\u2013cell membrane merge wi,23. The The content of exosomes reflects that of the cell of origin and, consequently, there is interest in characterizing it to obtain information on the cell of origin and the functions of exosomes, and to assess the potential of exosomes as drug delivery tools. The composition of exosomes depends on parental cell conditions, and includes lipids; proteins; and nucleic acids, such as DNA, non-coding RNA, rRNA and miRNAs (microRNAs) .The lipid composition of exosomes is characteristic and includes cholesterol, phosphatidylcholine, sphingolipid ceramide, and sphingomyelin that probably stabilize the exosomal bilayer membrane and maintain its integrity in the extracellular milieu . The sphVarious classes of proteins are found in exosomes, such as proteins involved in the vesicles\u2019 trafficking, cell surface receptors, and proteins involved in endocytic pathways . Also, in exosomes are proteins with specific post-translational modifications (PTMs) ,30, and The presence of mRNA and miRNDepending on the parental cells and their contents, exosomes may have many different functions. They are involved in cell-to-cell information transfer , immune Since exosomes can mediate transfer of molecules, it is very likely that they play a key role in intercellular interactions and in the maintenance of tissue homeostasis ,53. For The nervous system, composed by the central nervous system (CNS), and the peripheral nervous system (PNS), is implicated in the communication with both the external and internal environment of the organism by responding to chemical and physical stimuli .The main cell types found in the nervous tissue are neurons, or nerve cells, that have the ability to rapidly receive and transmit impulses to and from different parts of the body, and neuroglia, or glial cells, which assist in the propagation of the nerve impulses and provide nutrients to the neurons . Both neNeurons are highly specialized cells that receive, process, and transmit information through chemically-mediated electrical signals Figure .Despite the fact that neurons can be specialized and differ in a variety of features, they all share several characteristics. The key function of neurons is to communicate between them and with other cell types. When the nerve impulses travel along the axon in the form of an action potential, the vesicles at the axon terminal, which contain neurotransmitters or neuromodulators, release their content by exocytosis. These signals between neurons are passed via specialized connections called synapses , in whicThe term glia derives from the ancient Greek word \u201cgl\u00eda\u201d meaning \u201cglue\u201d in English, and may suggest a passive type of cell; however, glial cells are active, providing support and nutrition to the neurons, form myelin, and, by insulating axons, speed up electrical communication . A majorFor many years it has been argued that the number of glial cells in the brain was significantly higher than neurons, but recent work has revealed that neurons and glia are almost equal in number in the human cortex ,62,63. HNeuroglia of the CNS can be divided into macroglia and microglia . The macSpinal cord and brain contain different subclasses of oligodendrocytes (OLGs) which derive from multiple sources Figure . OLGs prWithin the oligodendrocyte linage, there exist the NG2-glia/oligodendrocyte cells. They are characterized by the presence on their surface of chondroitin sulfate proteoglycan and are considered an independent glial population, but their function in the adult brain is not yet fully established. NG2-glia maintains the physiological and homeostatic conditions of the nervous tissue generating mature myelinating oligodendrocytes; furthermore, it forms synapses with neurons of the hippocampus and probably in other parts of the brain, too . NotablyThe astrocytes are supportive glial cells in neural tissue with a star-like appearance because of their elaborate cytoplasmic processes ,69,70.Astrocytes play a role in a variety of complex and essential functions in the healthy CNS, such as the maintenance of water and ion homeostasis and blood\u2013brain barrier (BBB) integrity, as well as participation in tripartite synapses, all of which make astrocytes active actors in synaptic context ,71. FurtIn the functional regulation of the microenvironment, astrocytes and oligodendrocytes release EVs, in order to facilitate the cell\u2013cell communication and the activity of target cells ,74,75.The CNS macroglia cells are the ependymoglial cells derived from the neuroepithelium. They populate the interface between the brain parenchyma and the cavity of the ventricles in the CNS, and the central canal of the spinal cord. Macroglial cells appear with various shapes from cuboidal to columnar with cilia and microvilli on the apical surfaces to enhance absorbance and circulation of cerebral spinal fluid (CSF).Ependymoglial cells are of three types: ependymocytes, which make contact with the basal lamina labyrinths (remnants of embryonic blood vessels) and with the ventricles where they contribute to the CSF movement ,77; chorDifferently from the previously described cells of the nervous system, the microglial cells originate from mesodermal hematopoietic cells that in mammals come from the yolk sac ,81. TheyTo define the microglial cells just as macrophages would be an oversimplification because, in addition to their role in defending against bacterial and viral infections, they play a crucial role in the maturation of neural circuits by their \u201csynaptic pruning\u201d function ; they alMicroglial cells have a great morphological plasticity and with their highly motile processes without moving their bodies constantly explore their environment. By screening the brain parenchyma these cells rapidly search for pathogens, signs of injury, or homeostatic disturbances ,86. FinaThe functions of the nervous system and immune system are often considered independent from one another, however, this is a simplistic distinction, because in the regulation of the organism homeostasis, they are in constant communication ,88. ThisIn the PNS there are two types of neuroglia: Schwann cells (SCs) that myelinate axons; and satellite glial cells, that regulate nutrient and neurotransmitter levels in ganglia .SCs are recognized as the PNS counterparts of the oligodendrocytes in CNS, as they are involved in the neuromuscular synapse formation and in wrapping myelin around neuronal axons to form the myelin sheath. This SC activity promotes the efficient and energy low-cost propagation of axon potentials via saltatory conduction by maintaining internodal\u2014each myelin segment is flanked by unmyelinated nodes of Ranvier\u2013myelin sheath thickness and length relative to the diameter of the corresponding axon . SCs canSatellite glial cells (SGCs) have the same origin as Schwann cells. Sensory ganglia of the dorsal roots of the spinal cord are composed of afferent neurons without a myelin sheath but lined by SGCs and connective tissue cells.SGCs share many features with astrocytes, like the expression of glutamine synthetase and various neurotransmitter transporters. They cover axon terminals that make synaptic contacts on, or near, the neuronal somata, wrap around dendrites that emerge from neuronal somata to control the microenvironment and, similarly to astrocytes, influence synaptic transmission ,99. SGCsThe roles of exosomes in the CNS may be as follows: on the one hand, they can be active components necessary for the development and protection of the CNS under physiological conditions ,102,103,Since exosomes are involved both in healthy and pathogenic state of CNS, it is not surprising that these vesicles are released by most of the CNS cells, including neurons, microglia, oligodendrocytes, astrocytes, and neural embryonic stem cells, Cells from malignant gliomas, i.e., primary tumors that arise from neuroglial stem or progenitor cells, produce and release in circulation exosomes with potential to induce malignant transformation of normal cells . It has In amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), FTD-ALS, tauopathies, and Parkinson\u2019s (PD) and Alzheimer\u2019s (AD) diseases, exosomes migrate via blood and CSF carrying misfolded proteins or pro-inflammatory molecules ,110,111.Modified rabies virus glycoprotein (RVG)-targeted EVs were used to transport siRNA across BBB , that caFurthermore, it was demonstrated that the uptake of EVs by neurons in vitro and in vivo neurons and microglial cells of mouse, is more efficient than that of other traditional carriers, i.e., liposomes .The development and maintenance of neuronal circuits in the CNS requires a complex series of events involving coordinated short- and long-distance communication between numerous cell types. Neurons interact continuously with each other and with glial cells through electrical signals and through chemical mediators. In chemical synapses, more common than electrical synapses in the human CNS, the transmission of signals is carried out by chemical mediators, named neurotransmitters. The neurotransmitters are secreted by the pre-synaptic cells inside vesicles that reach the post-synaptic cells, where multiple downstream events, both electric and molecular, are triggered by binding of the neurotransmitter to specific receptors . In viewCNS neurons secrete exosomes to control the complex and coordinated communication among them and, with astrocytes and microglia, thus exosomes mediate a generalized cross-talk, in order to regulate neuronal regeneration and synaptic functions in development and adult life ,117.To the best of our knowledge, the first report regarding exosomes produced by neural cells is relatively recent. Glial cell lines overexpressing a prion protein (PrPsc) released exosomes as a way to spread the PrPsc and, these exosomes bearing PrPsc were infectious, contributing to the spread of prions throughout different areas of CNS and the whole organism . SuccessMicroglia-derived exosomes can modulate neuronal activity also via enhanced sphingolipid metabolism . InflammIn CNS, oligodendrocyte progenitor cells secrete exosome-like vesicles carrying myelin proteolipid protein (PLP), 2\u20193\u2019-cyclic-nucleotide-phosphodiesterase (CNP), myelin basic protein (MBP), and myelin oligodendrocyte glycoprotein (MOG) . The oliThe fact that exosomes can reach the circulation and the CSF makes these vesicles likely means of long-distance communication and transport for bioactive molecules to be delivered to selected targets. Because of their capability to cross the BBB ,130, andOn the other side of the matter, peripheral organs can influence the functions of CNS through exosomes ,133. TheThe intestinal microbiota-derived EVs can also enter the systemic circulation and pass through the BBB, inducing neuroinflammation that could be implicated in the pathogenesis of depressive disorders and affeProgress in the medical sciences has been steady over the last few decades, encompassing the discovery of etiological agents, elucidation of pathogenic mechanisms, and development of new diagnostic techniques and therapeutic strategies. One of the major obstacles to improving patient management has been the heterogeneity of any given disease, which varies from patient to patient. Thus, personalized medicine has emerged to develop means of diagnosis and treatment for the management of each patient in accordance with its specific characteristics. Theranostics is one advance in this direction that also aims at combining diagnostic and therapeutic capabilities in a single agent . ExampleAs described earlier, exosomes derived from different nervous system cells contain specific molecules or cell markers, e.g., oligodendrocyte-derived exosomes contain proteins of the myelin sheath; neuronal exosomes contain cell-adhesion proteins and receptor subunits; microglial-derived exosomes carry peptidases and cytokines. This suggests that exosomes have the ability to regulate and maintain functional cell homeostasis during health and under disease conditions. But on the other hand, exosomes can favor the disease mechanism rather than stop it, when they carry and deliver pathogenic molecules from the cell of their origin. This type of pathogenic role of exosomes has been observed in neuronal disorders with misfolded proteins , as will be discussed later.Neurologic disorders are numerous and diverse and can be caused by a variety of etiologic agents with many of the disorders being the consequence of the convergence of more than one etiopathogenic factor. An important group of neurological disorders are inherited, i.e., a mutated gene, or group of mutated genes are present in the genome of an individual which transmits it to its descendants. Some of these mutations are now well characterized ,142,143.The nervous system cells, Neuroinflammatory disorders often include cases also classified within other groups. Neuroinflammation occurs as a direct response of the glial cells against injury, microbial infection, chemical substances, autoimmunity, or neurodegeneration of nervous tissue, but when the activation of microglial or macroglial cells becomes aberrant it can trigger acute inflammatory responses that can progress toward chronicity and have serious pathogenic consequences. Chronic inflammation is typically associated with some neurodegenerative diseases such as AD and PD. These and other disorders, for instance MS and ALS, differ in pathophysiology and can cause memory and cognitive impairments or affect a person\u2019s ability to move, speak, and breathe. The outcome of a neurodegeneration is the loss of structural and functional neuronal integrity. Since there are several types of neurons and glial cells , their iNeurovascular diseases, owing to defects of blood vessels supplying blood to CNS, can increase the risk of stroke. These neurovascular deficits are involved in pathogenic mechanisms in various neurodegenerative diseases, as for instance in AD .Tumors are benign and malignant neoplasias of the CNS, PNS, autonomic nervous system, cranial nerves, and meninges . GenomicStudies on exosomes have contributed to increasing our current understanding of the pathogenesis of neurodegenerative disease. Since exosomal proteins were found accumulated in amyloid plaques in the brain of AD patients , the invThe likelihood of exosome involvement in AD pathogenesis was also suggested by the finding of hyperphosphorylated tau protein in exosomes from neural tissue in culture and in human CSF . Tau proCurrently, it is possible to diagnose AD when the disease is already established, e.g., when the patient has already developed dementia. The previous stages often remain asymptomatic, and these are the stages in which the patient could benefit most from treatment. Therefore, some sort of early diagnostic procedure is needed and, for this purpose, exosomes could be considered potentially useful biomarkers. Exosomes could act as A\u03b2 scavengers binding A\u03b2 to their surface and, subsequently, microglial cells would internalize \u201ccharged exosomes\u201d and process them for degradation ,159. ConAbnormalities in miRNA molecules are found in inflammatory cell populations or pathological samples of autoimmune disease . It was Tumor cells, derived from primary brain tumours or from metastases, use exosomes as packages to spread proteins and other molecules associated with malignancy ,170. ExoThe current diagnostic approaches for most neurological disorders are limited to evaluation of clinical symptoms and radiologic signs. Consequently, the diagnosis can be tardive and treatment often produces negligible benefits. Therefore, it is necessary to find biomarkers that can be measured with minimally invasive procedures if progress in early diagnosis and reliable and timely assessment of response to treatment are to be achieved. Within this context, exosomes appear as suitable biomarker candidates as the key specimens of liquid biopsies. Efforts should be made to standardize assays with high specificity and sensitivity that would extract as much clinically relevant information as possible from exosomes. This approach is promising, considering that exosomes are a showcase of molecules present in their cells of origin.Some properties of exosomes make them, in principle, convenient for use as drug carriers for delivery to the CNS. For example, exosomes can cross physiological barriers and can interact with plasma-cell membranes, which may eventually lead to their penetration into target cells. Current knowledge suggests that exosomes may have advantages in comparison with other drug delivery agents such as liposomes, for example, in what concerns safety and selectivity, but more research is needed to determine their practical value in clinics. Some of these issues are discussed below.In the last few years, research efforts have been focused on the manipulation of the exosomes\u2019 content and their targeting to the CNS pathological sites for treating specific pathologies. The potential application of exosomes and EV in general, as therapeutic tools, has led to the development of new and advantageous therapies, particularly for brain tumors. Illustrative examples pertain to exosomes from bone marrow and mesenchymal stem cells (MSCs) that werIn an animal experimental model of stroke, it has been shown that the intravenous administration of MSC-derived exosomes enhanced neurite remodelling, neurogenesis, and angiogenesis, leading to functional recovery . The effIn what pertains to the exosome-based strategies for the treatment of PD, the engineering of exosomes by electroporation with catalase can be mentioned . In a moIn a brain ischemia mouse model, engineered exosomes loaded with curcumin reached the target brain lesion after intravenous administration, supressing inflammation and apoptosis . The effThe possible toxicity of exosomal preparations and the side effects of their administration in neuronal tissue are still to be explored further and various technical hurdles need to be overcome. However, exosomes have a great potential to be part of a versatile strategy to treat neurological disorders for all the reasons discussed above, such as the requirement of minimally invasive techniques, low immunogenicity, and ability to cross the BBB and reach the target pathological cells.Incidence of CNS disorders is increasing worldwide, but no parallel progress in prevention and treatment occurs . Novel tDespite the range of available information about exosomes as potential disease biomarkers and the increasing number of clinical trials on exosome-based drug delivery strategies, in cancer, for example ,192, com"} +{"text": "As a nanoscale subset of extracellular vehicles, exosomes represent a new pathway of intercellular communication by delivering cargos such as proteins and nucleic acids to recipient cells. Importantly, it has been well documented that exosome-mediated delivery of such cargo is involved in many pathological processes such as tumor progression, cancer metastasis, and development of drug resistance. Innately biocompatible and possessing ideal structural properties, exosomes offer distinct advantages for drug delivery over artificial nanoscale drug carriers. In this review, we summarize recent progress in methods for engineering exosomes including isolation techniques and exogenous cargo encapsulation, with a focus on applications of engineered exosomes to target cancer metastasis. Consequently, they possess a similar fraction of the membrane phospholipid PE (phosphatidylethanolamine) in exosomes as their parent cells.97via negative staining. Size distribution and concentration, as well as sample purity, can be measured with DLS using a Zetasizer instrument and nanoparticle tracking analysis .via flow cytometry.21The characterization of exosomes can be performed visually, by using various dynamic light scattering (DLS) and microscopy techniques, and flow cytometry. Due to the small size range of exosomes, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) are frequently used to visualize exosomes It has been shown that cancer cells secrete more exosomes than their non-cancerous counterparts.36+) counts.+) into regulatory T cells.regs aids in the ability of the tumor microenvironment to suppress and evade an immune system response. T cell activation has also been inhibited by exosomes.via targeting TGF-\u03b2.+ T cell function.15Exosomes have been found to exhibit the ability to induce immune suppression.The process of EMT occurs when tissue epithelial cells possess altered biochemical factors that leave them with a more mesenchymal phenotype, aiding in functions such as migration and invasion.122via tail vein, intracardial or retro-orbital route.44Organotropism is defined as the non-random process which results in distant metastasis to specific organs.Angiogenesis and vascular permeability are activated and upregulated by exosomes derived from cancer cells. These exosomes carry pro-angiogenic factors such as VEGF, TIMP-1, IL-6, and FGF and cause their upregulation in recipient cells.96One crucial step in the study of exosomes is to isolate exosomes from a complex mixture of cell culture media, tissues or bodily fluids that contains cells, cell debris, other particulate components and macromolecules. An optimal method for exosome isolation is expected to exhibit high recovery yield and high purity of exosomes, and high efficiency as well. Several isolation techniques have been utilized in published exosome studies, each exploiting particular properties of exosomes, such as their density, shape, size, and unique surface proteins to aid their isolation Table\u00a0.Table\u00a01Sg) is performed to remove cells and large cellular debris from an exosome containing sample, e.g., conditioned cell culture media or biological fluid. Next to be removed are the smaller debris and intact organelles at 10,000\u201320,000\u00d7g. The final step is ultracentrifugation of the supernatant to form a pellet of exosomes.23Ultracentrifugation is considered the \u201cgold standard\u201d for exosome isolation and makes up\u2009>\u200950% of all exosome isolation techniques utilized in reported exosome research.Ultrafiltration, including syringe-driven filtration, may be the most straightforward method for exosome isolation.130g to remove particles much larger than exosomes before being concentrated with a concentrator with a uniform pore size of 13\u00a0nm or a ~\u2009100\u00a0kDa\u00a0MW cutoff (MWCO).77A commonly used protein concentrator has been reported to concentrate exosomes from urine samples with high yield and efficiency.For exosome isolation from cell culture supernatants, a 3-step sequential filtration has been reported. The first step is to remove floating cells and large cell debris using a 100-nm polyethersulfone (PES) filter.61Exosome isolation by ultrafiltration is much faster than that by ultracentrifugation and can be completed without using any other special equipment. However, exosomes may partly become deformed or broken when they are forced through nanoscale filters.61Size exclusion chromatography (SEC) is a promising method for exosome isolation because of its capability to separate nanoscale particles based on their hydrodynamic size.66Another size-based separation technique that has been applied to exosome isolation is Asymmetric Flow Field-Flow Fractionation (AF4).Surface proteins and other molecules that are unique to, or highly concentrated on, exosomes in exosome-containing samples offer opportunities for specific isolation by designing antibody-mediated immunoaffinity interaction.82et al. evaluated several commercially available kits for immunoaffinity-based isolation and modified their protocols to increase the purity of exosomes obtained.Zarovni et al. evaluated the feasibility of magnetic beads for isolating exosomes. As little as 1.0\u00a0mL of cell culture supernatant can be handled with a similar capture efficiency to that of ultracentrifugation.48The efficiency of immunoaffinity-based isolation may be further improved by using antibody-modified magnetic beads.Compared to ultracentrifugation for exosome isolation from cell culture medium, immunoaffinity-based isolation exhibits comparable yield with higher purity and advantages of ease of operation and much higher efficiency.37via a so-called polymer-based precipitation method, which is a widely used method to precipitate viruses and other macromolecules.90Exosomes can also be isolated Compared to other methods of isolation, polymer-based precipitation is easy to use, scalable for large sample volumes and does not require any specialized equipment or lengthy run time.83As a rapidly-growing engineering field, microfluidics has been widely used for the separation of particles ranging from nanoscale to microscale such as cells and nanoparticles.32et al. demonstrated a microchip that selectively traps exosome-like lipid vesicles 40\u2013100\u00a0nm, while sieving out proteins and cellular debris.119Techniques that have been incorporated in microdevices for exosome isolation include immunoaffinity, sieving, and trapping exosomes on porous structures.As mentioned above, the exosome isolation techniques are based on particular properties of exosomes, each with its advantages and disadvantages. Combining two or more technique could further improve the isolation of exosomesExosomes play significant and diverse roles in intercellular communications, particularly in long-distance intercellular signaling. This mechanism of communication is highly robust and efficient in exchanging information between cells.47Inspired by their physiochemical properties and natural cargo-delivering capability, researchers have also explored the potential of exosomes to deliver various exogenous therapeutics.via exosomes or (B) loading parental cells with a drug, which is then released in exosomes.7Due to the availability of various cellular engineering methods, most modifications of exosomes have been performed on parental cells which are then cultured to secrete modified exosomes.via Lipofectamine) to load cargo or result in the desired genetic expression.5Protein sequences, along with different types of RNA, are frequently used for cell transfection, ultimately altering the phenotype of the released exosomes.via transfection.135TRAIL is a therapeutic that has been loaded into cells via electroporation with doxorubicin (Dox) and injected intravenously into the murine model, which resulted in reduced breast cancer tumor growth.110Transfection has been employed to alter exosomes derived from murine immature dendritic cells (DCs). For pre-isolation, immature DCs were transfected with a viral vector to express Lysosome-associated membrane protein 2 (Lamp2b) fused to the \u03b1\u03b3 integrin-specific iRGD peptide, a membrane protein of exosomal origin, to enhance targeting efficiency to the tumor site.One way that exosome transfection has been accomplished is through miRNA expression vectors, which can result in exosomes carrying miRNA.For harnessing exosomes for cancer treatments, a study was performed by modifying an invasive triple negative breast cancer cell line (Hs578T) to overexpress miR-134.84et al., used electroporation to transfect marrow stromal cells with a miR-146b expression plasmid.Similarly, in an effort to analyze the efficacy of exosomes as carriers for anti-tumor microRNAs, Katakowski In vitro, HepG2 and Huh7 hepatoma cells were exposed to 122-Exo and chemotherapeutic drugs, revealing a decrease in cell viability when compared to controls. In vivo, intratumoral injections of 122-Exo increased the sensitivity of HCCs to sorafenib, as observed by reduced tumor size in the mouse model when compared to controls.67To heighten the sensitivity of Hepatocellular carcinoma (HCC) cells to chemotherapeutic agents, it was explored whether exosomes could be delivered to enhance expression of miR-122, a microRNA found to increase the chemosensitivity of HCCs.via lipofection with miRNA let-7a, a microRNA reduced in various cancers including breast cancer, whose expression suppresses tumor growth. Exosomes isolated from these cells were intravenously injected into mice with HCC70 breast cancer, successfully delivering the microRNA to the tumor site, as evidenced by reduced tumor growth when compared to control.85Exosomal delivery was also used to deliver microRNA to breast cancer cells expressing epidermal growth factor receptor (EGFR).et al.71Viral packaging can result in exosomes loaded with nucleic acids. This method employs Adeno-associated vectors (AAV) to load exosomes with a viral vector, and has been termed as \u201cvexosomes\u201d (vector-exosomes) in a study by Maguire A major benefit of exo-AAV is that they are able to evade neutralizing antibodies, compared to wild type AAV vectors that have the potential to evoke an immune response that can block their delivery.46Exosomes can be preloaded with a protein or drug of choice when parental cells are loaded with exogenous cargo. The biogenesis process results in the preloading of the exosomes released from the cells, and is especially beneficial for oligonucleotides.5Extracellular vesicles, characterized as exosomes, were conditioned to bear heat shock proteins (HSPs) and isolated from resistant hepatocellular carcinoma cells (HepG2) that had been previously treated with resistant or sensitive anti-cancer cells.70sn-glycero-3-phosphoethanolamine) or solid HSPC-based (hydrogenated soy phosphatidylcholine). Exosomes were collected and isolated from the culture media, their protein contents analyzed and broken up into categories . The resulting exosomes were then delivered to the B16BL6 melanoma cell line, as well as a murine colorectal cancer cell line (C26), and analyses were performed for studying how exosomal expression of the different proteins impacted their uptake by cancer cells.Liposomes formed the basis of a study involving exosome delivery of proteins to cancer cells, where they were co-incubated with a murine melanoma cell line (B16BL6).via \u201cliposome-based cellular engineering\u201d, which attempts to engineer parental cells via membrane fusogenic liposomes (MFLs).60Drug preloading for exosomal delivery of cancer treatments can be accomplished in vitro and in vivo to the human pancreatic adenocarcinoma cell line (CFPAC-1) inhibited cancer cell growth and proliferation both in vitro and in vivo.88Macrovesicles (MVs) and exosomes secreted from these MSCs have been frequently studied for therapeutic benefits in regenerative medicine due to their paracrine secretions, in the form of extracellular vesicles such as MVs and exosomes.via liposome delivery to parent cells.60As in the case of hydrophilic drug delivery, membrane fusogenic liposomes can also be used for the delivery of hydrophobic drugs. This method of parental cell engineering was used for anti-tumor drug loading when EVs containing the chemotherapeutics PTX and tirapazamine were co-incubated in Transwell experiments with B16F10 (melanoma) or MDA-MB-231 (late-stage breast cancer) cells. MFLs were able to reduce cell viability when compared to controls, demonstrating the efficacy of preloaded anti-tumor exosomes via valence bond or other specific conjugation methods.104In situations where engineering of exosomes at the\u00a0cellular level is not feasible, exosomes derived from various origins can be engineered to carry functional molecules after being isolated.via chemical methods. in vivo.102A few reports have demonstrated the feasibility of exosome surface modification et al. conjugated fluorescent molecules to the surface of exosomes derived from mouse 4T1 breast cancer cells using click chemistry, a highly efficient and widely-used bioconjugation method.et al. transfected human embryonic kidney 293T exosomes to express a surface luciferase with a fused biotin domain which was then coupled with fluorescent Alex Fluor\u00ae 680-Streptavidin. in vivo and analysis\u00a0of their blood circulation life.58To label exosomes for an imaging modality, Smyth Genetic engineering has also been reported as a method to modify the surface of exosomes.110Exosome membranes can be loaded with hydrophobic therapeutics to increase drug solubility and stability while hydrophilic therapeutics such as RNA can be encapsulated in exosomes to improve cellular delivery.via sucrose gradient centrifugation. Characterization of curcumin-loaded exosomes revealed higher solubility, stability, and bioavailability than free curcumin. In another study, the same group loaded JSI-124, a potent inhibitor of JAK/STAT3 signaling pathway with anti-tumor activity, into EL-4 exosomes, suggesting the potential of exosomes as a general delivery vehicle for hydrophobic compounds.100Similar to their passive encapsulation into parental cells, hydrophobic drugs can be inserted into the membrane simply by incubation with exosomes. One of the earliest studies of this kind is the exosomal delivery of curcumin, an anti-inflammatory agent.To develop exosome-encapsulated PTX to overcome multiple drug resistance (MDR) in cancer cells, Batrakova\u2019s group compared three different encapsulation methods, incubation at room temperature, electroporation, and sonication to encapsulate PTX into exosomes derived from mouse macrophage RAW 264.7 cells.55et al. dispersed purified exosomes derived from\u00a0an immature mouse DC line in a Dox solution before electroporation was applied.110Electroporation is more commonly used to load small hydrophilic compounds or nucleic acids into isolated exosomes.et al. reported the first siRNA delivery by isolated exosomes in 2011.et al. demonstrated that B cell-derived exosomes can deliver an exogenous miRNA-155 mimic into hepatocytes or macrophages to inhibit malignant growth. Unlike in parental B cells, baseline levels of miRNA-155 was found to be very low in B cell-derived exosomes. The authors optimized the loading efficiency of miRNA-155 by electroporation at various RNA-to-exosome ratios. Exosomes loaded with miRNA-155 mimic significantly increased miRNA-155 levels in primary mouse hepatocytes and the liver of miRNA-155 knockout mice.79As exosomes naturally deliver nucleic acids to recipient cells, exosomes have been expected to deliver exogenous siRNA in an efficient and targeted manner. Similar to their loading into cells, loading of siRNA into exosomes can be achieved by electroporation. Alvarez-Erviti As mentioned above, previous reviews have summarized exosome engineering to target various diseases, in particular cancer.Phase I clinical trials have demonstrated the feasibility of large-scale production of DC-derived exosomes and the safety of the exosomes in patients with colorectal\u00a0cancer, lung cancer, and melanoma.80Encouraging results were also obtained in a Phase II trial testing DC-derived exosomes as maintenance immunotherapy after induction chemotherapy in patients with metastatic lung cancer.et al. combined vaccination with tumor exosome-loaded DCs (DC-TEX) with drugs affecting myeloid-derived suppressor cells (MDSC). In the study, autologous DCs were loaded with PaCa cell-derived exosomes to vaccinate for PaCa in xenograft mice together with drugs such as Gemcitabine (GEM), all-transretinoic acid (ATRA) and sunitinib.128A promising option in cancer immunotherapy is active vaccination with autologous DCs loaded with tumor-associated peptides. However, the immune response of pancreatic cancer (PaCa) by this strategy is often found suppressed. To overcome this issue, Xiao et al. to develop exosomes derived from normal fibroblast-like mesenchymal cells to carry siRNA specific to oncogenic KRAS. Compared to control liposomes, the engineered exosomes exhibited an enhanced retention in circulation and a subsequent enhanced efficacy that is dependent on CD47. It was confirmed that the enhanced retention of exosomes is due to CD47-mediated protection of exosomes from phagocytosis by monocytes and macrophages. The engineered exosomes suppressed cancer in multiple mouse models of PaCa and significantly increased their overall survival. To take this discovery into human translation, the same lab reported bioreactor-based procedures employing good manufacturing practice (GMP) standards to generate large scale production\u00a0of siRNA-loaded clinical-grade exosomes. The clinical-grade GMP exosomes were tested in multiple in vitro and in vivo studies to confirm suppression of oncogenic Kras and an increase in the survival of several mouse models with PaCa.76The mutant form of the GTPase KRAS is a key driver of PaCa, which controls macropinocytosis in PaCa cells and increases exosome uptake. This led Kamerkar MDR1 and its sensitive counterpart MDCKWT.To overcome MDR in cancer cells, PTX-loaded exosomes were used to treat a drug resistant cancer cell line, MDCKExosomes are specialized intercellular messengers that alter the functional state of their target cells by delivering cargo such as proteins and nucleic acids from their parental cells. The role of exosomes in cancer including metastasis has been intensively investigated. Understanding of their properties and activities have provided a\u00a0solid foundation to engineer exosomes for the targeting of metastasis, which could significantly increase survival among cancer patients. After a decade of research, many engineered exosome engineering methods including those for isolation and cargo incorporation have proven to be successful for modifying exosomes with desirable diagnostic and therapeutic functionalities. Application of engineered exosomes to target metastasis have yielded encouraging results that support further development toward clinical practice."} +{"text": "Exosomes are membrane-enclosed entities of endocytic origin, which are generated during the fusion of multivesicular bodies (MVBs) and plasma membranes. Exosomes are released into the extracellular milieu or body fluids; this process was reported for mesenchymal, epithelial, endothelial, and different immune cells (B-cells and dendritic cells), and was reported to be correlated with normal physiological processes. The compositions and abundances of exosomes depend on their tissue origins and cell types. Exosomes range in size between 30 and 100 nm, and shuttle nucleic acids , microRNAs), proteins, and lipids between donor and target cells. Pathogenic microorganisms also secrete exosomes that modulate the host immune system and influence the fate of infections. Such immune-modulatory effect of exosomes can serve as a diagnostic biomarker of disease. On the other hand, the antigen-presenting and immune-stimulatory properties of exosomes enable them to trigger anti-tumor responses, and exosome release from cancerous cells suggests they contribute to the recruitment and reconstitution of components of tumor microenvironments. Furthermore, their modulation of physiological and pathological processes suggests they contribute to the developmental program, infections, and human diseases. Despite significant advances, our understanding of exosomes is far from complete, particularly regarding our understanding of the molecular mechanisms that subserve exosome formation, cargo packaging, and exosome release in different cellular backgrounds. The present study presents diverse biological aspects of exosomes, and highlights their diagnostic and therapeutic potentials. The existence of exosomes as extracellular vesicles (EVs) was first reported by Harding et al. and Johnstone et al. ,2. UnlikJournal of Extracellular Vesicles, and the establishment of the Exocarta and Vesiclepedia databases, which are dedicated to extracellular vesicles. The extracellular RNA (exRNA) research portal is the result of an initiative by the Extracellular RNA Communication Consortium (ERCC) that provides a catalog of extracellular RNAs , and reports on the mechanisms of exRNA generation, secretion, transport, therapeutic uses, and their uses as biomarkers of disease. In addition to their importance as signaling molecules for intercellular communication and regulation, exosomes offer potential means of ameliorating pathogenic immune responses, diagnosing different diseases, and delivering therapeutics. Here, we summarize essential findings of exosome biology, and provide an up-to-date account of their diverse physiological and pathological functions.Exosome discovery is bearing fruit, as evidenced by an enormous increase in the number of studies on exosome biology and the establishment of scientific societies like the American Society for Exosomes and Microvesicles (ASEM) and the International Society for Extracellular Vesicles (ISEV). This explosive growth of exosome biology even resulted in a journal g is routinely used to obtain exosomes from culture supernatants. Although the technique excludes contamination by dead cell debris, it results in mixed fractions of exosomes, protein aggregates, and vesicular structures. Other isolation methods include serial filtration [Exosomes made their entrance into the scientific world three decades ago. Late endosomes were considered as pre-degradative compartments, and the vesicular structures secreted by late lysosomes were considered as the membranous entities of dying cells. As technical problems in the isolation methods hamper their separation from other vesicle types, no consensus was reached on their isolation methods. The existence of exosomes became apparent when reticulocyte culture supernatant was purified by ultracentrifugation, and they were only recognized as functional entities by electron microscopy . Ultraceltration , immunoaltration , and comltration ,18,19,20Observations of exosomes by whole-mount electron microscopy revealed them to be \u201csaucer-like\u201d or \u201cdeflated-football\u201d shaped, believed to be due to vesicle collapse during sample preparation . AlthougThe budding of interluminal vesicles from endosomal compartments and their joining together results in the production of multivesicular bodies (MVBs) . Though Fluorescence-activated cell sorting (FACS), Western blotting, and mass spectrometry are commonly employed to decipher the exact compositions and to identify the molecular constituents of exosomes ,19,37. DN-ethylmaleimide-sensitive factor (NSF) attachment protein receptors (v-SNAREs) and target SNAREs (t-SNAREs), the secretion of exosomes with Wingless (Wnt) as a signature depends on arginine (R)-SNARE Ykt6 [Exosome secretion into the extracellular milieu modulates gene expression, function, and even cellular differentiation programs. The protein content and genetic material of exosomes can even change the morphology of a recipient cell by interfering with its signaling components. Though little is known of the mechanisms driving MVB to plasma membrane fusion, a study of reticulocytes revealed that exosome secretion is dependent on vesicular-associated molecular pattern 7 (VAMP7) function . DespiteARE Ykt6 . The SNAARE Ykt6 . AdditioARE Ykt6 . ExosomeARE Ykt6 ,43.+ or CD8+ T cells [Over the past decade, exosomes were implicated in diverse activities in biological systems, possibly by modulating intercellular communication or action at a distance ,44. Thei T cells ,51. Howe T cells . By infl T cells . Further T cells . By actiS-transferase, which are responsible for the detoxification of toxins and drugs in target cells [Exosomes released by epithelial cells were found to be involved in antigen presentation during inflammatory conditions. It may be that these extracellular vesicles are responsible for providing fixed cells with the ability to act at a distance . In the et cells ,64,65.Exosomes containing oncogenic materials are referred to as oncosomes, and they mediate the intercellular transport of these mutant molecules in a systemic manner ,67,68,69Given their wide-ranging functions, exosomes have huge diagnostic and therapeutic potential. By regulating physiological functions, such as angiogenesis, intercellular communication, coagulation, immune response, and cell survival, exosomes are of immense interest to the scientific fraternity worldwide. Their secretions from cells into body fluids , saliva, and others) are widely reported . ExosomeWith specific protein signatures , RNA (mRNA and miRNA), and characteristic lipid contents, exosomes in body fluids were investigated as diagnostic markers for the early detection of various diseases ,119. ExoIncreases in the miRNA content released as part of exosomes into body fluids provide insight into the progression of the disease ,121. CanHuman saliva, which is another indispensable source of exosomes, contains nucleic acids and proteins, and provides diagnostic signatures for different diseases . Levels Urine also serves as an efficient source of exosomal markers of urogenital diseases. Proteomic analysis of urine helped identify eight proteins useful for the detection of bladder cancer . ExosomeThe properties of exosomes, which include bioavailability, distribution, and stability under in vivo and in vitro conditions, and their abilities to cross the blood\u2013brain barrier (BBB) and regulate gene expression via the transfer of miRNA and small interfering RNA (siRNA) to target cells means they are preferred over other EVs as potential therapeutics . Further4 or H2O2 and effectively rescued recipient mice from CCl4-induced liver failure. The mechanism of rescue by hucMSC-Ex-derived cells was attributed to glutathione peroxidase 1 (GPX1), which detoxifies CCl4 and H2O2, thereby reducing oxidative stress and apoptosis. It was also shown that a knockdown of GPX1 in hucMSCs abolished the antioxidant and anti-apoptotic abilities of hucMSC-Ex, and reduced the hepato-protective effects of hucMSC-Ex in vitro and in vivo.Mesenchymal stem cells (MSCs) are stromal cells with well-known therapeutic potentials. These cells are present in bone marrow, umbilical cord, and adipose tissue . The exoTGFBR1 in ECs, following delivery of functional let-7b-5p via PF exosomes. Downregulation of let-7b-5p miRNA in PF exosomes impaired the angiogenic response by ECs. At a functional level, the authors reported that PF exosomes enhanced survival, proliferation, and networking of cultured endothelial cells (ECs), and restored the pro-angiogenic function of ECs depleted of their endogenous miRNA content.By aiding the adhesion of hematopoietic stem-cell progenitor cells to endothelia, microvesicles derived from platelets (PMVs) supported the engraftment of transplanted stem cells in lethally irradiated mice . BeltramCardiosphere-derived cells (CDCs) induce the therapeutic regeneration of the infarcted human heart by stimulating angiogenesis and causing functional improvements of infarcted myocardium. Interestingly, CDCs reduced scar sizes and the growth of new functional myocardium, which was previously considered an irreparable form of injury . IbrahimExosomes are known to have neuroprotective effects, that is, they aid neuron healing and the regeneration of peripheral nerves, and also act as mediators of neurodegenerative diseases. Furthermore, their ability to cross the blood\u2013brain barrier makes them indispensable neurotherapeutic carriers of drugs and therapeutics. In the nervous system, exosome-mediated neuronal communication facilitates cell-to-cell interactions. The transfer of miRNAs and protein entities from glial cells to neural exons was reported a few years ago . SulfatiExosomes are importantly involved in intercellular communication and in the pathogeneses of various human diseases. Because exosomes are readily accessible in body fluids, their genetic profiles provide new diagnostic and prognostic tools and open new therapeutic possibilities. Their abilities to carry mRNAs, miRNAs, and non-coding RNAs provide an efficient means of controlling protein expressions at a distance in different target cells. The use of exosomes as delivery vehicles offers significant advantages over existing delivery systems due to their small size, non-toxic natures, and target specificities . InteresAs our understanding of the biology of exosomes increases, so will our knowledge of design principles and exosomal conjugates. DC-derived exosomes engineered to express rabies virus glycoprotein showed positive results in the delivery of siRNA across the BBB in murine models . To faci"} +{"text": "Exosomes are cell-derived nanovesicles that are involved in the intercellular transportation of materials. Therapeutics, such as small molecules or nucleic acid drugs, can be incorporated into exosomes and then delivered to specific types of cells or tissues to realize targeted drug delivery. Targeted delivery increases the local concentration of therapeutics and minimizes side effects. Here, we present a detailed review of exosomes engineering through genetic and chemical methods for targeted drug delivery. Although still in its infancy, exosome-mediated drug delivery boasts low toxicity, low immunogenicity, and high engineerability, and holds promise for cell-free therapies for a wide range of diseases. For the first time, these vesicle-like structures were named exosomes In 1981, Trams et al. discovered a group of vesicle-like structures with diameters 40-1000 nm smaller than those of multivesicular bodies by transmission electron microscopy Exosomes mediate intercellular communications. Through exosomes, donor cells can transfer exogenous substances, such as proteins, mRNAs, microRNAs (miRNAs), and lipids, to recipient cells. Consequently, these naturally-equipped nanocarriers have been used for drug delivery Figure ) Originating from the endocytic pathway, exosomes are released from MVBs, which contain multiple vesicles generated through exocytic fusion with the cell membrane. Exosome generation includes four steps: budding, invagination, MVB formation, and secretion. As a form of budding, invagination of the cell membrane forms clathrin-coated vesicles, and clathrin vesicles enter the cytoplasm to form early endosomes. Proteins, lipids, and nucleic acids are specifically sorted and encapsulated to form multiple intraluminal vesicles (ILVs), which are the precursors of exosomes. After late endocytosis, multiple ILVs further develop into late endosomes (LEs). Then, a proportion of the LEs enters the lysosomal pathway, while the others fuse with the cell membrane, ultimately releasing multiple vesicle structures into the extracellular matrix in the form of exosomes . The ESCRT system contains four protein complexes: ESCRT-0, ESCRT-1, ESCRT-2, and ESCRT-3 together with auxiliary proteins such as VTA\u20431, ALIX, and VPS4. These proteins act synergistically in the stepwise formation of MVBs Figure ) The mechanism by which exosomes enter a recipient cell can involve multiple pathways at the same time. Exosomes can directly fuse with the plasma membrane, and can also be taken up through phagocytosis, micropinocytosis, and endocytosis mediated by lipid raft, caveolin, or clathrin . To date, the ExoCarta exosome database (http://www.exocarta.org) has collected 9769 proteins, 3408 mRNAs, 2838 miRNAs, and 1116 lipids that have been identified in exosomes from different types of cells and from multiple organisms. Although the cell source determines the kinds of proteins secreted by exosomes, 80% of the proteins in exosomes are highly conserved among different cells. Some are used as biomarkers of exosomes, including ALIX, TSG101, heat shock proteins, and the tetraspanins CD63, CD9, and CD81. These transmembrane proteins account for the targeting and selective entry of exosomes to recipient cells Table 1). Nucleic acids are abundant in exosomes, including mRNAs, miRNAs, mitochondrial DNA and piRNAs, lncRNAs, ribosomal RNAs, snRNAs, and tRNAs. Although the nucleic acids encapsulated in exosomes are degraded fragments of approximately 200 bp, they can still impact protein synthesis inside the recipient cell. Lipids in the membrane of exosomes include cholesterol (chol), phospholipids, phosphatidylethanolamines, polyglycerols, and diglycerides. Notably, exosomes have different lipid composition, distribution, and content than the cytoplasmic membrane. The lipid molecules are involved not only in maintaining the morphology of exosomes, such as their stability and structural rigidity, but also in many biological processes. For example, they can serve as signaling mediators by interacting with prostaglandin and phospholipase C and D Exosomes can transfer various types of cargoes, including DNAs, RNAs, lipids, metabolites, and proteins Despite being vehicles of natural origin, exosomes can be conveniently surface modified. One obvious goal of surface engineering is to confer cell type targeting specificity. Modification strategies include genetic engineering and chemical modification (Table 1). First, ligands or homing peptides are fused with transmembrane proteins that are expressed on the surface of exosomes. Subsequently, donor cells transfected with plasmids encoding the fusion proteins secrete engineered exosomes bearing targeting ligands on their surface. Examples of genetically engineered exosomes as drug delivery systems are summarized in Table 2.Genetic engineering of exosomes is a convenient method for imparting exosomes with new properties Currently, LAMP-2B is the most widely used exosomal surface protein to display a targeting motif. LAMP-2B, a member of the lysosome-associated membrane protein (LAMP) family, predominantly localizes to lysosomes and endosomes, with a smaller fraction circulating to the cell surface. It has been reported that LAMP-2B protein is abundantly expressed on dendritic cell-derived exosomes. Researchers proved that the N-terminus of LAMP-2B is displayed on the surface of exosomes and can be appended with targeting sequences KRAS gene knockdown and tumor growth suppression Figure 5) Cell-specific binding peptides targeting specific organs or tissues can be screened and selected by phage display and genetically modified at the N-terminus of LAMP-2B to realize their targeting effects. Donor cells transfected with the plasmid produce exosomes displaying the engineered peptide ligands. For example, rabies virus glycoprotein (RVG) peptide (TIWMPENPRPGTPCDIFTNSRGKRASNG) shows selective binding to acetylcholine receptors and has been used to develop neuro-specific exosomes to deliver drugs to the central nervous system KD values in the low nanomolar range towards receptors on target cells. For example, to improve therapy of chronic myeloid leukemia (CML), researchers modified the surface of exosomes by fusing interleukin-3 (IL-3) to the N-terminal of LAMP-2B. IL-3 is the native ligand of interleukin-3 receptor \u03b1 (IL-3R\u03b1), which is highly expressed on CML blasts. IL-3 exosomes loaded with imatinib and BCR-ABL siRNA effectively killed CML cells and extended the overall survival rate of xenografted mice. In vivo tracking of these engineered exosomes showed rapid migration to the CML xenograft tumors . developed cardiac cell-targeted exosomes expressing ischemic myocardium-targeting peptide in MSCs. IMPT exosomes specifically localized to the ischemic myocardium area and exerted cardioprotective effects in acute myocardial infarction LAMP-2B can also be genetically fused with targeting proteins or antibody fragments to display antibodies on exosomes. Compared with peptides, which have modest binding affinities, antibodies or affibodies can achieve higher affinities, with Besides LAMP-2B, the transmembrane protein platelet-derived growth factor receptor (PDGFR) is commonly used for membrane display. In one example, GE11 (YHWYGYTPQNVI) was genetically fused to the PDGFR transmembrane region using a pDisplay vector to produce GE11 exosomes in 293T cells. GE11 exosomes showed high affinity for epidermal growth factor receptor (EGFR)-overexpressing cancer cells and low mitogenic activity, making them a promising delivery system for EGFR-targeted therapy. In animals, GE11 exosomes loaded with the antitumor nucleic acid inhibitor miRNA let-7 significantly inhibited tumor growth, consistent with their tumor-targeting ability The tetraspanin superfamily CD63/CD9/CD81, with their two extracellular loops, can also be engineered to display targeting sequences or probes. For example, the fluorescent protein pHluorin was inserted into the small extracellular loop to probe for exosome secretion and uptake Exosomes with surface-displayed antigens can also be used as anticancer vaccines. In one study, fusion of ovalbumin (OVA) antigen to CD63 produced OVA exosomes that improved the immunogenicity of DNA vaccines and prevented tumor growth in a xenograft model Exosome membrane attachment via a signal peptide is also a readily achievable method for membrane display Table 3. For example, the amine groups of exosomal proteins can be easily modified with alkyne groups. Then, the alkyne-labeled exosomal proteins can be bio-orthogonally couple to azide-containing reagents through copper-catalyzed azide-alkyne cycloaddition (CuAAC) \u201cclick\u201d reactions. In a proof-of-concept study, this approach was utilized to modify the exosome surface with both a small molecule dye and a larger azide-containing model protein Although less explored, the surface of exosomes can be modified via chemical methods. Examples of chemically modified exosomes for targeted drug delivery are summarized in sn-glycero-3-phosphoethanolamine (DSPE-PEG) can accumulate in the exosome membrane. This method was successfully used to immobilize DSPE-PEG-RGD on exosomes. When combined with the tumor-specific targeting ligand folate, nearly all injected RGD exosomes concentrated in tumor areas. Chemically modified RGD exosomes were also combined with photothermal therapy and chemotherapy for pro-angiogenic treatment. As sigma receptors are overexpressed in lung cancer, they have been proposed as receptors for targeted exosome delivery. Anisamides, including aminoethyl anisamide (AA), are high-affinity sigma-selective ligands. AA linked to DSPE-PEG was successfully conjugated to the exosome membrane Insertion of amphipathic molecules into the lipid bilayer of exosomes represents another chemical modification strategy. Previous research has demonstrated that polyethylene glycol (PEG)-grafted 1,2-dioleoyl-Paclitaxel (PTX)-loaded AS1411-chol exosomes were efficiently delivered to target cancer cells, providing a promising delivery platform for cancer therapy. Another study labelled exosomes with ApoA-1 mimetic peptides conjugated to lipids, which significantly enhanced their selective internalization by primary glioma cells . used electrostatic interactions to induce fusion of cationic lipids with exosomes The lipid bilayer membrane of exosomes can spontaneously fuse with other types of membrane structures. In one study, fusogenic exosomes harboring the viral fusogen vascular stomatitis virus (VSV)\u2010G protein directly delivered membrane proteins into target cells, thus providing a new tool for membrane protein therapy Figure 6. In genetic engineering approaches, a targeting ligand is genetically fused with an exosome membrane protein and subsequently overexpressed in the donor cells. The donor cells thereby produce genetically engineered exosomes that display the targeting ligand. Genetic engineering of exosomes represents a highly accessible strategy for the display of functional ligands on the exosomal membrane, but requires plasmid construction and overexpression of the proteins in the donor cells. Lipids or a bioconjugation reaction can also anchor a targeting moiety to the exosome membrane. Chemical methods rely on bioconjugation of the targeting ligand with surface proteins, but inactivation of the surface protein or aggregation of the exosome may occur during the chemical manipulation. Despite their limitations, both approaches have been successfully implemented.The procedures for creating cell-specific exosomes by genetic engineering or surface chemistry approaches are summarized in Exosomes can transfer encapsulated proteins and genetic information to recipient cells and act as information messengers between cells. The surface molecules anchored on exosomes from different cell sources vary, which endows them with selectivity for specific recipient cells. Surface engineering aims to increase the local concentration of exosomes at the diseased site, thereby reducing toxicity and side effects and maximizing therapeutic efficacy. Recently, a series of transactions by large pharmaceutical companies have shown that the industry expects exosomes to deliver drugs to hard-to-reach tissues Cell-targeting exosomes can be used for imaging studies or diagnosis. For example, bioluminescence and fluorescence tracking of exosomes were realized by enzymatic biotinylation of exosomes with biotin ligase"} +{"text": "Exosomes, nanometer-sized lipid-bilayer-enclosed extracellular vesicles (EVs), have attracted increasing attention due to their inherent ability to shuttle proteins, lipids and genes between cells and their natural affinity to target cells. Their intrinsic features such as stability, biocompatibility, low immunogenicity and ability to overcome biological barriers, have prompted interest in using exosomes as drug delivery vehicles, especially for gene therapy. Evidence indicates that exosomes play roles in both immune stimulation and tolerance, regulating immune signaling and inflammation. To date, exosome-based nanocarriers delivering small molecule drugs have been developed to treat many prevalent autoimmune diseases. This review highlights the key features of exosomes as drug delivery vehicles, such as therapeutic cargo, use of targeting peptide, loading method and administration route with a broad focus. In addition, we outline the current state of evidence in the field of exosome-based drug delivery systems in systemic lupus erythematosus (SLE), evaluating exosomes derived from various cell types and engineered exosomes. Autoimmune diseases are among the leading causes of morbidity and mortality associated with chronic disease worldwide, especially in women, who comprise more than 90% of affected patients ,2. AutoiExosomes are lipid-bilayer-enclosed extracellular vesicles released by many cell types in both normal and pathological conditions, and which transport nucleic acids, lipids and proteins between cells ,19. As dCurrent evidence shows that exosomes are able to modulate the immune system, as shuttles for antigen presentation ,21,22,23Due to their suitable proprieties, as well as their known therapeutic effects, exosomes are attracting the curiosity of researchers to develop exosome-based drug delivery systems. This review centers first on the biogenesis of natural exosomes and their function in SLE. Next, we will narrow our focus to identify the key components enabling successful exosome-based drug delivery, to unravel the potential of exosomes for disease treatment, finally covering new insights in exosome-based drug and gene delivery for future clinical use in SLE. Extracellular vesicles are small spherical lipid bilayer-coated vesicles, secreted by multiple cell types and present in many body fluids, which mediate intercellular communication ,34. NowaThe most broadly studied EVs are exosomes, 40\u2013150 nm in diameter, endosome-derived and originating from intraluminal vesicles (ILVs) that reside inside multivesicular bodies (MVBs) . NucleicWhen released to the extracellular space, exosomes interact with their acceptor cells in different ways. Information transmission can occur at the cell surface without delivering any cargo (surface binding), as occurs during immune responses ,42, but Indeed, due to their involvement in normal and pathological cellular physiology, EVs have been recognized as good biomarkers in a wide variety of diseases including cancer, cardiovascular diseases, nephropathies and autoimmune diseases ,62,63,64Systemic lupus erythematosus is a prototypic autoimmune disease characterized by diverse immune disturbances. One of the serologic hallmarks of SLE is the production of antibodies to nuclear molecules (antinuclear antibodies (ANA)) ,4. TheseSeveral reports indicate higher levels of circulating IC-carrying MPs in SLE; as an example, Ullal et al. demonstrated that MPs display DNA and nucleosomal molecules in an antigenic form and could represent a source of ICs in SLE . Another+ in patients with SLE [Given their composition and immune properties, EVs act as proinflammatory mediators in SLE . In one with SLE . Anotherwith SLE . Finallywith SLE . These fSeveral studies reported EVs as reliable biomarkers of disease activity beyond their role in regulating immune responses, offering a valuable complement to classical laboratory markers ,78. NielMany recent studies highlight exosomal-derived miRNAs levels to identify distinct miRNA to discriminate LN. Our group revealed increased urinary exosomal miRNAs levels in patients with SLE, discriminating LN , and in In summary, quantity and phenotype of circulating EVs may be useful as novel biomarkers of activity and progression of SLE, providing a new therapeutic approach.Research interest in exosomes has grown dramatically during the last few years due to their unique properties. Transport inside exosomes allows concurrent intercellular communication by delivering different signals simultaneously. Unlikely free circulating soluble factors, they have the ability to release large amounts of functional molecules to recipient cells . MoreoveThe potential use of exosomes in therapy is based on the cargos they deliver to recipient cells. Endogenous cargos with known beneficial properties or artificially modified molecules can be loaded using a range of methods. Likewise, different exosome administration routes for therapy purposes have been described, with novel methods being developed during recent years .Since of exosomes have an inherent ability to carry different types of active molecules, they can be biologically or chemically loaded to deliver enhanced or broadened therapeutic compounds, representing an informed way to treat many diseases. Research as drug delivery vehicles is based mainly on the transfer of small RNAs, although other possibilities are being explored.Exosome based-miRNA therapy is the most developed area so far, since miRNAs function as gene expression regulators and have been implicated in the pathophysiology of numerous diseases. In addition, exosomal ability to carry therapeutic miRNAs has been extensively analyzed, demonstrating high effectiveness . This thSimilarly, mesenchymal stem cell (MSC)-based therapies have also demonstrated protective effects by promoting tissue repair, apoptosis and fibrosis inhibition or stimulation of cell proliferation ,92. MSCsPreliminary studies of the time laid the groundwork for pursuing exosomes as the best drug delivery system option for miRNA therapy, and paved the way to develop multiple strategies for refinement of methods and for direct targeting of exosomes to cell types.Small RNA interference (siRNA) has become a promising therapeutic strategy for knocking down targeted genes; however, their use as therapeutic compounds has limitations, as they are extremely hydrophilic and easily degraded . MoreoveThere is emerging interest in long non-coding RNAs not only as biomarkers but also as therapeutic agents due to their ability to interact with miRNAs. lncRNAs are naturally carried by EVs and are been increasingly documented as key mediators of EV biological effects . TherapeAs shown before, exosomes are promising targets for therapeutic delivery based on small nucleic acids such as siRNAs and miRNAs, which are successfully loaded inside EVs. However, the use of exosomes as drug delivery systems of exogenous DNAs remains poorly researched despite the widely reported presence of different DNA species inside exosomes ,117. LoaIn addition to different nucleic acids, exosome transported proteins are also being used as therapeutic cargos, albeit less commonly ,124. AspApart from the natural cargos of exosomes , their excellent biological properties have driven their use as drug delivery systems of synthetic compounds have been attempted to improve the efficiency and specificity of EVs delivery. Exogenous loading after EV isolation, which can be divided into passive and active processes, includes incubation as a passive option and sonication, extrusion, freezing and thawing cycles, electroporation and chemical-based transfection as active processes where the use of electric field or surfactants are required for EV membrane permeabilization. Endogenous manipulation refers to introducing the cargo of interest into EV-producing cells, commonly transfecting cells with expression vectors. In addition, modification or attachment of molecules and receptors to EV surface are key ways to enhance delivery of targeted therapy .Passive loading of EVs can be simply performed by incubating exosomes with the cargo of interest. The differing concentration inside and outside the EVs will drive diffusion of hydrophobic cargos through EVs lipid-bilayer. Several chemotherapeutic drugs such as doxorubicin and paclitaxel have been loaded into exosomes by incubation ,134, demSaponification allowed up to 11-fold higher drug loading of large or hydrophilic compounds like porphyrins compared to passive methods . In thisAn additional loading technique is to apply sound energy with a sonicator to exosomes solution, which induces mechanical shear forces affecting exosome membrane integrity and enhancing the incorporation of the cargos present in the solution to vesicles . This prExtrusion is a process by which exosomes are loaded by membrane disruption when extruded through small polycarbonate porous membranes, allowing to incorporate cargos present in the solution to the vesicles . DespiteThis method consists of freezing and subsequently thawing a solution of vesicles with the cargo of interest. The mix is incubated at 37 \u00b0C, frozen rapidly at \u221280 \u00b0C and then thawed at room temperature, repeating at least three times. One inconvenience of this method, however, is aggregation of vesicles into large-sized particles ,138.Permeabilization of EV membrane by applying an electrical field is one of the most common techniques employed to enhance uptake of the cargo of choice. Disruption of the phospholipid bilayer of EVs allows the hydrophilic compounds to be diffused into exosomes such as small DNAs ,139, miRUse of chemical reagents for siRNA loading into exosomes has also been described. Wahlgren et al. employed liposome-based transfection reagent to introduce MAPK-1 siRNA into exosomes by incubating the mixture for 10 min at room temperature . LikewisAnother interesting approach is the generation of engineering cells to produce loaded exosomes with specific cargos. One approach is transfection of donor cells to overexpress a particular gene, allowing the generation of specific gene-loaded exosomes during their biogenesis. Jiang et al. investigate the therapeutic effect of TSG-6 modified MSC-derived exosomes on a mouse full-thickness wound model, providing evidence of a possible mechanism to prevent from scar formation . In addiIn addition to isolating, loading and/or modifying exosomes, selecting the exosome administration route and dose are also important parameters to consider to achieve clinical relevance in the future. In this respect, a key challenge is enhancing the biodistribution, stability and therapeutic effect of exosomes. In the following subsection we summarize some of the most frequently used exosome administration routes. Systemic injection is one of the most commonly administration routes used in exosome therapeutics ,106. AmeSeveral studies provide evidence that exosomal subcutaneous injections are effective in several pathologies ,158, butIntranasal injection provides a practical option for administration of therapeutic compounds to the brain. One of the first studies reported that curcumin-loaded EL-4-derived exosomes were efficiently delivered to microglia cells by intranasal administration, showing protection against lipopolysaccharide (LPS)-induced brain injury . In addiAlthough not commonly used as intravenous injection, intraperitoneal delivery of exosomes is another administration route option . InteresIn addition to the administration routes previously described, oral treatment offers a simple non-invasive route of exosome delivery. This method, like others summarized above, has been employed to administrate chemotherapeutic drugs such as curcumin, resulting in three- to five-fold higher levels of curcumin in several organs compared to free delivery of the compound . RegardiFrom our literature review, we found a paucity of data comparing exosome loading by these different methods. Thus, it will be a vibrant area of research to assess the optimum doses required in vivo. In addition, the efficacy and safety of cargo-exosome delivery should be characterized in association with other available methods for gene and drug delivery to discover significant breakthroughs as well as find innovative applications of this novel approach.The properties of exosomes alone are not sufficient to guarantee the specific delivery of drugs and the enrichment of drugs in disease tissues. Indeed, a wealth of literature supports that exosomes need targeting strategies to improve the therapeutic effect of drugs .Targeted delivery of exosomes with increased biodistribution and specificity can be assessed by transfection of parent cells. The fragment is transfected by fusing the ligand of interest to the coding sequence of exosomal signaling peptide, allowing the protein of interest to attach to the vesicle surface . The mosDirect functionalization of exosomes is another option for exosome surface modification. Functionalization strategies tested, include covalent and non-covalent chemical. Covalent modification includes copper-catalyzed azide-alkyne cycloaddition (CCAAC) click chemistry, the reaction of an alkyne and an azide chemical group to form triazole linkage . The proAlong with the surface modifications described, stable modification of the exosome surface by non-covalent alteration of the EV membrane has been also developed and is summarized below. Multivalent electrostatic interactions involve binding highly cationic species to negatively charged groups located in the exosome membrane . HoweverDevelopment of modified exosomes has an auspicious future in drug delivery research. Innovative targeting moieties should be designed to promote enhanced delivery to a specific cell type. Although researchers have made impressive progress in the modification of engineered exosomes, most of the above-mentioned techniques are based on the modification of exosome donor cells. These changes may affect the protein composition and function of exosomes. Therefore, it is necessary to explore new techniques for exosome-targeted modifications to minimize changes in their composition. Maintaining progress in these areas will bring breakthroughs in the field and transform exosomes from promising candidates into smart nanoscale therapeutics.Both in themselves and as vehicles of drug and gene delivery, exosomes are under active development as therapeutic agents ,186. ExoThe use of EVs as a cell-free therapeutic alternative offers several distinct advantages over parent stem cells. A major advantage is that EVs, depending on their source, may be less immunogenic than their parental cells, likely due to a lower abundance of transmembrane proteins such as MHC complexes . Unlike Immune-therapeutic exosomes include naturally occurring exosomes, exosomes secreted by modified cells, and exosomes loaded with exogenous cargos .Several studies have shown that EVs from various cell sources have a therapeutic effect through their intrinsic content , which iOne major role of MSCs is to suppress proliferation and function of cells in both innate and adaptive immunity responses . Thanks MSC-EVs can exert immunosuppressive effects on T cells , inhibitLoading therapeutic cargos into exosomes is based on their biological processes, cargo is loaded into donor cells or overexpressed certain gene products which are then packaged into exosomes , route 2MSC is the most well-suited for mass production of exosomes for drug delivery . For exaExogenous cargos can be directly loaded into exosomes by different methods see , and theEncouraging recent studies show exosomes as potential therapeutic vehicle systems for small molecule drugs to reduce toxicity and enhance target disease tissue in autoimmune diseases. Currently, GCs are the most effective anti-inflammatory drugs available for SLE , but cliAnother potential drug to load in exosomes is curcumin, which has immunomodulatory potential in addition to anti-oxidant and anti-inflammatory effects. In both acute and chronic immune nephritis curcumin ameliorated kidney disease reducing proteinuria, glomerulonephritis, tubule-interstitial disease and renal infiltration . In addiDelivering specific functional nucleic acid drugs to targeted cells by exosomes can regulate gene expression and maintain the physiological balance of the cells ,231. LiuExosomes also transported protein cargos, including shock proteins, major histocompatibility complexes, cytoskeletal proteins and proteins related to signal communication and membrane transport ,236. In In summary, these works illustrate the undeniable potential of exosomes as a therapeutic drug carrier in autoimmune diseases . NeverthMost autoimmune disorders such as SLE are characterized by a chronic inflammatory state, the making reduction of inflammation essential in order to treat patient conditions. Drug therapies for controlling SLE have made significant progress in recent decades. However, dosing limits of therapeutic indications in current SLE therapeutics delay their optimal use, and SLE patients may tolerate severe negative side effects produced by non-specific organ toxicity from frequent and long-term treatment. Compared with conventional therapeutic approaches, exosome-based drug delivery system has many advantages, such as high stability, compatibility, low immunogenicity and toxicity, longer circulation time and targeting ability for specific drug delivery to inflamed tissue. Therefore, exosome-based nanocarriers potentially have a bright future as next-generation drug delivery vehicles. This review has provided the current state of the art in the field of exosome-based drug delivery in SLE, evaluating the beneficial effects of natural and engineered exosomes on immunosuppression, cartilage repair and renoprotection.However, the clinical manufacturing of exosome-based therapeutics requires that comply with good manufacturing practice (GMP) to ensure reproducibility, stability and purity . TherefoDespite the challenges facing the exosome-based drug delivery system, these endogenous vesicles have great potential in the biomedical field. Furthermore, developing artificial exosome mimetics with similar homing properties and low side effects to exosomes could be an interesting concept to pursue to obtain clinical scale production of nanocarriers. In summary, the application of nanotechnology and nanomedicine provides great potential for prevention and treatment of human autoimmune diseases, especially in SLE therapy."} +{"text": "Periodontal disease, one of the most prevalent human infectious diseases, is characterized by chronic inflammatory tissue destruction of the alveolar bone and the connective tissues supporting the tooth. Such pathological inflammation is caused by the host innate and adaptive immune response to a constellation of periodontal biofilm-related multiple microorganisms. The current knowledge of the pathophysiology of periodontal diseases is involves identifying keystone bacteria in the biofilm, the host immune response (including genetic factors), and environmental stressors. Based on these accumulated lines of evidence, novel diagnosis, preventive, or therapeutic remedies are under development or envisioned. Briefly, the sum of all these scientific endeavors will deepen the understanding of more detailed molecular mechanisms of periodontitis, thereby contributing to advances in therapeutic strategies for periodontitis. Thus, this Special Issue, entitled \u201cMolecular Mechanisms of Periodontal Disease\u201d, ten articles, three systematic, and six literature review papers are presented, highlighting recent advances in \u201cthe relation between keystone bacteria and immune system\u201d, \u201cthe role of bacterial/host metabolites\u201d, \u201cthe mechanism of osteoclastogenesis\u201d, \u201cthe genetics, epigenetics, and environmental factors\u201d, \u201cthe potential liquid biopsy for periodontitis\u201d, and \u201cthe novel strategy for the regenerative therapy\u201d.As described above, periodontitis is elicited by the host immune response against periodontal pathogenic microorganisms . Recent It is also well accepted that not only PAMPs but also several metabolites secreted from bacteria or host cells are associated with the progress of periodontal disease. For instance, short-chain fatty acids (SCFAs), including acetate, propionate, and butyrate, which are Gram-negative bacterial metabolites, are frequently detected in the periodontal pocket of periodontitis patients. Magrin et al. revealed that, among the SCFAs, butylate might exacerbate the gingival epithelium inflammatory response in their in vitro study and systReceptor activator of nuclear factor-kappa B ligand (RANKL) is a master cytokine of osteoclast differentiation and function. Although it is still controversial, osteoblasts or stromal cells are thought to be the cellular source of RANKL in the context of periodontitis. Thus, much scientific attention has been paid to the study of osteoblasts and osteoclasts. Kelder et al. reported that osteoblastic cells isolated from long bone showed more significant osteogenic activity than those of alveolar bone . As the The genetic background of the host immune system seems to be associated with the onset of periodontitis caused by pathogenic bacteria. A case\u2013control study by Borilova et al. indicated one possibility that the polymorphism IL-10-1087G/C (rs1800896) and specific IL-10 haplotypes may play a role in the development of periodontal disease . In addiGiven its intrinsic nature, gingival crevicular fluid (GCF) and saliva can be an attractive liquid biopsy for periodontitis. In combination with the advanced technology for proteomics analysis, Preian\u00f2 et al. proposed a future reliable diagnostic system using GCF . Han et To date, several regenerative therapies using stem cells, growth factors, and biomaterials have been tested with the aim of successful periodontal tissue regeneration. In any case, emerging evidence implies that both osteogenesis and angiogenesis are indispensable for successful tissue regeneration. This relationship between osteogenesis and angiogenesis is well documented in the review article by Diomede et al. . RecentlFor decades, scientific endeavors in periodontology have made several paradigm shifts, including keystone bacteria-mediated dysbiosis, immune regulatory T/B cells, RANKL-producing stromal cells, and genetic and epigenetic inflammatory responses. Moreover, following these discoveries, novel promising therapies are under development. In this Special Issue, nineteen cutting edge scientific papers in this study field are introduced. We are confident that scientists, including all the authors involved in this Special Issue, will continue to provide unexpected discoveries that will lead us to understand the molecular mechanisms of periodontal disease. This, in turn, will encourage us to develop novel promising preventive and therapeutic strategies for periodontitis patients."} +{"text": "In this work, we propose a new model describing the relationship between the analyte concentration and the instrument response in photoluminescence sensors excited with modulated light sources. The concentration is modeled as a polynomial function of the analytical signal corrected with an exponent, and therefore the model is referred to as a polynomial-exponent (PE) model. The proposed approach is motivated by the limitations of the classical models for describing the frequency response of the luminescence sensors excited with a modulated light source, and can be considered as an extension of the Stern\u2013Volmer model. We compare the calibration provided by the proposed PE-model with that provided by the classical Stern\u2013Volmer, Lehrer, and Demas models. Compared with the classical models, for a similar complexity , the PE-model improves the trade-off between the accuracy and the complexity. The utility of the proposed model is supported with experiments involving two oxygen-sensitive photoluminescence sensors in instruments based on sinusoidally modulated light sources, using four different analytical signals . M parameters) and some calibration data, the calibration procedure can be mathematically formulated as the multidimensional search of the M parameters minimizing the error between the calibration data and the model and positive values for the rest of parameters), and, when parameter x in the L or D models takes the value x parameter in the L and D models is equal to 1). The PE models provide a good robustness against instrumental bias, with the lowest relative error in experiments 1, 2, 5, and 6 and results very close to those of the best models in experiments 3 and 4.In order to study the effect of an instrumental bias (either in the phase-shift or in the modulation-factor) over the accuracy provided by the calibration model, a sensitivity analysis has been performed. The plots in The comparison of the results for the 3-parameter P2, L, and PE1 models reveals more robustness against instrumental bias and better performance with the PE1 model, suggesting that the inclusion of the exponent m and The comparison of the errors in m or m or According to the description of the photoluminescence provided in the literature ,5,6,7, tC as a p-th degree polynomial of N-sites Demas model requires a N-dimensional search (of the N Stern\u2013Volmer constants) for fitting the In this paper, we propose a new model for describing the relationship between the analytical signal and the analyte concentration in chemical sensors based on photoluminescence with modulated light sources. This model represents the concentration The proposed model (for degrees 1 and 2) has been compared with the classical calibration models in six experiments of phase-resolved photoluminescence. The proposed model provides the best trade-off between accuracy and complexity: the PE1 model (with only three parameters) provide an accuracy that is closer to the 4-parameter models than to the 3-parameter models. The PE2 model (with four parameters) provides better results than the 2-sites Demas model .K can be derived from the model parameters. Consistent results are observed when these parameters estimated for the proposed model are compared to those estimated with the classical calibration models.Even though the parameters of the proposed model do not have a direct physical interpretation, the response of the model at null concentration and the sensitivity According to the analysis included in this manuscript, for photoluminescence sensors close to the Stern\u2013Volmer model or excited at low modulation frequency, the Lehrer and Demas models are expected to be consistent. However, for higher modulation frequencies and particularly when the photoluminescence sensor significantly deviates from the first order differential equation, the Lehrer and Demas models exhibit inconsistencies and an alternative like the proposed polynomial-exponent model could be an interesting approach for calibrating the photoluminescence sensors."} +{"text": "The atmospheric concentrations of many non-methane hydrocarbons (NMHC) in the UK have been observed to fall over this period in broadly similar proportions. The relative contribution to emissions from solvents and industrial processes is estimated to have increased from approximately 35% in 1990 to approximately 63% in 2017. In 1992, UK national monitoring quantified 19 of the 20 most abundant individual anthropogenic VOCs emitted , but by 2017 monitoring captured only 13 of the top 20 emitted VOCs. Ethanol is now estimated to be the most important VOC emitted by mass followed by n-butane (52.4\u2009kt\u2009yr\u22121) and methanol (33.2\u2009kt\u2009yr\u22121). Alcohols have grown in significance representing approximately 10% of emissions in 1990 rising to approximately 30% in 2017. The increased role of solvent emissions should now be reflected in European monitoring strategies to verify total VOC emission reduction obligations in the National Emissions Ceiling Directive. Adding ethanol, methanol, formaldehyde, acetone, 2-butanone and 2-propanol to the existing NMHC measurements would provide full coverage of the 20 most significant VOCs emitted on an annual mass basis.Volatile organic compounds (VOCs) are a broad class of air pollutants which act as precursors to tropospheric ozone and secondary organic aerosols. Total UK emissions of anthropogenic VOCs peaked in 1990 at 2,840\u2009kt\u2009yrThis article is part of a discussion meeting issue \u2018Air quality, past present and future\u2019. More ref asthma leads tof asthma or changf asthma . There if asthma . A conseNMVOCs comprise all organic compounds except methane which at 293.15\u2009K show a vapour pressure of at least 0.01\u2009kPa (i.e. 10\u2009Pa) or which show a comparable volatility under the given application conditions.The term VOC, used frequently and interchangeably with the longer abbreviation non-methane VOCs (NMVOCs), is a catch-all term for any organic compound found as a gas in the atmosphere. In urban air, NMVOCs can encompass many thousands of different organic compounds including non-methane hydrocarbons (NMHCs), oxygenated, nitrated and halogenated species . Since tA slightly different definition of VOCs is given within the 2004/42/EC Paints Directive , which r2 to C14 are thought of as VOCs. Longer-chain hydrocarbons (>nC14) may fall outside of the definition, as may more highly functionalized organic compounds such as organic acids or organic peroxides. The majority of persistent organic pollutants (POPs) have lower vapour pressures than this definition, although some two and three ring polycyclic aromatic hydrocarbons such as naphthalene are considered as VOCs as well as POPs. The EC Air Quality Directive 2008/50/EC [\u2018volatile organic compounds\u2019 (VOC) shall mean organic compounds from anthropogenic and biogenic sources, other than methane, that are capable of producing photochemical oxidants by reactions with nitrogen oxides in the presence of sunlightThe practical realization of both definitions is that most simple NMHCs with a carbon number falling within the range CThe reporting of emissions as set out in treaties such as NECD and CLRTAP, in principle, takes into account all species that fall within the relevant technical definition. In the case of the UK, this is done through the evaluation of hundreds of different sources where statistical data must be collated for the purposes of reporting a single annual VOC emission total. The majority of countries report totals of emissions from various different sectors. The UK is unusual in then mapping the sectoral emissions onto nearly 700 different species.Pragmatic decisions are obviously needed with respect to those VOCs that might be measured as part of any verification or regulatory activities. Measurements may be needed to inform directly on attainment of specific health-related VOC concentration targets , or those VOCs that may act as an independent check that estimated emissions changes are being reflected in the ambient atmosphere.Since all VOC analytical methods have boundaries on specificity and sensitivity, a very limited range of VOCs are typically monitored in ambient air, compared with those actually emitted. Ideally, the species that are routinely monitored in air should reflect those that are indicative of the most significant emissions, and those that cause the greatest harm to health. Across Europe routine monitoring of VOCs has followed guidelines in Annex X of the Directive on Cleaner Air for Europe 2008/50/EC. This recommends the measurement of 31 individual VOCs, all NMHCs, ranging in molecular weight from ethane, through 1,3,5-trimethylbenzene. From these recommendations have followed a range of commercial monitoring instruments based around thermal desorption and the development of traceable standards for calibration to mole and kilogram . The cho(a)1 Both the NECD and CLRTAP set emissions ceilings with milestone targets for particular dates. For example, the NECD sets a ceiling for 2020 that requires a 32% reduction in total UK emissions relative to 2005 levels, excluding agricultural sources. This equates to 724\u2009kt in 2020 based on the latest NAEI estimates for 2005. The NECD then requires a 39% reduction in emissions relative to 2005 levels by 2030, equivalent to 649\u2009kt.The National Atmospheric Emissions Inventory (NAEI) estimates UK VOC emissions from anthropogenic sources following methods in the EMEP/EEA Emissions Inventory Guidebook for submThe NECD and CLRTAP define those VOC sources to be included and excluded from the national inventory (e.g. emissions of NMVOCs from biogenic sources are not included) and the technical definition (see earlier). The Guidebook provides estimation methodologies and default emission factors for each source category, although countries can use country-specific emission factors where these are deemed relevant. Key requirements for inventory reporting are Transparency, Completeness, Consistency, Comparability and Accuracy, and in this respect, it is important to provide a national inventory with time-series consistency going back to at least 1990 and forward to 2030.The NAEI uses a combination of emission factors from the EMEP/EEA Guidebook and emission rates provided by industry and regulators in the UK. The emission factors represent either (i) the total mass of all individual VOCs when added together or (ii) a metric defined as total hydrocarbons (THCs), a non-speciated mass of VOCs, used, for example, in the case of tailpipe road transport emissions. The speciation of the total emissions into individual VOCs is undertaken separately. Where THC factors are used, the methane emissions calculated separately are subtracted out.2 including many consumer products.The NAEI uses three basic approaches for estimating total VOC emissions\u2014top down, point source and industry-reported. In the first case, an emission factor approach (top-down) combined with relevant activity statistics is used for many combustion sources including crude oil refineries, other industrial sites and in residential buildings. Factors are also used for transport sources, as well as for some processes in the food and drink industry and for some uses of solvents,A point-source approach (bottom-up) can be used where the sum of emissions is estimated/measured and reported by process operators for each emitting site within a sector. The UK estimate is generated simply by summing the emissions reported for all of the sites within a given sector. This approach is used for refinery processes, chemicals industry, oil and gas production and certain types of solvent use in industry such as for printing of flexible packaging, and coating of road vehicles. Key sources for data on point-source emissions are the regulators in England, Scotland, Wales and Northern Ireland who maintain inventories for the processes they regulate, and the BEIS Environmental & Emissions Monitoring System (EEMS) for the offshore oil and gas sector.ad hoc basis and usually cover a limited number of years, and thus, a time-series may need to be generated by splicing together various datasets, possibly from different data providers. Therefore, there is a risk that a time-series might not be fully consistent and estimates for different years may be subject to different levels of uncertainty. However, industry estimates are essential for the VOC inventory: they provide data for sources where the lack of public domain data means that emission factor or point-source approaches cannot be used. Further details of the methods used in the NAEI are provided in the UK's national inventory report submitted annually to the UNECE and NECD. The latest version of the inventory is for years up to 2017 (the 2017 NAEI) as submitted in early 2019 .In the case of solvent use, the third approach is used for UK emission estimates taking data provided directly by industry. However, this may come with little additional information on emission factors or activity data. Data are often supplied on an (b)The largest contribution to VOC emissions in 2017 was from solvents. Overall emissions have decreased for this sector, but by significantly smaller amounts than road transport and fugitive emissions. There have also been relatively smaller reductions in emissions from industrial processes such that this sector contributed 15% of total emissions in 2017; these emissions now mostly come from the food and drink industry which have been increasing since 1990.The period since around 2000 has seen a substantial re-ordering of the relative contributions from each of the major source sectors. \u22121 by 2030 based on current projections. Meeting those targets is likely to require an increased policy and regulatory focus on those VOCs from the solvent and industrial usage sectors, and so the next section examines which VOC species are significant contributors.Overall, a small exceedance of the 649\u2009kt NECD target for 2030 is predicted for the UK unless there are further actions to reduce emissions. The size of additional reduction needed to meet the 2030 target is estimated to be approximately 30\u2009kt\u2009yr(c)There is no statutory requirement to report national inventories of individual VOC species to the NECD or CLRTAP, but a comprehensive speciated inventory is very valuable for other purposes. Given the different chemical reactivities of each component, a speciated inventory is essential for atmospheric models of ozone and SOA formation. It is also necessary for the interpretation of ambient measurements of VOCs and how such information can be used to verify inventory trends.13 aromatics\u2019.The UK is one of the few countries to have developed a comprehensive speciated inventory for VOC emissions, and this is widely used in atmospheric modelling. The speciated inventory was first developed in the mid-1990s, but the lack of significant new data sources means that development essentially finished in the early 2000s when the methodology was published . The NAEThe information used in the development of the species profiles came from various sources, for example industry trade associations in the 1990s provided some speciated estimates including the Solvent Industry Association, the British Coatings Federation and the British Aerosol Manufacturers Association. The solvent industry also helped with speciation of white spirit and other hydrocarbon mixtures. The Environment Agency's Pollution Inventory and similar inventories compiled by other UK regulators include some details of speciation, although the amount of detail is now much less than was the case in the early 1990s. Some analysis was undertaken of fugitive emissions at petrol stations, and species profiles were also provided by the refinery sector. The profiles for vehicle exhaust emissions were taken from the EMEP/EEA Emissions Inventory Guidebook. An important source for other sectors was the US EPA SPECIATE database.As much of the data used to develop the speciation profiles were gathered during a short period in the late 1990s and early 2000s, and since more recent data are not available, it is assumed that the species emitted by a particular source are the same in all years. While this is likely to be true for sources such as bread baking or gasoline distribution, it is probably not true of technologically evolving sources such as industrial coating processes, chemicals manufacture or the formulation of consumer and household products.There are approximately 360 different individual VOC source sectors or processes included within the NAEI, and each of these has a representative chemical speciation associated with it, so it is possible to interrogate the NAEI for annual amounts and therefore emission trends of individual VOCs. Although in total the NAEI contains data on nearly 700 VOCs, a much smaller subset of approximately 40 VOCs represents typically approximately 70% of the total mass of emissions. The 10 most significant individual VOCs in terms of mass emitted represented 45.3% of UK national emissions based on 2017 estimates. Ethanol was the most important VOC comprising 16.8% of all emissions. The recent trends in the 10 highest VOCs by mass emission are shown in n-butane, for example, is currently the second most abundant VOC in the UK inventory; in 1990, n-butane was emitted overwhelmingly from gasoline extraction and fugitive distribution losses (139.8\u2009kt\u2009yr\u22121). However, by 2017, the largest anthropogenic source of n-butane in the inventory was from its use as an aerosol propellant (25.5\u2009kt\u2009yr\u22121) with the gasoline/fugitive losses having been reduced to 23.3\u2009kt\u2009yr\u22121.Even for simple alkanes there have been some notable changes in the major contributing sources. (d)a shows the estimated emission trends of 12 VOC functional group types, plus a further \u2018other\u2019 category for all other minor functionalized VOCs that do not fall within these 12 classes. Figure\u00a04b shows the same data but expressed as a group contribution to the annual emissions in percentage terms for each year. The most striking feature is the increased significance of alcohols generally, with significant contributions to 2017 emissions made by 1-propanol, 2-propanol and 1-butanol as well from methanol and ethanol highlighted in the previous figure.Figure\u00a04a shows the trends in total VOC emissions for the UK (by mass) and the normalized POCP for each year derived from the top 40 emitted VOCs in that year using Derwent et al. [x and all other relevant photochemical parameters are held constant.Understanding the exact location- and condition-specific impacts of a change in VOC speciation on the tropospheric ozone can only be done fully using explicit modelling and is beyond what we report here. It is possible, however, to evaluate how a change in VOC speciation may impact on the ozone-forming potential of national emissions in the atmosphere by considering the photochemical ozone creation potential (POCP) ,20 of thFigure\u00a05t et al. POCP valOver the 1990\u20132017 period, the normalized POCP of the UK VOC mixture declined slightly, by approximately 4%. A complex set of changes lie behind this; the largest change in any single VOC is the reduction in ethane emissions (a low POCP compound) and the growth in ethanol (a species of intermediate POCP reactivity). However, there are also concurrent reductions in a range of other high POCP alkenes and aromatic compounds that offset the growth in ethanol. The net change is to an average emission mixture that is very slightly lower in POCP per unit national emissions in 2017 than the mixture being emitted in 1990, but likely too small a change to materially affect ozone formation rates.(e)3 Since 2001, there have been four continuous GC-FID systems measuring NMHCs in the UK and of these the monitoring station at Marylebone Road in Central London has the most complete data record. Although Marylebone Road is formally a roadside location, it does experience the full range of urban emissions given its central position within the city. Other publications have reported data from the UK up to around 2008, for example Von Schneidemesser et al. [Continuous measurements have been made of a range of non-methane hydrocarbons in the UK Automated Hydrocarbon Network since 1992. The shape of the network has changed over the years, although the methodology has always been based around thermal desorption\u2014GC-FID. Further details can be found on the UK-AIR online resource.r et al. . Here, wi- and n-butane, ethene, benzene, ethyne and propene. This is potentially rationalized as arising from initial falls in concentrations through the reduction of emissions of these species from their road transport source, but that other urban sources also exist that have not declined and that now dominate ambient concentrations.Ambient concentrations measured at the roadside cannot be expected to directly reflect inventory changes, since roadside monitoring sites of this kind are skewed to detecting changes in the locally dominant road transport source. However, it is still instructive to examine the general scale of reductions seen and the degree of agreement between observation and inventory. Making direct like-for-like trend comparisons between observations and inventories is generally not appropriate, given multiple contributing sources that define the ambient concentrations of any given VOC. Urban monitoring stations can be sensitive to a particular subset of sources along with reflecting broader patterns of long lifetime VOCs which are influenced by the regional transport. However, 1,3-butadiene is one VOC where some direct comparisons can potentially be made. The lifetime of butadiene during daytime with respect to hydroxyl radical oxidation is very short, around 30\u2009min, and so its measured concentration essentially reflects local sources only. The only major urban source of 1,3-butadiene is thought to be road transport tailpipe emissions. In (f)Previous sections have shown how at a national level, current emissions in the UK are now significantly influenced by VOCs released from solvent use, both industrial and domestic. As further reductions in emissions are required across Europe, and most reductions from the fossil fuel and transport sectors have likely already been achieved, reducing solvent emissions appear the most feasible route to meet future NECD objectives. Specific policies and interventions that might achieve these aims are beyond this paper, but there will likely be an overarching requirement for ambient observations to continue to provide some external verification that changes in emissions as determined by inventory reporting processes have occurred. This situation creates a measurement challenge since the solvent sector is heavily influenced by oxygenated compounds, rather than NMHCs which current online monitoring infrastructure is generally best configured to detect . There aThe current speciation of VOCs from the class of \u2018solvents and related products' in the NAEI is likely imperfect due to lack of up to date speciation information from manufacturers, but it provides some guide to the key species that are released. An alternative way of visualizing the impacts of changing emission speciation on monitoring strategies is to consider the most significant VOCs emitted each year and then whether there is coverage in terms of ambient air measurements. We evaluate this by taking a time slice at 5-year intervals from the VOC speciation in the NAEI and rank order the top 40 VOCs emitted in each year.13). Over time, however, the observational coverage has declined, to 2017 when only 13 of the top 20 VOCs were routinely monitored, including omission of two of the top three most significant VOCs emitted by mass.VOCs in It is also instructive to note those VOCs which are not as significant as they once were. Ethyne is one of the most challenging VOCs to measure using thermal desorption techniques due to its very low boiling point, and this can be a defining factor for much of the analytical instrumentation for analysis . By 2017, ethyne had fallen to 82nd when ranked by mass of emissions from a position in 1990 of 21st. An argument can also be made that the various isomers of butenes and pentenes that were once important as contributors to both total mass emissions and to reactivity and ozone creation potential in the 1990s have been reduced so significantly that the effort expended on their measurement in networks may not be particularly productive.(g)et al. [Since there is no standardized international requirement to report speciated VOCs as part of emissions control treaties, it is difficult to make direct comparisons between the UK-specific conclusions we report here and other global locations. We make some attempt to reality-check whether the influence of oxygenated VOCs is as great as suggested by UK inventories by examining the most significant VOCs detected in ambient air in the UK and in other countries, using data collected from short periods of research observations and process studies. Ambient data are, of course, not directly comparable to emission inventories at a national scale, since large point sources may not be detected at a given measurement location and there may be important seasonal factors not captured in short-term measurements that are reflected in annual averages. Many oxygenated VOCs are also produced through atmospheric oxidation, so they may be elevated in ambient air not solely because of direct anthropogenic emission, but also as a by-product of the degradation of other VOCs. This is particularly important for acetaldehyde and formaldehyde which are common degradation products, including also from natural emissions such as isoprene. We exclude those from this analysis. From the recent literature, we note the work of MacDonald et al. which shWe note that since research observations exist for many oxygenated VOCs, it demonstrates that there are no insurmountable technical obstacles to improving routine measurement coverage . This pa2.Inventory estimates of total VOC emissions in the UK show significant decreases over the past 30 years from a peak around 1990, which is mirrored in ambient measurements of some NMHCs. The most significant sources in the 1990s were NMHCs related to fossil fuel usage, in particular from natural gas extraction and distribution, gasoline tailpipe emissions and fugitive fuel losses. Policies and regulations across multiple sectors to reduce emissions have been very effective, particularly from road transport, which now contributes only a small amount (approx. 4%) to total UK VOC emissions. There has been little change in overall emissions from industrial and domestic solvent usage, and this has resulted in a growth of this sector as a proportion of national emissions. Along with this change has been a change in both the relative and absolute amounts of individual VOCs emitted. Ethanol is now the most abundant VOC emitted in the UK and overall, short-chain alcohols are the most important functional group, measured by mass. The shift in functional groups has not, however, had an appreciable impact on overall average POCP (a 4% decline) since the growth in ethanol has been balanced by losses in reactivity in other species, notably alkenes and mono-aromatics.If future changes in emissions are to be independently tested against external atmospheric monitoring, then a revised analytical strategy is needed, both in terms of species quantified and locations of the measurement themselves. The current online VOC networks used across Europe predominantly focus on the measurement of NMHCs, with only rather limited coverage of certain functionalized VOCs using off-line methods, for example to measure aldehydes. This results in many of the major oxygenated VOCs emissions going undetected and skewed in geography towards VOCs emitted from transport sources. Without some change to this position, it will not be possible to evaluate how successful future policies and technical interventions have been in reducing solvent emissions, noting that reductions in VOCs are needed in many countries to meet NECD and CLRTAP obligations for 2030. The addition of ethanol, methanol, formaldehyde, acetone, 2-butanone and 2-propanol to the existing suite of NMHC measurements would provide for a full observational coverage of the 20 most significant VOCs emitted on an annual mass basis. It may be possible for a change in future observational strategy to be brought closer to cost-neutral in operational terms by ceasing observations of certain analytically challenging non-methane hydrocarbons, such as acetylene, butenes and pentenes, which have seen their emissions fall very significantly and that are now often close to or below instrumental detection limits in UK ambient air."} +{"text": "Olfactory dysfunctions, including hyposmia and anosmia, affect ~100 million people around the world and the underlying causes are not fully understood. Degeneration of olfactory sensory neurons and incapacity of globose basal cells to generate olfactory sensory neurons are found in elder people and patients with smell disorders. Thus, olfactory stem cell may function as a promising tool to replace inactivated globose basal cells and to generate sensory neurons.Methods: We established clonal expansion of cells from the murine olfactory epithelium as well as colony growth from human olfactory mucosa using Matrigel-based three-dimensional system. These colonies were characterized by immunostaining against olfactory epithelium cellular markers and by calcium imaging of responses to odors. Chemical addition was optimized to promote Lgr5 expression, colony growth and sensory neuron generation, tested by quantitative PCR and immunostaining against progenitor and neuronal markers. The differential transcriptomes in multiple signaling pathways between colonies under different base media and chemical cocktails were determined by RNA-Seq.Results: In defined culture media, we found that VPA and CHIR99021 induced the highest Lgr5 expression level, while LY411575 resulted in the most abundant yield of OMP+ mature sensory neurons in murine colonies. Different base culture media with drug cocktails led to apparent morphological alteration from filled to cystic appearance, accompanied with massive transcriptional changes in multiple signaling pathways. Generation of sensory neurons in human colonies was affected through TGF-\u03b2 signaling, while Lgr5 expression and cell proliferation was regulated by VPA.Conclusion: Our findings suggest that targeting expansion of olfactory epithelium/mucosa colonies in vitro potentially results in discovery of new source to cell replacement-based therapy against smell loss. The National Institute on Deafness and Other Communication Disorders (NIDCD) estimates that ~1.4% of the United States population experience chronic olfactory dysfunction and smell loss + cells generate multiple cell lineages in the OE after methimazole-caused injury Two subtypes of basal cells, horizontal basal cells (HBCs) and GBCs, are responsible for OE regeneration in vitro as cloned neurospheres from rat Olfactory stem cells have been established + cells sorted from the murine OE in vitro using a Matrigel-based three-dimensional system. Colonies derived from adult Lgr5+ stem cells have been established in vitro, including cells from the tongue + cells were regulated under different growth conditions + olfactory sensory neurons in OE colonies. Morphological change from filled to cystic colony was observed in different base media with cocktail treatments, which was associated with differential Lgr5 expression and transcriptional alteration in multiple signaling pathways. Furthermore, we also established colonies cultured from human olfactory mucosa, and colony proliferation and neuronal differentiation were regulated by defined chemical stimulations. Thus, this study put forward a new source for cell-based transplantation therapy against smell loss.In this study, we cultured Lgr5Genetically targeted heterozygous Lgr5-EGFP-IRES-CreERT2 mice , Rosa26-floxed STOP-TdTomato mice and OMP-Cre mice were purchased from the Jackson Laboratory. Both male and female mice were used in this study and the data were grouped together because no sex difference was evident. The procedures of animal handling and tissue harvesting were approved by the Shanghai Medical Experimental Animal Administrative Committee (Permit Number: 2009-0082).All reagents were purchased from Sigma Aldrich unless specified. Chemicals were prepared as stock solutions and diluted freshly before using. The final concentration of each chemical was: 50 ng/mL bFGF , 50 ng/mL EGF , 50 ng/mL IGF-1 , 3 \u03bcM CHIR99021 (#SML1046), 1 mM VPA , 100 \u03bcg/mL pVc , 2 \u03bcM 616452 , 10 mM Nicotinamide (#N0636), 12.5 mM N-Acetyl-L-cysteine (#A9165), 10 \u03bcM SB431542 (#S4317), 5 \u03bcM LY411575 (#SML0506), 10 \u03bcM SB202190 (#S7067), 100 ng/mL SHH and 500 nM A83-01 (#SML0788).+ cells, the OEs from Lgr5-EGFP-IRES-CreERT2 mice at 3-month-old age were minced into small pieces using scissors in Tyrode's solution , then treated with 0.25% Trypsin-EDTA and DNase I for 20-30 min at 37 \u00b0C. Cell suspension was collected by centrifugation at 200 g. Cell pellet was suspended in HBSS buffer using a fire-polished glass pipette. Single-cell suspension was filtered using 70-\u03bcm nylon mesh to remove large aggregates, followed by further filtering with 35-\u03bcm nylon mesh . Cells were sorted using BD FACS Aria III cell sorter (BD Biosciences), according to the green fluorescent EGFP signal . Cells were sorted into a low-attached 1.5-mL microcentrifuge tube containing 0.5 mL colony growth medium described below.To obtain Lgr5+ cells from the OE were cultured according to previously reported protocol with some modifications in vitro culture and passaged every 7-10 days by using 0.25% Trypsin-EDTA to make single cell suspension. Approximately 5000 cells were reseeded in each well of low-attached 24-well plate.Sorted Lgr5TM , 2 \u03bcM 616452, 500 nM A83-01 and/or 10 \u03bcM SB431542. Approximately 10000 single cells were seeded per well in low-attached 24-well plate. Colonies were visible on Day 7 and then were passaged every 10 days or based on colony growth status.The written consents were obtained from patients before experiment. The procedures of human tissue harvesting and handling were approved by the Ethics Committee of Eye, Ear, Nose & Throat Hospital, Fudan University (Permit Number: 2019081). Human olfactory mucosae were dissected from patients with olfactory neuroblastoma. When tumors were resected, connected olfactory mucosae were isolated from tumor tissues and kept in iced PBS. Blood cells were removed by incubation with Red Blood Cell Lysis Solution . Tissues were minced into small pieces and digested with 0.25% Trypsin-EDTA and 50 \u03bcg/mL DNase I. Cell pellets were collected in growth medium that was used in murine OE colony culture without addition of Matrigel and then filtered with 35-\u03bcm nylon mesh . Cell suspension was resuspended in growth medium supplemented with 100 \u03bcg/mL Primocin5 and 2.5\u00d7105 TU lentivirus, respectively (MOI = 50). Apparent red fluorescence was observed on Day 3 post infection and colonies were analyzed on Day 10.The shLgr5-mCherry lentivirus targeting the mouse Lgr5 and shCtrl-mCherry control lentivirus were prepared by GENECHEM . Viral infection in murine OE colonies was conducted when passaging. Approximately 1000 and 5000 single cells were seeded into each well of low-attached 96- and 24-well plates, infected with 0.5\u00d710+ cells in OE colonies derived from Lgr5-CreERT2/Rosa26-TdTomato mice, 4-Hydroxytamoxifen at final concentration of 400 ng/mL was added into cultures on Day 0 post passaging and kept for 7 days. Colonies were subjected to further analysis on Day 10. Chemicals including small molecules and growth factors were added on Day 0 post passaging. A scheme describing the lineage tracing protocol was shown as For lineage tracing of Lgr5TM-A medium containing N2 (1%), B27 (2%), GlutaMAX\u2122 Supplement, HEPES (10 mM) and Matrigel [3% (Vol/Vol)]. To induce neuronal differentiation, NeurobasalTM-A differentiation medium was added on Day 10 post in vitro culture. LY411575 and/or CHIR99021 was added on Day 11 and colonies were incubated with these chemicals for 7 days. The scheme for differentiation protocol was shown as Figure The differentiation medium was based on NeurobasalThe growth medium [known as NWR (Noggin/Wnt3a/R-Spondin 1)-based medium] was replaced by DMEM/F12 medium containing B27, N2, GlutaMAX\u2122 Supplement, HEPES and 3% Matrigel, supplemented with 50 ng/mL EGF, 50 ng/mL bFGF and 50 ng/mL IGF-1 (known as EFI-based medium). The EFI-based medium was added on Day 4 post colony passaging.The protocol was described previously TM blocking buffer containing 2% (Vol/Vol) donkey serum and 0.3% Triton X-100 at room temperature for 1 h. Primary antibody incubation was performed overnight at 4 \u00b0C. After washing three times with PBS, appropriate secondary antibodies were used to visualize staining. The nuclei were counterstained with DAPI . Slides were mounted in Vectashield Antifade Mounting Medium . The primary antibodies used in this study were as follows: chicken anti-OMP olf , rabbit anti-mOREG/MOR174-9 and rabbit anti-PGP9.5 . Fluorescent images were captured under a SP8/Leica confocal microscope with LAS AF Lite software. Contrast and brightness of images were set in an equal level when captured. All captured images were z optical sections.For the immunostaining on cultured colonies, the cryosection (prepared as described above) were washed by PBS, followed by incubation with SuperBlock2O. Reaction conditions included an initial denaturation at 95 \u00b0C for 1 min, followed by 40 cycles of 95 \u00b0C for 20 s, 60 \u00b0C for 20 s and 72 \u00b0C for 30 s. The relative expression level was calculated using the 2-\u0394\u0394Ct method. Primers sequences were as follows: mouse Lgr5: CCTGTGGCTAGATGACAATGCTCTC and AAGGCGTAGTCTGCTATGTGGTGTA. Human Lgr5: ACCTATCGTCCAACCTCC TGTCGTC and GCACAGCACTGGTAAGCATAAGGCA. Human OMP: AGTCTGT GTACCGCCTCAACTTCA and TCTATGGCATCCGAGTCCTCCTTG. Human Krt5: GAGATCGCCACTTACCGCAAGC and CATAGCCACTGCCACTGCCATAT C. Human GAPDH: GGAGCGAGATCCCTCCAAAAT and GGCTGTTGTCATACT TCTCATGG. Mouse GAPDH: TCAATGAAGGGGTCGTTGAT and CGTCCCGTAG ACAAAATGGT.Total RNA was extracted by using E.Z.N.A.\u00ae Total RNA Kit I according to the manufacturer's manual. The extracted RNA was immediately dissolved in RNase-free water, and purity and concentration were determined on Biophotometer . First-strand cDNA was synthesized using a PrimeScript\u2122 RT Master Mix . Primers used in this study were synthesized by Ruidibio . Quantitative real time PCR was performed on an Analytikjena Real-Time PCR System . The reaction mixtures included cDNA template, 0.2 mM primers, SYBR qPCR SuperMix and ddHProteins were extracted using a Total Protein Extraction Kit . Samples were prepared by boiling in 5\u00d7SDS Loading Buffer for 5 min. SDS-PAGE was used to separate total protein, and 3 \u03bcg sample was loaded into each lane. The proteins were transferred to PVDF membrane and the membrane was blocked in 5% non-fat milk for 1 h. The following primary antibodies were used to detect protein expression: Lgr5 and \u03b2-actin . After incubation in primary antibodies at 4 \u00b0C overnight, membrane was incubated with secondary HRP antibodies at room temperature for 1 h. Protein bands were visualized using ECL Substrate .http://www.geneontology.org/) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis (http://www.genome.jp/kegg/) were performed. All the sequence data was analyzed on I-Sanger (www.i-sanger.com).RNA-Seq analysis was conducted by Majorbio Corp. . Sequencing reads were mapped to the mouse genome using HISAT2. Transcriptomes from RNA-seq reads were reconstructed by StringTie. Expression differences were evaluated using DESeq2. Pearson's coefficient was calculated to determine the correlation among different groups. The clustering analysis of global gene expression pattern in different samples was carried out using K-means clustering algorithm by RSEM software. Gene Ontology and 0.04% (w/v) PluronicTM F-127 for 45 min at 37 \u00b0C. Excess dye was removed by washing with HBSS. Imaging was carried out using confocal microscope (OLYMPUS IX83-FV3000-OSR) with an excitation wavelength of 494 nm and an emission wavelength of 516 nm by OLYMPUS FV31S-SW software. Odorant solution was applied by bath application. Acquired sequences of images were converted to \u0394F/F0, where \u0394F was the real-time fluorescent intensity change relative to F0, and F0 was the averaged baseline fluorescence values before odorant onset. Colonies of interest were labeled with a fixed area of region of interest (ROI) using Image J software and fluorescence change over time in this ROI was calculated using GraphPad Prism software.OE colonies on Day 14 post + cells. Counting was carried out by someone who was blinded to the experimental conditions design to eliminate bias. Data were presented as mean \u00b1 SEM from at least three independent experiments. Band intensity in western blot analysis was measured by Image J software. Statistical difference between two groups was measured by unpaired t test, and among multiple groups was determined by one-way ANOVA or two-way ANOVA using GraphPad Prism 6.0 software.Cell counting from the confocal images was performed using Image J software. Ratio of positively stained cells in each colony was defined as the number of positively stained cells versus the number of DAPIin vitro, we cultured Lgr5-EGFP+ cells sorted from the OEs of Lgr5-EGFP-CreERT2 mice. On Day 10 after in vitro culture, typical sphere-like structures were observed, similar to cultures obtained from liver and taste Lgr5+ progenitor cells + cells in colonies expressed progenitor cell marker Sox2 . In colonies at Passage 3 and Passage 4, CHIR99021 treatment also led to increases in the ratio of Lgr5-EGFP+ cells (p < 0.05 at P3 and p < 0.001 at P4), while addition of VPA did not significantly increase the ratio of Lgr5-EGFP+ cells at Passage 3 (p > 0.1 at P3 and p < 0.05 at P4). This was also confirmed by the quantitative PCR, showing that CHIR99021 treatment enhanced Lgr5-mRNA level by 2.6 \u00b1 0.8 fold compared to NWR control . Besides, colonies were treated with other chemicals including IGF-1 (I) and bFGF (F) without Noggin, by 43 \u00b1 9% without Wnt3a, and by 48 \u00b1 4% without R-Spondin 1 . Thus, NWR were required to maintain Lgr5 expression in OE colonies. To further demonstrate the necessity of NWR, we measured Lgr5-mRNA level in the presence of EFI instead of NWR. EGF, bFGF and IGF-1 are three basic factors maintaining the growth of Lgr5+ cells from mammalian cochlea in vitrop < 0.01, n = 4). Besides, N-Acetyl cysteine and VIAFSNT cocktail increased Lgr5-mRNA level by 2.5 \u00b1 0.9 and 3.6 \u00b1 0.1 fold, respectively . However, compared to NWR-based medium with chemical supplements, culturing in EFI-based medium with corresponding single chemicals and VIAFSNT cocktail showed significant reduction in Lgr5-mRNA levels , suggesting that EFI-based medium was not optimal for maintaining Lgr5 expression in OE colonies. Thus, we concluded that supplementing NWR into culture medium was preferable to maintain Lgr5 expression in OE colonies.The culture medium maintaining OE colony growth contained Noggin (a bone morphogenetic proteins inhibitor p < 0.05) and 193 \u00b1 32% (p < 0.01), respectively, while other chemicals did not change colony number (p < 0.001). Immunostaining against Ki67 demonstrated that addition of CHIR99021 and 616452 significantly increased the ratio of Ki67+ cells by 175 \u00b1 37% (p < 0.001) and 82 \u00b1 22% (p < 0.01) compared to NWR control while VPA led to an increase by 22 \u00b1 2% compared to NWR control (p < 0.05), demonstrating that these two chemicals affected generation of progenies from Lgr5+ cells. However, treatment with CHIR99021, pVc or bFGF did not significantly alter the ratio of TdT+ cells-containing colonies . Notably, chemical cocktail -VPA changed the morphology most significantly, while other cocktails such as -IGF-1, -bFGF and -SB431542 also increased the ratio of cystic colonies compared to EFI control (p < 0.001). In the absence of pVc (-pVc), the ratio of cystic colonies was significantly decreased compared to that in VIAFSNT-treated colonies . When colonies were cultured in NWR-based medium, only N-Acetyl cysteine (Cys) enhanced the ratio of cystic colonies, while other single chemicals did not significantly induce morphological alteration (p < 0.001). By comparison, chemical cocktails such as VIAFSNT (p < 0.001), -VPA (p < 0.05), -IGF-1 (p < 0.01), -bFGF (p < 0.001) and -Nicotinamide (p < 0.001) significantly increased the ratio of cystic colonies in contrast to NWR control . Culturing in EFI-based medium increased the colony size by 47 \u00b1 6% compared to in NWR-based medium. Chemical treatments such as Cys, IAFSNT (-VPA) and VIAFSNT did not significantly alter the colony size in contrast to NWR control . This was further validated by western blot, demonstrating the Lgr5 protein level was significantly reduced in OE colonies infected with Lenti-shLgr5-2 compared to the level in colonies infected with Lenti-Ctrl . Lgr5 downregulation increased the ratio of cystic colonies by 1.8 \u00b1 0.4 fold . By contrast, there was no significant alteration in the ratio of Ki67+ cells between mCherry- Lenti-shCtrl- and Lenti-shLgr5-infected colonies . Thus, these data demonstrated that Lgr5 downregulation hampered proliferation in OE colonies.Considering that Lgr5s Figure A. The efin vitro culture expansion p < 0.001). In both cystic and filled colonies, we observed positive staining against neuronal markers PGP9.5, Tuj1 and OMP as well as proliferative progenitor markers Ki67 and Sox2 , while treatment with A83-01 and SB431542 significantly increased the ratio of PGP9.5+ cells in cystic colonies . However, the ratio of Tuj1+ neurons was not significantly changed in cystic colonies under chemical stimulations and 616452 (p < 0.05) significantly increased Lgr5- and OMP-mRNA levels and by 44 \u00b1 8% in the presence of VPA (p < 0.001), but was not significantly changed with treatment of IGF-1, pVc or bFGF (+ cells by 2.7 \u00b1 0.6 (p < 0.01) and 3.7 \u00b1 0.6 fold (p < 0.001) A-C, 8G. + colonies from the murine OE and found that CHIR99021 and VPA induced highest Lgr5-mRNA level in colonies cultured in NWR- and EFI-based media, respectively. LY411575 significantly enhanced sensory neuronal generation in murine OE colonies. Culturing with EFI-based medium led to cystic appearance of murine colonies and was accompanied by significant alteration in transcriptomes involved in multiple signaling pathways. In colonies from human olfactory mucosa, addition of VPA and 616452 was favorable to Lgr5 expression and cell proliferation, while A83-01/SB431542 promoted neuronal differentiation. Thus, this study optimized the culture condition of olfactory epithelium/mucosa colony culture, potentially providing a new source for cell-based transplantation therapy against smell loss.In this study, we explored chemicals facilitating the expansion of Lgr5+ cells from multiple tissues form 3D cultures in vitro with regulation of Wnt signaling and through HDAC inhibition. CHIR99021 and VPA enhanced self-renewal of Lgr5+ stem cells in mouse small intestinal colonies + supporting cells + cells in colonies derived from Lgr5+ cells in the murine OE while Lgr5-mRNA level was increased by CHIR99021 in NWR-based medium (Figure Lgr5+ embryonic liver cells-formed organoids represented different cellular subtypes + cells (Figures Cystic structure is a typical morphology in organoids Figures -6. These+ stem cells from other organs Human olfactory stem cells under 2D culture are promising in the treatment of degenerative disease In summary, this work established culture of murine and human colonies from nasal epithelium and olfactory mucosa under defined chemical treatments, potentially providing source to stem cell therapy against loss of smell caused by sensory neuronal degeneration.Supplementary figures.Click here for additional data file."} +{"text": "Trypanosoma rangeli is a protozoan that infects several domestic and wild mammals and shows significant distribution in Latin American countries. T. rangeli infection is similar to Chagas disease, both in diagnostic and prophylactic terms. Thus, the objective of this work was to review the diagnostic aspects and use of T. rangeli as an immunogen for Trypanosoma cruzi infection. Trypanosoma rangeli\" and diagnostic aspects of T. rangeli infection in humans and/or research on the possible vaccines developed using T. rangeli strains for T. cruzi infection. For this elaboration, Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines were adopted with descriptors derived from the Medical Subject Headings (MeSH) platform in the PubMed/MEDLINE and SciELO databases. The inclusion criteria were defined as original articles on \"T. rangeli for T. cruzi infection in vertebrates and the remaining 14 predominantly dealt with the diagnostic aspects of T. rangeli infection in humans.After applying the inclusion and exclusion criteria, 18 articles were procured, of which 4 addressed research on the possible vaccines developed using In this study, we formulated a compilation of the essential literature on this subject, emphasizing the need for more accurate and accessible techniques for the differential diagnosis of infections caused by both protozoa, and underscored several prospects in the search for a vaccine for Chagas disease. Trypanosoma rangeli,T. rangeli,Rhodnius prolixus, can propagate the protozoan while feeding on these vertebrate hosts. This can provoke intense immune response, producing high levels of antibodies in the infected speciesT. rangeli infection stands out as a differential diagnosis for Trypanosoma cruzi (etiologic agent of Chagas disease) infection due to overlapping geographical distribution of the causative microorganisms, allowing the existence of simple and/or associate infections in both invertebrate and vertebrate hosts,T. rangeli, when presented in blood cultures of vertebrate hosts, possesses an undulating membrane of length 34 \u03bcm from one end to its free flagellum (T. cruzi membrane is more pleated)T. rangeli makes it difficult to distinguish it from T. cruzi,,T. cruzi in mammals,The protozoan T. rangeli infection in human hosts has been estimated to be approximately 18 monthsT. cruzi antibodies are of great significance,T. cruzi have prophylactic potential and broad medical relevancy because Chagas disease is a parasitic infection with a high mortality rate in humans, notably in the Americas, showing an estimation of 10,000 deaths annually. Moreover, its main clinical manifestation, chronic Chagas cardiomyopathy (CCC), can be particularly lethal in affected individuals. Recent studies have indicated that this disease has affected approximately 7 million people,,The period of spontaneous resolution of T. cruzi and T. rangeli infections and (ii) investigate the capability of T. rangeli in developing a vaccine for Chagas disease.Thus, due to the geographical overlap and morphological and immunological similarity of these two protists, it can be inferred that there is a need for techniques that allow the correct diagnosis of suspected Chagas disease cases. Therefore, the objectives of this article were to (i) address the diagnostic differentiation of Trypanosoma rangeli\" identified in the \"Title\u201d, and a \"human infection\" filter was explicitly delimited on the PubMed/MEDLINE platforms. On the SciELO platform, the same descriptor and filters that were specific to the platform, \"Health Sciences\u201d, \"Tropical medicine\u201d, and \"Infectious Diseases\u201d, were used in Thematic Areas. We emphasize that although the search strategy included \"human infection\u201d, the articles with non-human animal models were also considered for further evaluation because some of these texts showed clear epidemiological, clinical, diagnostic, or prophylactic aspects of T. rangeli infection in humans.We used the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines for the preparation of this review, as shown in There was no elaboration of the chronological or language restrictions in either of the platforms used. Duplicate reviews and studies were removed after conferring the authors, titles, years, and publication journals. When queries arose, the particular publications were downloaded and evaluated. The articles were initially selected through the analysis of several \u2018Titles\u2019 and \u2018Abstracts\u2019, according to the data presented in in vitro, human, and murine models and showing implications for human studies, such as vaccine-related or clinical/epidemiological/prophylactic studies. After the assessment of the \u2018Titles\u2019 of the non-duplicated screened studies, the exclusion criteria were used to classify the included studies into (i) review articles; (ii) articles in which the clinical/epidemiological effects were not related to human populations; and (iii) articles that sought protein fragment analysis, molecular assays, or genetic sequencing of T. rangeli and were unrelated to the diagnostic aspects and use of this species as an immunogen for T. cruzi infection.The inclusion criteria included (i) original articles, case reports and series, and clinical trials and (ii) experimental studies using T. rangeli, for T. cruzi infection for a systematic review. After initial screening, all potentially relevant studies were downloaded in full text and evaluated for eligibility.We examined the \u2018Abstract\u2019 of each article and selected only those texts that highlighted the diagnostic implications in humans and/or comprised research on possible vaccines, developed using Trypanosoma rangeli\", applying the filters that limited the subject to \u201cHealth Sciences\u201d and \u201cHuman Studies\u201d (84 by PubMed and 38 by SciELO), and eliminating duplicates (1 duplication), 121 publications were submitted for the evaluation of \u2018Titles\u2019, leaving only 50 papers for further evaluation. The \u2018Abstracts\u2019 of these papers were read, and 23 articles were selected for the composition of this review. However, 5 manuscripts could not be included in the study because they could not be downloaded in full text. Thus, 18 articles were selected for the present analysis.Of the 122 articles retrieved by searching the descriptor \"T. rangeli and T. cruzi) in almost all texts (17 out of 18), (iii) a predominance of the type of descriptive experimental study in almost all texts (15 out of 18); research with human animal model (12 out of 18) and (iv) research in Latin American countries , belonging to the epidemiological distribution of Chagas disease, could be observed.T. cruzi infection by inoculating mice with T. rangeli strains,,,T. cruzi populations in the control group and the group immunized with T. Rangeli. Even though Pal\u00e1u et al. (2003)T. rangeli strains, respectively, both articles presented correlated results, underlining parasitemia reduction and increased survival rates of the immunized mice, compared to the control mice. Moreover, analysis of the histological preparations of the skeletal and cardiac muscles of the mice in the study by Basso et al. (2008)T. cruzi-infected vaccinated group, whereas many amastigote nests and severe inflammatory infiltrates were detected in the non-vaccinated group.Four articles identified the experiments proposed for the development of vaccines against T. cruzi infection in mice inoculated with T. rangeli strains. Marini et al. (2011)T. cruzi epitopes, similar to those of T. rangeli, in the vaccine as an elementary aspect for the early elimination of T. cruzi and the low production of histological lesions in vaccinated mice.Marini et al. (2011)19,20,21,22,23,24,25,26,27,28,29,30.31.32. Tanoura et al. (1999)T. rangeli parasitemia, which was characterized by the gradual reduction of the acute infection, in human and murine hosts, suggesting that T. rangeli trypomastigotes could survive in the host\u2019s blood for long periods without proliferating. Sousa et al. (2008)T. rangeli stocks from two chronic chagasic patients and confirmed the identification of T. rangeli by visualizing the parasite in laboratory samples. Parada et al. (2010)T. rangeli infection in a blood donor in Europe, concluding that the possibility of T. rangeli infection should also be considered in patients with a history of previous positive tests, even if antibody serology for Chagas disease was negative. Additionally, 11 of these articles showed greater emphasis on improving the techniques and strategies of diagnostic differentiation of T. rangeli and T. cruzi infections,,,,,,,,,,It could be observed that 14 articles included in the systematic review showed a clinical, diagnostic, and/or epidemiological focus. (1984)Schottelius et alT. rangeli with T. cruzi in laboratory tests, Anthony et al. (1979). (1987)T. rangeli-infected patients and showed the presence of similar antibodies in Chagas disease patients and serum of T. rangeli-infected patients taken for serological diagnoses.On the other hand, considering the possibility of cross-reactivity of . (1987)T. cruzi or T. rangeli infections in immunoassays. Of these patients, 4 were diagnosed with T. rangeli infection, 4 showed mixed infections, and 6 were infected with only T. cruzi. Ross et al. (1993)T. cruzi.Furthermore, Guhl et al. (1994)T. rangeli than against T. cruzi. Moreover, V\u00e1squez et al. (1997)T. cruzi and T. rangeli in a survey, demonstrating a greater serological immunoreactivity to T. rangeli preparations than to T. cruzi preparations in the studied population.O'Daly et al. (1996)T. rangeli. Avila et al. (1990)T. rangeli-infected patients (56% of the patients); however, antiC IgM antibodies were also present in patients with chronic chagasic disease (30%) and human visceral (57%) and cutaneous (20%) leishmaniasis.To establish a more specific form of laboratory diagnosis, other tests were also studied. Salda\u00f1a & Souza et al. (2010)T. rangeli species to identify intraspecific polymorphisms and target PCR diagnostic methods, outlining the several primers capable of categorically recognizing the genomic DNA of both species. This seemed to be an efficient method for the identification of these protozoa during the acute and chronic phases of infection in vertebrate hosts and vectors, respectivelyBotero et alT. rangeli analysis in the medical field, encompassing queries about zoonosis, and the formulation of a suitable vaccine for Chagas disease. Both Trypanosoma species not only share vectors and habitat, but also show strong similar antigenic responses,T. rangeli.The proposed area of study in this systematic review was focused on the significance of T. rangeli infection might elicit a humoral and/or cellular immune response and partial protection against subsequent T. cruzi infectionT. rangeli inoculation of domestic dogs in rural areas of Argentina demonstrated the induction of an important antibody response against T. cruzi over prolonged periods and a significant reduction in parasitemia in the inoculated dogs. This suggested that antibodies against T. rangeli might be involved from the inception of T. cruzi eliminationAnimal testing revealed that T. rangeli does not induce acute parasitemia in humans on its own, the understanding of its morphological forms of infection is advantageous to differentiate it from T. cruzi in vertebrate and invertebrate species,Though R. prolixus. The metacyclic trypomastigotes are inoculated when the arthropod feeds on the host\u2019s blood,,T. rangeli demonstrated pathogenicity in experiments with this vector,,T. rangeli in the hosts and evaluated the metacyclogenesis process in vitro and the terminus of metacyclic trypomastigotes after infection in mice. They also examined fibroblast cultures, establishing that the parasitemia process was characterized by a gradual reduction of the acute infection and the hemolytic trypomastigotes could survive in the blood for long periods without proliferating.The disease is transmitted to vertebrate hosts mainly through the bite of invertebrate reservoirs, such as T. cruzi infection due to possible cross-reactions.Regarding the etiopathological diagnosis of both protozoa in Chagas disease patients, strategies for differentiating these parasites are required. The data obtained indicated that although ELISA was sufficiently sensitive for sero-epidemiological studies, the serological cross-reactivity between the two species was a possibility that must be considered, even in the cases of double infection. Ross et al. (1993)T. rangeli infections in humans. T. rangeli strains inoculated in mice appeared to be favorable stimuli for innate immune response against the subsequent T. cruzi inoculations. We underlined a few notable aspects including, the significant reduction in parasitemia, high survival rates of the immunized mice compared to the control organisms, and reduction in the infectious and inflammatory processes of T. cruzi in the groups immunized with T. rangeli,Together, the referenced articles were the result of meticulous analyses of the pertinent studies related to It was also noted that PCR and minicircle amplification, followed by DNA probe hybridization and RFLP analysis, were viable alternatives for more specific diagnosisT. rangeli strains for the diagnosis and immunoprophylaxis of T. cruzi infection.Our research could be a preliminary study and catalyst for more in-depth analyses in this area. Additionally, we emphasize the need for future experimental investigations to clarify the possibility of using"} +{"text": "N-terminal part of ARHGAP18. We present strong evidence that PKN3-ARHGAP18 interaction is increased upon ARHGAP18 phosphorylation and that the phosphorylation of ARHGAP18 by PKN3 enhances its GAP domain activity and contributes to negative regulation of active RhoA. Taken together, we identified new set of potential PKN3 substrates and revealed a new negative feedback regulatory mechanism of Rho signaling mediated by PKN3-induced ARHGAP18 activation.Protein kinase N3 (PKN3) is a serine/threonine kinase implicated in tumor progression of multiple cancer types, however, its substrates and effector proteins still remain largely understudied. In the present work we aimed to identify novel PKN3 substrates in a phosphoproteomic screen using analog sensitive PKN3. Among the identified putative substrates we selected ARHGAP18, a protein from RhoGAP family, for validation of the screen and further study. We confirmed that PKN3 can phosphorylate ARHGAP18 in vitro and we also characterized the interaction of the two proteins, which is mediated via the PKN3 (protein kinase N3) is a serine/threonine kinase belonging to the PKN family of kinases that act downstream of small Rho GTPases. While the expression of PKN1 and PKN2 is ubiquitous in most of the adult tissues , osteoclSince PKN3 downstream signaling still remains largely understudied, we decided to perform a phosphoproteomic screen to identify new PKN3 substrates using a chemical genetic approach based on the mutation in gatekeeper residue of the kinase ,17. AmonARHGAP18\u2014also known as SENEX\u2014is a member of a RhoGAP protein family which plays a role in regulation of activity of small Rho GTPases. Interestingly, ARHGAP18 exhibits different specificity towards individual Rho GTPases. In endothelial cells, ARHGAP18 was shown to act preferentially on RhoC . HoweverIn our study, we found that PKN3 interacts with ARHGAP18 and we present strong evidence that PKN3 phosphorylation of ARHGAP18 leads to the activation of its GAP domain, resulting in a decrease of RhoA activity.p-nitrobenzyl mesylate) and immunoblotted with an antibody recognizing thiophosphate-ester (clone 51-8). As expected, both PKN3 WT and PKN3 AS were able to use ATP\u03b3S but N6-Bn ATP\u03b3S could only be used by PKN3 AS . This approach is based on mutation of the gatekeeper residue allowing the kinase to use synthetic ATP analogs with a bulky group in the N6 position, thus providing specificity to the screen . Therefo PKN3 AS b. The ph PKN3 AS c. The li PKN3 AS . We subjSince PKN3 is a kinase acting downstream of active Rho GTPases and was shown previously to interact with two RhoGAP proteins and phosphorylate them , we deciIn the phosphoproteomic screen we identified two phosphorylated residues in the sequence of ARHGAP18 \u2013 Ser156 and Thr158. Interestingly, we noticed that although phosphorylation of Thr154 did not appear in our phosphoproteomic results, the region surrounding this residue strongly resembles the PKN3 phosphorylation consensus motif , mainly C-terminal region following the GAP domain (\u0394C525) and subjected to co-immunoprecipitation with Flag-PKN3. We observed a significant increase in binding of the GFP-ARHGAP18 N200 with PKN3 when compared to full-length GFP-ARHGAP18 (PKN3 was shown to directly interact with two other Rho-GAP proteins from the Graf family via their SH3 domain . Therefo (\u0394C525) a. Co-imm (\u0394C525) b. This sARHGAP18 c. To furARHGAP18 d. To furARHGAP18 e. To finARHGAP18 f. Taken Two isoforms of ARHGAP18 have been described with the only difference between the two being, that isoform 2 (Iso2) is missing the first 45 amino acids . Since wTo assess whether the phosphorylation of ARHGAP18 could have some effect on its function we first focused on the levels of active RhoA. U2OS cells expressing individual variants of GFP-ARHGAP18 were subjected to RhoA-GTP pull-down assay using immobilized GST-Rhotekin. Interestingly, a significant decrease in the levels of active RhoA was observed in cells expressing GFP-ARHGAP18 TST-DDD when compared to WT suggesting that phosphorylation of ARHGAP18 leads to activation of its GAP domain a,b. To sAlthough PKN3 is an important effector kinase of small Rho GTPases and a key player in regulation of processes such as cytoskeleton organization , prolifeAmong the identified putative substrates of PKN3 we selected ARHGAP18 for further study and validation. In an in vitro kinase reaction using either truncated or full-length protein we confirmed that PKN3 is able to phosphorylate ARHGAP18 on Thr154, Ser156 and Thr158. Although there are 80 serine and threonine residues present in the 663 amino acids-long sequence of ARHGAP18, mutation in these three candidate sites alone was sufficient to substantially impair the phosphorylation of ARHGAP18 by PKN3, suggesting the candidate sites are phosphorylated in a highly specific-manner. However, since the decrease in phosphorylation was not complete, we expect there are another residues in the sequence of ARHGAP18 phosphorylated by PKN3. We next found that PKN3 is able to interact with ARHGAP18 and we mapped the interaction interface to the first 25 amino acids in the sequence of ARHGAP18. Importantly, this region is missing in ARHGAP18 Iso2 which is translated from an alternative downstream start codon . InteresNotably, PKN3 and ARHGAP18 share a lot of similar traits in the signaling of endothelial cells. After downregulation of either ARHGAP18 or PKN3 in endothelial cells, the cells lose the capacity of characteristic tube formation in both 2D and 3D environment ,10,28 anN-terminus is potentially sterically blocked and, therefore, inaccessible for interactions. ARHGAP18 phosphorylation induces structural change leading to activation of GAP domain and in case of ARHGAP18 Iso1 also to a great strengthening of ARHGAP18 and PKN3 interaction mediated by the release of its N-terminus, which becomes accessible for interaction with PKN3. Our data also suggest that phosphorylation of both ARHGAP18 isoforms could facilitate a negative feedback loop in regulation of signaling mediated by Rho GTPases. In case of ARHGAP18 Iso1 this feedback mechanism is further strengthened by formation of a ternary complex between ARHGAP18, PKN3 and Rho GTPases , MDA-MB-231 cells were obtained from Dr. Zadinov\u00e1 as described previously [All the cell lines were cultured in DMEM supplemented with 10% FBS and 10 \u00b5g/mL ciprofloxacin in humidified incubator with 5% COeviously . Unless \u00ae Site-Directed Mutagenesis Kit following the manufacturer\u2019s instructions with the corresponding primers (T639G F/ R) listed in Human Flag-PKN3 WT and KD (kinase dead) in pcDNA3, as well as StrepII-PKN3 in pcDNA3 were used previously . Analog-XhoI, SacI, BamHI and NcoI restriction sites. Synthesized sequence was cloned into pEGFP c1 vector using BglII and EcoRI sites. ARHGAP18 N200 constructs were created by PCR amplification with respective primers (ARHGAP18 N200 F/R), digested with EcoRI/BamHI and cloned into pEGFP c1 via EcoRI/BglII sites and into pGEX 2T bacterial expression vector via EcoRI/BamHI sites. All the mutants of ARHGAP18 WT or N200 in either pEGFP c1 or pGEX 2T were created by whole plasmid synthesis approach (WHOPS) with Pfu-X7 polymerase and subsequent DpnI treatment with the respective primers listed in the EcoRI/BamHI sites. All the created constructs were verified by sequencing. cDNA encoding human ARHGAP18 isoform 1 was newly synthesized using GeneArt Gene Synthesis and cultured to 0.8 OD595. IPTG was added to the final concentration of 0.4 mM and incubated overnight at room temperature. Proteins were purified from cleared lysates using Pierce\u00ae Glutathione Agarose .GST alone and GST-fused proteins ), were purified using BL21 (DE3) 6-benzyl-ATP\u03b3S . Afterwards, denaturation buffer , 10 mM TCEP , 100 mM NH4HCO3 , 2 mM EDTA ) was added to the samples to 6 M final concentration of Urea, incubated for 1 h at 55 \u00b0C and cooled to RT for 10 min. Samples were diluted using 50 mM NH4HCO3 to a 2 M final concentration of Urea and 1 M TCEP was added to final concentration of 10 mM. 50 \u00b5g of Trypsin was added to each sample and incubated overnight in 37 \u00b0C with gentle agitation. Samples were then acidified to the final concentration of 0.1% TFA and peptides were extracted using Oasis\u00ae PRiME HLB columns . Peptides washed with 0.1% TFA in water were eluted with 1 mL of 0.1% TFA in 50% Acetonitrile in water and concentrated to 50 \u00b5L using speed vacuum. For capture of thiophosphorylated peptides, 100 \u00b5L of UltraLink\u00ae Iodoacetyl beads per sample was washed with 200 mM HEPES pH 7.0 and blocked for 10 min in dark with 5 \u00b5L of 5 mg/mL BSA in 50% Acetonitrile 50% 20 mM Hepes pH 7.0. Samples adjusted to a final concentration of 20 mM HEPES pH 7.0 and 50% Acetonitrile were added to the beads and incubated in dark place overnight at RT. After incubation, the beads were washed in the following order with 1 mL of water, 5 M NaCl, 50% Acetonitrile, 5% Formic Acid and incubated in 10 mM DTT for 10 min. Finally, samples were eluted for 10 min in 1 mg/mL OXONE pH 3.5, desalted with ZipTip and analyzed by Thermo Orbitrap Fusion coupled with Thermo Ultimate 3000 HPLC. Raw data were analyzed using MaxQuant software with MaxLFQ algorithm and the MS/MS spectra were searched against Uniprot-SwissProt human database both in forward and reverse using Andromeda search engine. Search parameters were set to standard trypsin digestion with two missed cleavages, variable N-terminal carbamylation, variable methionine oxidation and variable serine/threonine phosphorylation with the maximum number of modifications per peptide set to 5. The identified peptides were filtered based on 1% FDR. For selection of putative PKN3 phosphorylation sites, all the phosphosites identified in the control samples were eliminated from the analysis, together with phosphopeptides with localization probability below the cutoff value 0.75.The screen was performed following the protocol published by Hertz and colleagues . BrieflyMDA-MB-231 cells were transfected using PEI . After 48 h, the cells were washed with ice-cold PBS, lysed using 1% Triton X-100 in TBS , 150 mM NaCl) and the lysates were cleared by centrifugation. Subsequently, 20 \u00b5L of Anti-Flag Affinity Gel was added to each lysate and rotated for 3 h in 4 \u00b0C. After the incubation, the beads were washed twice with ice-cold lysis buffer, once with TBS and resuspended in SDS-PAGE sample buffer. After the separation of the samples using gradient SDS polyacrylamide gels (6\u201315%), proteins were transferred to Amersham Protran 0.45 \u00b5m nitrocellulose membrane using Transblot Turbo in a buffer containing 300 mM Tris, 300 mM Glycine, 0.025% SDS and 20% EtOH. After the transfer, the membranes were stained in Ponceau S for total protein, washed in TBS and blocked with 4% BSA in TBS for 30 min in RT. The membranes were incubated with respective antibodies diluted in 1% BSA in TBST overnight in 4 \u00b0C. The antibody against thiophosphate ester was diluted 1:5000 in 5% milk in TBST. Secondary antibodies were diluted in 2% milk in TBST. The membranes were developed with either WesternBright ECL or SuperSignal\u2122 West Femto Maximum Sensitivity Substrate using Amersham Imager 600 .StrepII Tag antibody was purchased from Novus Biologicals. Anti-thiophosphate ester antibody and GFP antibody used for immunoblotting (#ab290) were purchased from Abcam . RhoA antibody was purchased from Cell Signaling Technology. GFP antibody used for immunoprecipitation was purchased from Invitrogen. Flag tag monoclonal antibody and GST antibody (#G7781) were purchased from Sigma.\u00ae Superflow\u00ae resin and eluted with 1.25\u00d7 Buffer E .Kinase assays were performed as described previously . BrieflyN-terminal fragments of ARHGAP18 were produced and purified as described above (see Protein expression and purification). For the kinase assays of full-length ARHGAP18 proteins, individual GFP-ARHGAP18 variants were immunoprecipitated from transiently transfected U2OS cells. Lysates were prepared with the lysis buffer containing 1% Triton X-100 in TBS as described above and proteins were precipitated using anti-GFP 3E6 antibody (Invitrogen) and Protein A Sepharose 4 Fast Flow . Immobilized proteins were eluted using 0.1 M Glycine pH 3.5 for 10 min in RT, corresponding volume of 1 M Tris pH 9.2 was added to adjust the pH to 7.5 and eluted proteins were used as a substrate in kinase reaction. As a positive control for testing of PKN3 AS, a peptide derived from GSK3 was used . GST-fus2 , 10 mM Glycerol-2-Phosphate and 5 mM DTT . N6-benzyl-ATP\u03b3S or ATP\u03b3S were used as indicated in the final concentration of 1 mM. Reactions were incubated for 45 min in 35 \u00b0C and stopped by adding 0.5 mM EDTA pH 8.0 to the final concentration of 20 mM. Thiophosphorylated proteins were then alkylated with 50 mM PNBM (p-Nitrobensyl mesylate) at RT for 2 h, subjected to SDS-PAGE and immunoblotted using anti-thiophosphate ester antibody . All the kinase reactions were carried out in the kinase buffer containing 30 mM Tris pH 7.5, 4 mM MgClg, 13 min) and the supernatants were equalized for total protein . Agarose-bound GST-Rhotekin (20 \u00b5g) was added to each lysate and rotated for 45 min in 4 \u00b0C. Afterwards, the beads were washed twice with ice-cold lysis buffer, once with TBS and resuspended in SDS-PAGE sample buffer. The samples were subjected to SDS-PAGE and immunoblotted with respective antibodies as described above. For the analysis of active RhoA, U2OS cells were transfected with individual variants of GFP-ARHGAP18 or GFP alone, starved for 3 h and then lysed in 1% Triton X-100 in TBS , 150 mM NaCl) with inhibitors of proteases and phosphatases after 5 min stimulation by DMEM with 10% FBS. The lysates were centrifuged and 1mM DTT supplied with inhibitors of proteases and phosphatases . The lysates were equalized for the total amount of GFP signal. Agarose-bound RhoA-CA (15 \u03bcg) was added to each lysate and rotated for 1 h at 4 \u00b0C. The beads were washed three times with lysis buffer and resuspended in SDS-PAGE sample buffer. The samples were subjected to SDS-PAGE and analyzed by immunoblotting as described above.The RhoA pull-down assay was performed using purified constitutively active form of RhoA as described in Garc\u00eda-Mata et al. . Cells t"} +{"text": "Absence seizures are associated with generalised synchronous 2.5\u20134 Hz spike-wave discharges causing brief and sudden alteration of awareness during childhood, which is known as childhood absence epilepsy (CAE). CAE is also associated with impaired learning, psychosocial challenges, and physical danger. Absence seizures arise from disturbances within the cortico-thalamocortical (CTC) network, including dysfunctional feed-forward inhibition (FFI); however, the precise mechanisms remain unclear. In epileptic stargazers, a genetic mouse model of CAE with chronic seizures, levels of \u03b3-aminobutyric acid (GABA), and expression of GABA receptors are altered within the CTC network, implicating altered GABAergic transmission in absence seizures. However, the expression of GABA synthesising enzymes (GAD65 and GAD67) and GABA transporters (GAT-1 and 3) have not yet been characterised within absence seizure models. We found a specific upregulation of GAD65 in the somatosensory cortex but not the thalamus of epileptic stargazer mice. No differences were detected in GAD67 and GAT-3 levels in the thalamus or somatosensory cortex. Then, we assessed if GAD65 upregulation also occurred in Gi-DREADD mice exhibiting acute absence seizures, but we found no change in the expression profiles of GAD65/67 or GAT-3. Thus, the upregulation of GAD65 in stargazers may be a compensatory mechanism in response to long-term dysfunctional FFI and chronic absence seizures. Childhood absence epilepsy (CAE) is one of the most prevalent paediatric epilepsies, accounting for 10\u201317% of all diagnosed epilepsy cases in school-aged children . AbsenceA lack of stargazin protein in the stargazer mouse model of absence epilepsy reduces the expression of \u03b1-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) at excitatory synapses onto feed-forward inhibitory interneurons of the CTC network i.e., those in the somatosensory cortex (SScortex) ,9,10,11 ARs are only increased in the VP region [AR upregulation in the stargazer VP thalamus occurs only after seizure onset at postnatal days (PN)17\u201318 [AR upregulation likely contributing to seizure maintenance rather than generation. In the SScortex, by contrast, GABA transmitter levels are increased [In the CTC network of epileptic stargazer mice, levels of GABA and its receptors (GABARs) show region-specific alterations. In the thalamus, GABA levels are significantly reduced in the ventral posterior (VP) nucleus but not P region ,18. HowePN)17\u201318 . This suncreased . Interesncreased . Thus, tGABA levels are controlled by the enzymes responsible for its production and transporters mediating its reuptake. Glutamate decarboxylases (GADs) synthesise GABA from glutamate and exist as one of two isoforms: GAD65 and GAD67. GAD65 is predominantly localised to axon terminals where it catalyses the on-demand GABA synthesis crucial for phasic inhibition . On the GABA levels are also regulated by the GABA transporters (GATs: GAT-1 and GAT-3) that mediate its reuptake from the extracellular space. GAT-1 is expressed in neurons and some astrocytic processes, where it regulates GABA levels during sustained neuronal activity. Conversely, GAT-3 is produced primarily in astrocytes and regulates GABA levels in extrasynaptic areas ,25.In the genetic rat model of absence epilepsy (GAERS), absence seizures are thought to be caused by excess tonic inhibition in the VP thalamus due to insufficient GAT-1-mediated reuptake of extracellular GABA. Increased tonic GABAergic inhibition is evident in the VP thalamus of GAERS after PN17 and is sustained up to PN30 when SWDs first appear . IncreasIn a recent study, GABA uptake assays were performed in primary cultures of astrocytes from the thalamus and cortex of GAERS. The uptake activity of each transporter was determined by selective pharmacological blockade showing functional alterations somewhat at odds with GAT expression changes. GAT-1 expression increased and GAT-3 expression decreased in GAERS thalamic astrocytes, which showed reduced GABA uptake by both GATs. Additionally, in GAERS cortical astrocytes, where GAT-3 mediated uptake was reduced, GAT-1 and GAT-3 were both upregulated . MoreoveCurrently, less is known about the role of GADs in epilepsy. GAD65 knockout animals are highly vulnerable to seizures ,31,32,33Clearly, there are many mechanisms that can impair normal GABAergic inhibition within the CTC network. Abnormalities in the expression of GADs and GATs may play a role in altering GABA levels and thus GABAergic inhibition within the CTC network, leading to absence seizures. However, to date, there have been no studies specifically analysing the relative expression levels of GADs and GATs within the CTC network of a genetic mouse model of absence epilepsy. Hence, in this study, we first aimed to investigate the expression profile of GADs and GATs in the stargazer mouse model of absence epilepsy, which is known to have altered GABA levels in the CTC network . In thisThe expression pattern of GADs and GATs in relation to PV+ interneurons of the SScortex and the GAD65 expression was uniform throughout the cortical layers appearing as small, punctate staining within the neuropil but not colocalised with PV+ cell bodies, which is consistent with its axon terminal restricted expression A. MagnifThe expression pattern of GAT-1 and GAT-3 was uniform across SScortex layers with both GATs showing dispersed punctate labelling throughout the SScortex neuropil A,B.In the RTN thalamus, GAD65 expression was restricted to punctate structures, likely the axon terminals of GABAergic interneurons. GAD65 staining did not colocalise with the cell bodies of PV+ interneurons, but magnified merged images showed GAD65-positive puncta surrounding PV soma A. In conThe expression of GAT-1 and GAT-3 was uniform throughout the RTN and VP thalamus A,B. ColoThe omission of primary antibodies eliminated immunolabeling in tissue sections A,B confiTogether, it can be concluded that the expression patterns of GADs and GATs in the SScortex and the thalamus are not observably different between epileptic stargazers and their non-epileptic counterparts and appear similar to published localisation profiles.Western blotting analysis was performed to analyse the relative levels of GADs and GATs in whole-tissue lysate of the SScortex and VP thalamus of epileptic stargazers and non-epileptic control littermates. The same antibodies with specificity verified by confocal immunofluorescence were used for semi-quantification of protein levels with Western blotting.n = 18; STG 1.24 \u00b1 0.11, n = 15; p = 0.024, n = 16; STG 1.217 \u00b1 0.11, n = 13, n = 12) compared to non-epileptic littermates , GAD67 , GAT-3 ). Unfortunately, GAT-1 was unable to be detected in Western blots using the original antibody or by antibodies from two different suppliers, despite using the highest recommended concentration. Hence, we were unable to analyse the relative levels of GAT-1 in stargazers compared to non-epileptic control littermates.The protein level of GAD65 was significantly increased in SScortex of epileptic stargazers by 24% above their non-epileptic littermates E,F. In t n = 17) A\u2013F .n = 12); vehicle-treated DREADD ; CNO-treated non-DREADD WT control animals ); ; vehicle treated DREADD ; CNO-treated non-DREADD WT control animals ); ; vehicle-treated DREADD ; CNO-treated non-DREADD WT control animals ).Expression levels of GAD65, GAD67, and GAT-3 in the SScortex were not significantly different between groups A\u2013F and GABA transporting proteins (GATs) was investigated. We found that GAD65 levels are significantly increased in the SScortex of epileptic stargazer mice compared to non-epileptic littermates, whereas levels of GAD67 and GAT-3 are unaltered. In contrast, none of these proteins are altered in the VP thalamus of epileptic stargazer mice. Acute administration of CNO into inhibitory Gi-DREADD animals, which silences FFI and is sufficient to generate absence-like seizures, did not change the SScortex expression levels of GAD65, GAD67, or GAT-3. This suggests that changes in GAD65 expression in the stargazer SScortex may occur in response to a sustained reduction of inhibitory activity during development.Western blotting showed significantly increased GAD65 levels in the SScortex of epileptic stargazer mice. This indicates the excess GABA seen in the SScortex of epileptic stargazers may be dIn contrast, GAD67, which is expressed throughout cells ,45,46,47Cre/Gi-DREADD animals [Proteins can be rapidly synthesised at synapses in response to stimulation levels . To asse animals . HoweverIn the thalamus, as in the cortex, GAD65 showed a uniform staining pattern across both RTN and VP regions, while GAD67 strongly colocalised with the PV+ soma in RTN but showed uniform labelling in VP. As RTN contains the inhibitory cell bodies that project to VP, GADs in the VP region are those expressed in the axon terminals. Thus, any alterations to GAD expression in this region may be indicative of changes in de novo protein synthesis at the axon terminals . HoweverARs are upregulated to increase postsynaptic sensitivity to GABA release. Further studies assessing the temporal changes in GABA and GAD65 levels across stargazer development will help clarify the drivers of altered GABA levels in different brain regions.As no changes in the expression of GADs or GAT-3 were found in VP thalamus, it is likely that these proteins are not responsible for the reduced VP thalamic GABA levels in stargazers . It is pAlternatively, excess GAT-1 in the VP thalamus, causing excess GABA uptake, may be responsible for lowered GABA levels. Unfortunately, this study was unable to quantify changes in GAT-1 levels via Western blotting. As studies have shown the involvement of GAT-1 in absence seizures ,48, its Cre/Gi-DREADD mice. Mice were bred and housed at the University of Otago animal facility at controlled room temperature (22\u201324 \u00b0C) with ad libitum access to food and water. All animal procedures were approved by the University of Otago Animal Ethics Committee under the AEC no. DET 32/17 and D94/16.Animals used in this study were stargazer mice and PVStargazer mice were obtained from Jackson Laboratories, USA. The breeding protocol for stargazer mice in this study was similar to our previously published work ,9,10. HeloxP-flanked STOP cassette designed to prevent transcription of the downstream HA-hM4Di-P2A-mCitrine coding region. The breeding protocol for PVCre/Gi-DREADD mice in this study was similar to our previously published work [PV-Cre knockin and hM4Di-flox mice were obtained from Jackson Laboratories, USA. PV-Cre knockin mice express Cre recombinase in PV+ interneurons without disrupting endogenous parvalbumin expression. hM4Di-flox mice have a hed work . We crosCre/Gi-DREADD mice or crossed with heterozygous hM4Di-flox males to generate litters with PVCre/Gi-DREADD and non-DREADD expressing wild-type (WT) control littermates [Cre/Gi-DREADD mice was reported in our previous published study [Homozygous female PV-Cre mice were crossed with either homozygous hM4Di-flox males to generate litters of PVtermates . Only feed study .Cre/Gi-DREADD mice colonies. They were mixed in DNA lysis buffer and proteinase K and digested overnight at 55 \u00b0C. The following day, samples were centrifuged and DNA extracted to process for PCR. Genotyping was performed using primers listed in Cre/Gi-DREADD mice, genotyping was performed separately to confirm Cre knockin and hM4Di-flox using Cre, hM4Di mutant, and wild-type primers . The PCR product was run in an agarose gel at 70\u201380 V for around two hours. Then, the gel was placed on a UV light source to view and photograph bands. Cre/Gi-DREADD and stargazer colonies.Genotyping was performed to verify the mouse genotypes. Ear notches were collected from the offspring of stargazer and PVn = 4; and non-epileptic controls n = 3) were deeply anesthetised with an intraperitoneal injection of 60 mg/kg of sodium pentobarbital. Then, transcardial perfusion was performed with 5% heparin in 0.1 M phosphate-buffered saline (PBS), which was followed by 4% paraformaldehyde (PFA) in 0.1 M Sorensen\u2019s phosphate buffer (PB). Brains were carefully extracted and post-fixed in 4% PFA overnight at 4 \u00b0C. The next day, brains were washed three times in 0.1 M PB followed by cryoprotection in increasing concentration of sucrose in PBS, i.e., 10% for 30 min, 20% for 30 min, and 30% at 4 \u00b0C until the brains sunk to the bottom of the glass container. Brains were sectioned into 30 \u00b5m coronal sections on a freezing cryostat , and sections were collected into 12-well plates containing PBS.Adult stargazer mice , 0.1% Bovine Serum Albumin (BSA), 0.1% Triton X-100 in PBS) for two hours at room temperature. Following blocking, sections were incubated with the appropriate primary antibodies diluted in PBS with 0.1% BSA and 0.3% Triton X-100 for 48 h at 4 \u00b0C. Primary and secondary antibodies used for immunofluorescence confocal microscopy are listed in Images were acquired using a Nikon A1+ inverted confocal laser scanning microscope. Channel configurations were set for the green channel (488 nm laser excitation) and red channel (568 nm laser excitation). During confocal imaging, the detector offset for each channel was kept at zero; the detector gain, laser power, scan speed, and image pixel size were optimised accordingly. Images were captured of the region of interest (ROI) in the SScortex and thalamus (RTN and VP) in sections from the brain.Animals were sacrificed by cervical dislocation. Brains were immediately extracted, snap-frozen on dry ice, and stored at \u221280 \u00b0C. The cerebellum was dissected from the rest of the brain and forebrain mounted in the cryostat chuck using embedding media ) . The temperature inside the cryostat chamber was always maintained at \u221210 \u00b0C. Then, 300 \u00b5m thick coronal sections were cut and thaw-mounted on glass slides. Slides were temporarily stored on dry ice to prevent degradation of the tissues. The primary SScortex and VP thalamus were identified using the Mouse Brain Atlas and punched out from sections with 1.0 mm biopsy punches under a dissecting microscope. Micro-punched tissues were collected into microtubes containing homogenisation buffer , 1% phenylmethylsulfonyl fluoride (PMSF), and 1% protease inhibitor . Samples were sonicated for five minutes, heated for two minutes at 100 \u00b0C, and then centrifuged at 13,000 rpm for five minutes at 4 \u00b0C. The supernatant from each microtube was transferred into a new microtube and stored at \u221280 \u00b0C until use.In the stargazer-based Western blotting experiments, 27 epileptic stargazers and 31 non-epileptic control animals from 20 litters were included. In the DREADD-based Western blotting experiments, 12 CNO-treated DREADD animals, 8 vehicle-treated DREADD animals, and 8 CNO-treated non-DREADD WT controls were used. In the DREADD experiment, animals were injected with CNO/vehicle (10 mg/kg) and sacrificed within 30 min after injection Timing of the sacrifices was based on results from our previous published work where siFirst, 1.5 mg of CNO was dissolved in 75 \u03bcL of dimethyl sulfoxide (DMSO). Then, the volume was adjusted to 1.5 mL by the addition of 0.9% sterile saline to prepare CNO of 1 mg/mL concentration. Vehicle was prepared by mixing DMSO with 0.9% sterile saline. CNO or vehicle was injected intraperitoneally at 10 mg/kg. For focal injections, 0.3 \u03bcL of CNO was infused into the specific brain region at a rate of 0.1 \u03bcL/min. It was delivered via a Hamilton microinjection syringe attached to polythene tubing and a 33-gauge internal cannula inserted into the previously implanted guide cannula.p < 0.05 .Statistical analyses were performed in GraphPad Prism 9.1.1. Data are presented as mean \u00b1 standard error of the mean (SEM). Statistical differences in the expression levels of protein in the Western blotting experiments were tested using the Mann\u2013Whitney unpaired rank test. Statistical significance was set at In the stargazer mouse model of genetic absence epilepsy, FFI neurons are less responsive to activation. The upregulation of GAD65 observed in this study may be induced as an attempt to compensate for a chronic lack of FFI, with acute silencing of FFI insufficient to trigger such a transcriptional response. Excess GAD65 would increase GABA synthesis at axon terminals, thus resulting in more GABA released per synaptic event, heightening SScortex GABA levels, as has been reported in stargazers . Excess"} +{"text": "Severe hypotonia during infancy is a hallmark feature of Prader Willi syndrome (PWS). Despite its transient expression, moto development is delayed and deficiencies in motor coordination are present at older ages, with no clear pathophysiological mechanism yet identified. The diverse motor coordination symptoms present in adult PWS patients could be, in part, the result of a common alteration(s) in basic motor control systems. We aimed to examine the motor system in PWS using functional MRI (fMRI) during motor challenge. Twenty-three adults with PWS and 22 matched healthy subjects participated in the study. fMRI testing involved three hand motor tasks of different complexity. Additional behavioral measurements of motor function were obtained by evaluating hand grip strength, functional mobility, and balance. Whole brain activation maps were compared between groups and correlated with behavioral measurements. Performance of the motor tasks in PWS engaged the neural elements typically involved in motor processing. While our data showed no group differences in the simplest task, increasing task demands evoked significantly weaker activation in patients in the cerebellum. Significant interaction between group and correlation pattern with measures of motor function were also observed. Our study provides novel insights into the neural substrates of motor control in PWS by demonstrating reduced cerebellar activation during movement coordination. Prader Willi syndrome (PWS) is a rare genetic disease that is due to the loss of expression of the paternal copy of chromosome 15q11-q13 . The disIn addition, alterations in the motor control in PWS entail an important part of the clinical expression that may be severe at birth. Neonatal hypotonia is a hallmark feature of PWS ,3 characNo clear pathophysiological mechanism has been yet identified accounting for the motor expression of the PWS. The extensive genetic defect in PWS is likely to interfere with brain development at multiple levels. However, the clinical characteristics of the symptoms and the evolution from a dominant hypotonia in infants to broad repercussions on the motor control in adults suggest that the motor symptoms could share, at least in part, a common discrete neural correlate rather than being a consequence of diffuse brain damage.Current imaging studies have identified anatomical alterations in both the cerebellum and the sensorimotor cortex e.g., ,18,19,20In the current study, we employed fMRI to investigate whether the brain systems involved in the motor control are dysfunctional in adults with PWS. Neural activity evoked in PWS patients during a series of motor tasks of different complexity was compared with the normal pattern identified in a reference group of healthy participants. A correlation analysis was also performed between brain activation and behavioral measures of motor performance including upper limb muscle strength, functional mobility and balance.We hypothesized that activity in the motor system would be abnormal in the adult with PWS, and that the cerebellum could be particularly affected, as a brain structure capable of influencing each aspect of motor control. Indeed, although there are other factors that may be associated with neonatal hypotonia, such as metabolic or muscle-related peripheral factors, which could also be present in PWS, neonatal hypotonia may perhaps express to some extent a severe cerebellar dysfunction partially compensated during development, but more subtly expressed in the form of motor coordination deficits in adult PWS patients. The anatomical alterations described in the cerebellum in adults may well indicate that the cerebellum does not develop normally despite partial functional compensations.The sample included 23 patients with genetically diagnosed PWS . Genetic testing showed 16 patients (70%) with interstitial deletion of the proximal long arm of chromosome 15, and 7 patients (30%) with maternal uniparental disomy or defect of the imprinting center. Patients under the age of 18 years or with non-stable medical conditions and those considered unable to follow MRI instructions were not eligible. Twenty-two healthy subjects matched by age and sex to the PWS group made up the control sample. The study was conducted according to the guidelines of the Declaration of Helsinki. The protocol was approved by the Institutional Ethics Committee of Institut Universitari Parc Taul\u00ed of Sabadell, Barcelona (ref. END-GH-2017). All participants or their parents/guardians provided written informed consent and all PWS patients who were not able to sign, gave drawn assent.Participants underwent selected behavioral tests of motor function in a single session prior to the MRI scanning day. The following tests were administered:Hand grip strength was measured as an indicator of general muscle strength using the Jamar hydraulic hand dynamometer . Participants were instructed to sit in a chair with their feet touching the ground, shoulder adducted and neutrally rotated, with the elbow bent to 90\u00b0, the arm against the trunk, and the wrist in a neutral position . Each paFunctional mobility and risk for falls was assessed by the Timed Up and Go (TUG) test . ParticiThe Berg balance scale (BBS) was usedParticipants were previously given instructions concerning fMRI testing procedures and the need to remain still during acquisition. Functional MRI testing involved the performance of three manual tasks of different motor complexity ,30: (1) A Philips Achieva 3.0 Tesla magnet , equipped with an eight-channel phased-array head coil and single-shot echo planar imaging (EPI) software was used for the fMRI assessment. The functional sequences consisted of gradient recalled acquisition in the steady state (time of repetition (TR), 2000 ms; time of echo (TE), 35 ms; pulse angle, 70\u00b0) within a field of view of 230 \u00d7 230 mm, with a 64 \u00d7 64-pixel matrix, and a slice thickness of 3.59 mm . A total of 34 interleaved slices were acquired to cover the whole brain. Three 3-min scans were acquired for each participant. Each functional time series consisted of 90 consecutive image volumes obtained during each of the 3-min assessments. The first four image volumes in each run were discarded to allow magnetization to reach equilibrium. All participants wore headphones and performed all three tasks.For each of the three fMRI tests, we used an identical 3AB block-design paradigm consisting of six 30-s blocks, totaling 3 min in duration, which corresponded to 3 blocks of 30 s of right hand movement alternating with 3 blocks of 30 s of left hand movement in the first task, and to 3 rest (baseline) blocks of 30 s alternating with 3 movement blocks of 30 s in the second and third tasks. The examiner visually controlled task performance and gave the commands \u201cnow move\u201d and \u201cnow pause\u201d at the beginning and end of the respective hand movement blocks. During the rest period, participants were asked to simply lie still and not think about hand movements.https://www.fil.ion.ucl.ac.uk/spm (accessed on 18 January 2021)) implemented in Matlab . Preprocessing involved motion correction, spatial normalization, and smoothing by means of a gaussian kernel of full-width half-maximum 8 mm. Data were normalized to the standard SPM-EPI template and resliced to 2 mm isotropic resolution in Montreal Neurological Institute (MNI) space. All image sequences were visually inspected for potential acquisition and normalization artifacts.Imaging data were processed using Statistical Parametric Mapping software time series were aligned to the first functional image (of each time series) in each participant using a least squares minimization and a 6-parameter (rigid body) spatial transformation. (ii) We included 6 motion-related regressors in the first-level (single-subject) analyses. (iii) Within-subject, censoring-based MRI signal artifact removal (scrubbing) was usedt test was used to compare mean differences between the PWS and the control group in terms of demographic characteristics and behavioral ratings. Pearson\u2019s chi-squared test was used to assess the relationships between categorial variables. Visual analyses and Shapiro-Wilk tests showed non-normal distributions in the scores of the TUG test and the BBS. Thus, the non-parametric Mann-Whitney test was used to study potential group differences in these variables. The statistical analyses on the behavioral data were conducted with the Statistical Package for the Social Sciences ).Student\u2019s t test designs were used to generate group activation maps. Two-sample t tests were used to compare brain activation between the PWS and the control group. In addition, whole-brain correlation analyses were performed to map the possible relationships between brain activation and maximal grip strength, time to complete the TUG test, and BBS scores separately and for each task.To obtain individual maps of brain activity evoked during motor challenge, a boxcar regressor was generated considering the three blocks of the baseline condition and the three blocks of the test condition, and applying a hemodynamic delay of 4 s. Contrast \u201cright hand < left hand\u201d and \u201cright hand > left hand\u201d images were estimated for each participant in the unimanual flexion-extension task. The contrasts \u201cbaseline < motion\u201d (activation) and \u201cbaseline > motion\u201d (deactivation) were estimated instead for both the bimanual anti-phase flexion-extension and the finger-to-thumb opposition tasks. Resulting first-level SPM contrast images for each subject were carried forward to group-level random-effects analyses. One-sample p < 0.005 were considered, which satisfied the family-wise error (FWE) rate correction of pFWE < 0.05, according to Monte-Carlo simulations.Thresholding criteria. In whole-brain analyses, clusters > 2.3 mL (290 voxels) at a height threshold of p < 0.001). The mean grip strength for each handle position for each group is shown in p < 0.001), and PWS patients\u2019 score in the BBS test was also significantly lower than that for controls . In the PWS group, there was a significant negative association between the scores in the TUG and BBS tests such that less time in the TUG test was associated with higher scores in the BBS. This association was not observed in the control group, probably due to a ceiling effect of the scores in the BBS test.Descriptive statistics for all the measures are presented in The \u201cacross-hand\u201d analysis of the first task consisted of the comparison of the right-versus-left hand activation. In both groups, repetitive opening and closing of the hand caused a localized significant increase in MR signal intensity in the contralateral Rolandic region that was strictly located in the primary motor and somatosensory cortices, anterior and posterior to the central sulcus respectively, with a peak maximum at the cortical representation of the hand, as well as in the contralateral putamen extending to the posterior insula, and cerebellum ipsilateral to the hand that was performing the task . No signThe analysis of the second task consisted of the comparison of brain activation during bimanual anti-phase flexion-extension movements with the resting condition. As opposed to the analysis of the first task, this analysis did not produce cancellation of shared activity and, therefore, it allowed further functional identification of non-primary cortex activations. The control group showed activation in the sensorimotor strip, premotor and supplementary motor areas, superior parietal cortex, central operculum, thalamus, putamen and pallidum in the basal ganglia, and cerebellum bilaterally. The pattern of activation in the PWS group was qualitatively similar, albeit less extensive particularly in subcortical regions, and included bilateral changes occurring in sensorimotor, premotor and supplementary motor cortices, and cerebellum . Figure In the finger-opposition task, neural activation was conspicuous in both groups in the contralateral (left) sensorimotor hand area and ipsilateral cerebellum, thus resembling the activation pattern observed in the simpler unimanual task, but similarly to the bimanual task, such a fine finger movements also recruited ipsilateral and contralateral areas surrounding the primary sensorimotor cortex, thalamus, basal ganglia extending to the insula and opercula bilaterally, and contralateral cerebellum. In the control group, activation was also found in ipsilateral and, to a lesser extent, contralateral prefrontal cortex and temporo-occipital cortex bilaterally. In the PWS group, the maximal activation was found in the posterior bank of the central sulcus in the region that contains the sensorimotor representation of the hand; however, subcortical activation was restricted to the contralateral thalamus, pallidum and putamen, and ipsilateral cerebellum . For then = 16) and UPD (n = 7). Groups did not differ in age, sex or IQ. No significant differences in brain activation were observed between the two groups.In order to study the effect of the genetic subtype, PWS patients were split in 2 groups: DEL positive correlation between left premotor cortex activation and BBS scores. Finally, PWS showed stronger (than controls) negative correlation between hand grip strength and activation in the left thalamus and cerebellum during the fingers-opposition task. Brain areas where the magnitude of activation during the fMRI tasks was associated with changes in behavioral measurements are summarized in In the present study, we aimed to investigate the brain\u2019s motor system in adult PWS patients using fMRI with motor activation paradigms of different complexity. Selected behavioral testing additionally served to assess general aspects of motor function . We found that patients displayed altered motor performance and reduced brain activation during motor tasks compared with healthy controls. In line with our hypothesis, statistically significant between-group differences involved the cerebellum. Significant interactions between group and correlation pattern with measures of motor function were also observed.In the simple task, consisting in the repetitive flexion-extension of one hand, control subjects demonstrated the expected finding of robust activations in primary sensorimotor cortex and thalamus contralateral with respect to the hand participating in the movement, and ipsilateral cerebellum. With increasing task complexity, that is, repetitive bimanual anti-phase flexion-extension movements and fingers-to-thumb opposition movements, a greater number of secondary regions and bilateral subcortical areas were additionally recruited, which is in accordance with the results of previous fMRI and positron emission tomography (PET) studies using equivalent tasks e.g., ,34,35,36The cerebellum has an important role in the control of fine voluntary movements, motor coordination and motor timing ,41,42. RPWS patients typically score below the normal range on standardized motor performance tests . ConsistNeuroanatomical and imaging studies have described cerebellar abnormalities ,50,51 anAccurate control of head motion effects may be a strength of our studies; however, fMRI activation findings can be complicated by the level of intellectual disability present in PWS individuals which may interfere with task complexity and represent a potential confound related to cognitive demand. Nevertheless, we selected only relatively complex tasks, and subjects were trained on and they practiced the tasks until the authors verified that all participants performed the tasks correctly so that they were considered well learned before the fMRI scan, which does not support that PWS\u2019 cerebellar dysfunction is simply the result of their IQ. Future studies using a control group of subjects with a similar intellectual disabilities may help elucidating this issue. The study is also limited by its cross-sectional nature with a relatively small sample size, which might prevent us from drawing general conclusions.In summary, our study provides novel insights into the neural substrates of motor control in PWS by demonstrating reduced cerebellar activation during movement coordination. Prominent hypotonia in infants and subsequent motor problems that continue into childhood are compatible with a poor cerebellar function as a key pathophysiological factor. Our findings reinforce the role of the cerebellar dysfunction in PWS and indicate that it may be also expressed in adulthood. Investigation of the neural mechanisms underlying motor development in PWS may help improving the effectiveness of current therapies and eventually designing new interventions for the motor difficulties that characterize these patients."} +{"text": "A despeito da comprovada efetividade do cardioversor-desfibrilador implant\u00e1vel (CDI), as terapias de choque deflagradas pelo dispositivo podem causar n\u00edveis elevados de ansiedade e depress\u00e3o, provocando efeitos delet\u00e9rios na qualidade de vida.Florida Shock Anxiety Scale (FSAS) para a l\u00edngua portuguesa falada no Brasil. Realizar a tradu\u00e7\u00e3o, adapta\u00e7\u00e3o transcultural e valida\u00e7\u00e3o do instrumento Nesse estudo psicom\u00e9trico, a validade de construto foi realizada pela an\u00e1lise fatorial explorat\u00f3ria (AFE) e confirmat\u00f3ria (AFC) e pela Teoria de Resposta ao Item. Os \u00edndices de ajustamento da AFC foram: Robust Mean-Scaled Chi Square/df NNFI, CFI (Comparative Fit Index), GFI (Goodness Fit Index), AGFI (Adjusted Goodness Fit Index), RMSEA (Root Mean Square Error of Approximation) e RMSR . A confiabilidade foi verificada pelo Alfa de Cronbach, \u00d4mega de McDonald e Greatest Lower Bound. As an\u00e1lises foram realizadas no SPSS 23.0 e Factor 10.8.01, com n\u00edvel de signific\u00e2ncia de 5%. A vers\u00e3o final em portugu\u00eas do FSAS foi administrada em 151 portadores de CDI, com idade m\u00e9dia de 55,7 \u00b1 14,1 anos e predom\u00ednio do sexo masculino. A an\u00e1lise paralela indicou que o FSAS \u00e9 unidimensional, com vari\u00e2ncia explicada de 64,4%. As correla\u00e7\u00f5es variaram de 0,31 a 0,77; as cargas fatoriais de 0,67 a 0,86 e as comunalidades de 0,46 a 0,74. Os \u00edndices de ajustamento da AFC estabeleceram-se acima dos limites de qualidade. Encontramos evid\u00eancias satisfat\u00f3rias de confiabilidade da escala FSAS. O instrumento FSAS-Br apresentou evid\u00eancias consistentes de validade e confiabilidade, podendo, portanto, ser utilizado em portadores de CDI do Brasil. :764-772) Terapias de baixa energia, conhecidas como estimula\u00e7\u00e3o antitaquicardia ou sobre-estimula\u00e7\u00e3o, n\u00e3o provocam dor. Terapias de alta energia s\u00e3o choques de at\u00e9 40 joules que, apesar de causar grande desconforto, usualmente, s\u00e3o aplicados ap\u00f3s o paciente ter perdido a consci\u00eancia, uma vez que s\u00e3o disparados cerca de 15 segundos ap\u00f3s o in\u00edcio da fibrila\u00e7\u00e3o ventricular ou de uma taquiarritmia ventricular r\u00e1pida. Em situa\u00e7\u00f5es pouco desej\u00e1veis, como \u00e9 o caso das arritmias resistentes \u00e0 sobre-estimula\u00e7\u00e3o ou nas tempestades el\u00e9tricas, choques de alta energia podem ocorrer em pacientes acordados.8 Por outro lado, quando o CDI \u00e9 utilizado para preven\u00e7\u00e3o secund\u00e1ria, a incid\u00eancia de terapias de choque pode variar de 35 a 53%, no primeiro ano ap\u00f3s o implante.8 A despeito do elevado n\u00edvel de sofistica\u00e7\u00e3o tecnol\u00f3gica dos CDIs, lamentavelmente, existe o risco de o paciente receber um choque inapropriado em decorr\u00eancia da discrimina\u00e7\u00e3o err\u00f4nea entre taquiarritmias supraventriculares e ventriculares. Nessas ocasi\u00f5es, a sensa\u00e7\u00e3o relatada \u00e9 de uma experi\u00eancia dolorosa e angustiante.14Estima-se que a chance de portadores de CDI necessitar de um choque apropriado para preven\u00e7\u00e3o prim\u00e1ria da morte s\u00fabita card\u00edaca varia de 2 a 15% ao ano.14Por outro lado, tem sido reportado que o implante do CDI pode conferir aos pacientes uma grande sensa\u00e7\u00e3o de seguran\u00e7a, considerando-se a capacidade do dispositivo para interromper epis\u00f3dios inesperados de taquiarritmias ventriculares potencialmente letais.14Portadores de CDI convivem com a expectativa de que, a qualquer momento, o dispositivo ir\u00e1 aplicar terapias de choque para interromper arritmias ventriculares decorrentes de sua doen\u00e7a card\u00edaca. Desse modo, apesar de reconhecerem os benef\u00edcios do tratamento, alguns pacientes podem apresentar quadros de ansiedade, depress\u00e3o, dist\u00farbios do humor, transtorno de estresse p\u00f3s-traum\u00e1tico, bem como medo do n\u00e3o funcionamento do dispositivo em momentos cruciais.16 Em pouco tempo, o instrumento Florida Shock Anxiety Scale (FSAS) alcan\u00e7ou grande aceita\u00e7\u00e3o internacional, tendo sido traduzido e validado em diferentes culturas , com resultados consistentes.Diante da preocupa\u00e7\u00e3o com os efeitos delet\u00e9rios do CDI na adapta\u00e7\u00e3o psicossocial, foi desenvolvido um instrumento especificamente destinado \u00e0 avalia\u00e7\u00e3o do n\u00edvel de ansiedade relacionado \u00e0 presen\u00e7a do CDI e aos choques aplicados pelo dispositivo, para uso na pr\u00e1tica cl\u00ednica e no cen\u00e1rio da pesquisa cient\u00edfica.A finalidade do presente estudo foi avaliar as propriedades psicom\u00e9tricas da vers\u00e3o brasileira do instrumento FSAS para portadores de CDI.Trata-se de um estudo transversal com duas fases: (1) Processo de tradu\u00e7\u00e3o e adapta\u00e7\u00e3o transcultural do instrumento FSAS; (2) Valida\u00e7\u00e3o de construto do instrumento FSAS.Esse estudo foi realizado em um hospital cardiol\u00f3gico de alta complexidade, tendo sido aprovado pelo Comit\u00ea de \u00c9tica em Pesquisa da Institui\u00e7\u00e3o. Todos os participantes assinaram o termo de consentimento livre e esclarecido (TCLE).16O instrumento FSAS foi desenvolvido em 2006 nos Estados Unidos para avaliar o n\u00edvel de ansiedade relacionado \u00e0 presen\u00e7a do CDI e aos choques aplicados pelo dispositivo. O instrumento consiste em 10 itens, com cinco op\u00e7\u00f5es de resposta , correspondendo a uma escala do tipo Likert de 5 pontos.As perguntas do question\u00e1rio est\u00e3o relacionadas ao medo e \u00e0 ansiedade dos pacientes em raz\u00e3o da expectativa de o dispositivo aplicar terapias de choque, bem como a mudan\u00e7as comportamentais adotadas para evitar a ocorr\u00eancia de terapias de choque, a partir da perspectiva do paciente.18 O instrumento pode ser autoadministrado ou aplicado na forma de entrevista.O escore total do instrumento FSAS \u00e9 determinado pela soma de todos os itens, podendo atingir pontua\u00e7\u00e3o m\u00e1xima de 50 pontos. Quanto maior a pontua\u00e7\u00e3o, maior o n\u00edvel de ansiedade, sendo que os itens que atingem tr\u00eas ou mais pontos devem ser considerados os aspectos mais cr\u00edticos.25 O comit\u00ea de especialistas foi constitu\u00eddo por profissionais da \u00e1rea da estimula\u00e7\u00e3o card\u00edaca artificial e enfermeiros.O processo de adapta\u00e7\u00e3o transcultural do instrumento FSAS seguiu as recomenda\u00e7\u00f5es internacionais e constou de cinco etapas: (1) Tradu\u00e7\u00e3o por dois colaboradores independentes; (2) Consolida\u00e7\u00e3o das tradu\u00e7\u00f5es pelo comit\u00ea de especialistas; (3) Retrotradu\u00e7\u00e3o; (4) Harmoniza\u00e7\u00e3o das vers\u00f5es pelo comit\u00ea de especialistas; (5) Pr\u00e9-teste com a popula\u00e7\u00e3o alvo; (6) Revis\u00e3o do pr\u00e9-teste e versionamento final.1 e T2) foram consolidadas em uma \u00fanica vers\u00e3o (T1-2), ap\u00f3s discuss\u00e3o com o comit\u00ea de especialistas. A partir dessa vers\u00e3o traduzida, foi realizada a etapa da retrotradu\u00e7\u00e3o (RT1 e RT2) por dois professores bil\u00edngues que n\u00e3o tinham tido contato pr\u00e9vio com o instrumento original. A finalidade dessa etapa foi aferir a consist\u00eancia sem\u00e2ntica e idiom\u00e1tica das tradu\u00e7\u00f5es realizadas na primeira etapa. Finalmente, foi realizada uma nova reuni\u00e3o com o comit\u00ea de especialistas para revisar todo o processo de adapta\u00e7\u00e3o transcultural e realizar a harmoniza\u00e7\u00e3o das vers\u00f5es, obtendo-se, assim, a vers\u00e3o pr\u00e9-final do instrumento. adultos, com idades entre 18 a 80 anos, de ambos os sexos e com qualquer n\u00edvel de escolaridade; (2) portadores de CDI h\u00e1 mais de seis meses; (3) capacidade de compreender e responder \u00e0s perguntas do question\u00e1rio utilizado no estudo; (4) concord\u00e2ncia em participar do estudo, com assinatura do TCLE. N\u00e3o foram inclu\u00eddos no estudo os indiv\u00edduos que apresentaram ao menos uma das seguintes situa\u00e7\u00f5es: (1) indica\u00e7\u00e3o de transplante card\u00edaco; (2) gesta\u00e7\u00e3o em curso; (3) neoplasias malignas.Os pacientes foram selecionados, de forma consecutiva, durante o atendimento ambulatorial ou mediante visitas realizadas \u00e0 Unidade de Interna\u00e7\u00e3o da nossa institui\u00e7\u00e3o. Os indiv\u00edduos que preencheram os crit\u00e9rios de elegibilidade foram convidados a responder o question\u00e1rio FSAS. Nesse mesmo momento, era realizado o levantamento do hist\u00f3rico cl\u00ednico e obtidas informa\u00e7\u00f5es sobre o implante do CDI, terapias de choque pr\u00e9vias e uso de medicamentos.tablets, utilizando-se formul\u00e1rios eletr\u00f4nicos desenvolvidos no sistema Research Electronic Data Capture (REDCap)26 hospedado no servidor da nossa institui\u00e7\u00e3o.Todos os dados foram coletados diretamente em 25O dimensionamento do n\u00famero de participantes nos estudos psicom\u00e9tricos usualmente \u00e9 feito sobre o n\u00famero de itens dos instrumentos avaliados. Alguns estudos demonstraram que uma rela\u00e7\u00e3o de 20:1 ou superior, ou seja, 20 participantes para cada item do instrumento, seria o ideal. Contudo, rela\u00e7\u00f5es de 10:1 j\u00e1 permitem an\u00e1lises adequadas. Desse modo, foi estabelecido o m\u00ednimo amostral de 150 pacientes.Foi realizada uma an\u00e1lise descritiva minuciosa utilizando medidas de tend\u00eancia central . O teste de Kolmogorov-Smirnov (KS) foi utilizado para a testagem isolada da normalidade de cada item do question\u00e1rio, enquanto que o Teste de Mardia foi empregado para verificar a normalidade multivariada.Todas as an\u00e1lises foram realizadas nos softwares SPSS 23.0 e Factor 10.8.01, adotando-se o n\u00edvel de signific\u00e2ncia de 5%.Neste estudo, foram realizadas as an\u00e1lises fatoriais explorat\u00f3ria (AFEs) e confirmat\u00f3ria (AFC) para verificar a dimensionalidade do instrumento FSAS em sua vers\u00e3o na l\u00edngua portuguesa.Optimal Implementation of Parallel Analysis (PA), com minimum rank factor analysis (MRFA), que minimiza a vari\u00e2ncia comum dos res\u00edduos.28A robustez do teste foi determinada a partir da associa\u00e7\u00e3o de um bootstrap com uma extrapola\u00e7\u00e3o amostral para 5000. A extra\u00e7\u00e3o dos fatores foi feita inicialmente com RULS (Robust Unweighted Least Squares), que reduz os res\u00edduos das matrizes.29A testagem da dimensionalidade foi realizada pela An\u00e1lise Paralela Robusta (APR), por meio da Foi utilizado o \u00edndice de discrimina\u00e7\u00e3o do item (a), que mede a for\u00e7a de associa\u00e7\u00e3o entre o item e a vari\u00e1vel latente e tem interpreta\u00e7\u00e3o similar \u00e0s cargas fatoriais da AFE.Para a adequa\u00e7\u00e3o dos itens e do modelo, foram considerados os seguintes crit\u00e9rios: vari\u00e2ncia explicada do modelo (60 a 70%), valores de cargas fatoriais , comunalidades e discrimina\u00e7\u00e3o do item, problemas de colinearidade e multicolinearidade .Robust Mean-Scaled Chi Square/df NNFI , CFI , GFI , AGFI , RMSEA e RMSR .29-31Os \u00edndices de ajustamento do modelo e os respectivos valores esperados foram: Greatest Lower Bound).Foram adotados tr\u00eas indicadores para a avalia\u00e7\u00e3o da confiabilidade da vers\u00e3o brasileira do question\u00e1rio FSAS: Alfa de Cronbach, \u00d4mega de McDonald e GLB . Houve predom\u00ednio do sexo masculino que correspondeu a 64% dos casos. A maioria dos pacientes era da ra\u00e7a branca e 49% haviam cursado o ensino fundamental .Dentre as doen\u00e7as card\u00edacas, chama a aten\u00e7\u00e3o a Doen\u00e7a de Chagas, que esteve presente em 30,5% dos casos, seguida pela cardiopatia isqu\u00eamica em 25,2%. A S\u00edndrome de Brugada e a S\u00edndrome do QT longo cong\u00eanita foram identificadas em 4,6 e 3,3% dos pacientes, respectivamente.New York Heart Association. A fun\u00e7\u00e3o ventricular esquerda determinada pelo ecocardiograma transtor\u00e1cico bidimensional variou de 18 a 77%, com mediana de 35%.A avalia\u00e7\u00e3o basal mostrou que a maioria dos pacientes estava em classe funcional I e II , segundo os crit\u00e9rios da Apenas 29,1% dos pacientes n\u00e3o apresentavam comorbidades associadas. Dislipidemia e hipertens\u00e3o arterial foram as comorbidades mais frequentes, estando presentes em 51,4% e 49,5% dos pacientes, respectivamente. Fibrila\u00e7\u00e3o atrial estava presente em 27,1% dos indiv\u00edduos estudados .Como j\u00e1 era esperado, 80,1% das indica\u00e7\u00f5es do CDI corresponderam \u00e0 profilaxia secund\u00e1ria da morte s\u00fabita arr\u00edtmica, visto que, na realidade brasileira, devido \u00e0 escassez de recursos, o implante de CDI ainda \u00e9 pouco utilizado para profilaxia prim\u00e1ria da morte s\u00fabita.Quanto ao tipo de CDI, sistemas unicamerais estavam presentes em 41,1% dos pacientes, seguidos pelos sistemas dupla-c\u00e2mara . Apenas 12,6% dos pacientes apresentavam CDI com ressincronizador card\u00edaco, por terem doen\u00e7a card\u00edaca mais avan\u00e7ada. O tempo de uso do CDI foi de 6,7 \u00b1 4,4 anos, variando de 6 meses a 16,9 anos. O n\u00famero expressivo de pacientes que haviam recebido terapias de choque do CDI foi condizente com a indica\u00e7\u00e3o cl\u00ednica do dispositivo, visto que terapias de choque s\u00e3o muito mais frequentes em indiv\u00edduos submetidos \u00e0 profilaxia secund\u00e1ria da morte s\u00fabita.Atrav\u00e9s da an\u00e1lise descritiva dos itens do instrumento, foi poss\u00edvel identificar que houve viola\u00e7\u00e3o da normalidade da distribui\u00e7\u00e3o, indicando, portanto, a necessidade da utiliza\u00e7\u00e3o de correla\u00e7\u00f5es polic\u00f3ricas, ao inv\u00e9s da correla\u00e7\u00e3o de Pearson.As m\u00e9dias dos itens do instrumento variaram de 1,5 a 2,9. O escore m\u00e9dio do FSAS foi de 22,8 \u00b1 11,1, com mediana de 20 pontos e varia\u00e7\u00e3o de 10 a 50 pontos. N\u00e3o houve efeito de valores extremos na m\u00e9dia .2= 565,5, df= 45; p< 0,001) e o determinante da matriz ) revelaram uma correla\u00e7\u00e3o significativa entre os itens, confirmando a adequa\u00e7\u00e3o da aplica\u00e7\u00e3o da AFE.Os valores obtidos do \u00edndice de Kaiser-Meyer-Olkin , do teste de esfericidade de Bartlett tamb\u00e9m apontou apenas uma dimens\u00e3o com um autovalor de 6,08. O fato de o instrumento ser unidimensional dispensou a necessidade de t\u00e9cnicas rotacionais da matriz fatorial. A unidimensionalidade indicou o uso da t\u00e9cnica Normal-ogive graded response model para a TRI, mais adequada para modelo unidimensional polit\u00f4mico.31A an\u00e1lise paralela indicou a exist\u00eancia de apenas uma dimens\u00e3o para o instrumento. Ademais, esse conjunto de itens consegue explicar 64,4% da vari\u00e1vel latente . cross-loading). As comunalidades variaram de 0,46 a 0,74, com todos os itens acima do limite de 0,40. Para a discrimina\u00e7\u00e3o do item (a), os valores variaram de 0,91 a 1,71, tamb\u00e9m demonstrando boa ader\u00eancia \u00e0 vari\u00e1vel latente e confirmando os dados obtidos atrav\u00e9s das cargas fatoriais.A Robust Mean and Variance-Adjusted Chi Square X2/ df (35) = 40,40; p < 0,243; NNFI= 0,997; CFI= 0,997; GFI= 0,986; AGFI= 0,982. Os indicadores de res\u00edduos ficaram em n\u00edveis bons , indicando pouca diferen\u00e7a entre a matriz original e a matriz gerada pela carga dos fatores.31A AFC revelou um bom ajuste ao modelo unidimensional, com valores semelhantes aos recomendados pela literatura: Encontramos evid\u00eancias satisfat\u00f3rias de confiabilidade da escala FSAS-Br, com coeficiente alfa de Cronbach de 0,92, \u00d4mega de McDonald de 0,92 e GLB de 0,98.25 A vers\u00e3o final da escala FSAS para a l\u00edngua portuguesa (FSAS-Br) falada no Brasil, apresentou equival\u00eancia conceitual, sem\u00e2ntica, cultural e de mensura\u00e7\u00e3o em rela\u00e7\u00e3o aos itens originais no ingl\u00eas.16No presente estudo, foi descrito o processo de tradu\u00e7\u00e3o e adapta\u00e7\u00e3o transcultural de uma escala de avalia\u00e7\u00e3o do n\u00edvel de ansiedade relacionado \u00e0s terapias de choque do desfibrilador implant\u00e1vel, atendendo ao rigor metodol\u00f3gico preconizado na literatura internacional.Esfor\u00e7os foram empreendidos para incluir pacientes com diferentes perfis sociodemogr\u00e1ficos e diferentes tipos de doen\u00e7a card\u00edaca para garantir uma representa\u00e7\u00e3o heterog\u00eanea, visando assegurar a melhor calibra\u00e7\u00e3o dos itens do instrumento. Desse modo, pacientes com diferentes tipos de CDI foram inclu\u00eddos, assim como pacientes com indica\u00e7\u00e3o de profilaxia prim\u00e1ria ou secund\u00e1ria da morte s\u00fabita card\u00edaca. N\u00e3o obstante, os principais grupos de cardiopatias comuns a esse perfil de pacientes tamb\u00e9m foram contemplados, com expressiva preval\u00eancia da Doen\u00e7a de Chagas, cardiopatia isqu\u00eamica e hipertr\u00f3fica.22 Desse modo, \u00e9 importante ressaltar que a escala FSAS n\u00e3o foi projetada para avaliar aspectos relevantes da adapta\u00e7\u00e3o ao dispositivo e o seu real impacto na qualidade de vida, fazendo-se necess\u00e1ria a utiliza\u00e7\u00e3o de outros instrumentos para complementar a avalia\u00e7\u00e3o desses pacientes. Nesse sentido, os autores da escala FSAS desenvolveram outro instrumento, o Florida Patient Acceptance Survey (FPAS),32 que visa avaliar a adapta\u00e7\u00e3o psicossocial dos portadores de dispositivos card\u00edacos implant\u00e1veis. Os resultados do processo de adapta\u00e7\u00e3o transcultural e valida\u00e7\u00e3o para a l\u00edngua portuguesa da escala FPAS ser\u00e3o publicados oportunamente.No cen\u00e1rio internacional, a escala FSAS tem sido amplamente utilizada em diferentes contextos, j\u00e1 que apresenta boa sensibilidade para identificar o n\u00edvel de ansiedade relacionado \u00e0s terapias de choque do CDI e tempo reduzido para preenchimento.31 Na publica\u00e7\u00e3o original do FSAS, os autores postulam que o instrumento \u00e9 bidimensional, apresentando uma dimens\u00e3o composta por 7 itens (\u201cConsequence\u201d) e outra com 3 itens (\u201cTrigger\u201d).15 Esse modelo n\u00e3o foi reprodut\u00edvel para a vers\u00e3o brasileira, visto que todas as an\u00e1lises realizadas no presente estudo sustentaram a unidimensionalidade da escala FSAS-Br. Revisitando o estudo de Kuhl et al.,15 \u00e9 importante ressaltar que a amostra foi constitu\u00edda por apenas 72 participantes, o que pode ter causado impacto nos resultados das an\u00e1lises psicom\u00e9tricas.A comprova\u00e7\u00e3o das evid\u00eancias de validade de um instrumento tem sido recomendada pela comunidade cient\u00edfica como forma de verificar se o instrumento realmente mensura a vari\u00e1vel latente de interesse com precis\u00e3o. Al\u00e9m disso, \u00e9 importante analisar se a estrutura fatorial do instrumento est\u00e1 adequadamente representada pela sua dimensionalidade, ou seja, o n\u00famero de dimens\u00f5es que comp\u00f5em o instrumento de avalia\u00e7\u00e3o.Shock anxiety\u201d), ou seja, as duas dimens\u00f5es identificadas previamente poderiam ser melhor explicadas pela sua associa\u00e7\u00e3o a um fator comum, que seria a dimens\u00e3o \u201cAnsiedade relacionada ao choque\u201d. Em virtude desses resultados, os autores passaram a recomendar a utiliza\u00e7\u00e3o da pontua\u00e7\u00e3o total da escala FSAS, ao inv\u00e9s de subdividi-la nas duas dimens\u00f5es anteriormente descritas. Esses resultados corroboraram a estrutura fatorial identificada em nosso estudo.Posteriormente, a escala FSAS foi submetida a um processo de avalia\u00e7\u00e3o de suas propriedades psicom\u00e9tricas, dessa vez, com uma casu\u00edstica de 443 participantes.16 A AFC mostrou que as duas dimens\u00f5es anteriormente identificadas eram altamente correlacionadas a um fator de segunda ordem for prevention of sudden cardiac death, especially among patients with ventricular dysfunction and arrhythmogenic genetic diseases.6The primary purpose of ICD is to correct potentially fatal ventricular arrhythmias by delivering low- or high-energy therapy. Low-energy therapy, known as antitachycardia pacing or antitachycardia pacing (ATP), is a painless method. High-energy therapy delivers shocks of up to 40 J which, in spite of causing major discomfort, usually occur after the patient has lost consciousness, since they are applied about 15 seconds after the initiation of ventricular fibrillation or fast ventricular tachyarrhythmia. In undesirable situations, such as arrhythmias resistant to overstimulation, or in electrical storm, high-energy discharges can occur in awake patients.8 On the other hand, when the ICD is used for secondary prevention, the incidence of shock therapies may vary between 35 and 53%, within the first year after implantation.8 Despite the high level of technological sophistication of ICDs, unfortunately, there is the risk that the patient may receive inappropriate shock deliveries as a result of erroneous discrimination between supraventricular and ventricular tachyarrhythmias. On these occasions, the sensation reported is a painful and distressing experience.14It is estimated that the chances of ICD patients will need appropriate electric shocks for primary prevention of sudden cardiac death varies between 2 and 15% per year.14 On the other hand, ICD implantation has been reported to provide the patient with a great sense of safety, considering the device\u2019s capacity to interrupt unexpected episodes of potentially fatal ventricular arrhythmias.14ICD patients live with the expectation that, at any moment, the device will deliver shock therapies to interrupt ventricular arrhythmias resulting from their heart disease. Thus, although they recognize the benefits of the treatment, some patients may present with anxiety, depression, mood disorders, post-traumatic stress disorder, as well as fear that the device will not operate in crucial situations.16 The Florida Shock Anxiety Scale (FSAS) quickly achieved wide international acceptance, and has been translated and validated in several countries , with consistent results.In face of the concern about the deleterious effects of ICD on psychosocial adaptation, an scale was specifically developed to assess the level of anxiety related to the presence of ICD and to the shocks delivered by the device, for use both in clinical practice and in the context of scientific research.The purpose of the present study was to assess the psychometric properties of the Brazilian version of the FSAS for ICD patients.This study was conducted in a high-complexity cardiology hospital and it was approved by that hospital\u2019s Committee of Ethics in Research. All subjects signed a free and informed consent form.This was a psychometric study of cross-cultural adaptation and validation of the FSAS.16The FSAS was developed in 2006 in the United States to provide a quantitative measure of ICD shock-related anxiety. The instrument consists of 10 items, with five response options , corresponding to a 5-point Likert scale.Questions are related to patients\u2019 fear or anxiety caused by the expectation that the device may deliver shock therapies and to the behavioral changes to avoid the occurrence of ICD therapies.18The instrument can be self-administered or administered by interview.The FSAS total score is determined by the sum of all items, with a maximum score of 50 points. The higher the score, the higher the anxiety level. The items receiving three points or more should be considered the most critical aspects.25The expert committee was composed by professionals of the area of artificial cardiac stimulation and by nurses.The cross-cultural adaptation process of the FSAS followed international guidelines and included five stages: (1) Translation by two independent translators; (2) Synthesis of the translations; (3) Back-translation; (4) Harmonization of the translations by the expert committee; (5) Pretest with the target population; (6) Pretest review and final translation.1 and T2) were reconciled into one version (T1-2), after discussion with the expert committee. Working from the final version and blind to the original version, two bilingual teachers carried out the back-translations (RT1 and RT2). The aim of this stage was to measure the semantic and idiomatic consistency of the translations produced in the first stage. Finally, a new meeting was held with the expert committee to review all the cross-cultural adaptation process and undertake the harmonization across versions, thus obtaining the pre-final version of the instrument adults, aged between 18 and 80 years, of both sexes and with any education level; (2) ICD implanted for more than 6 months; (3) capable of understanding and answering the questionnaire used in the study; (4) having agreed to participate in the study by signing the informed consent form. We did not include in the study patients presenting at least one of the following situations: (1) indication for cardiac transplantation; (2) ongoing pregnancy; (3) malignant neoplasia.26 (Research Electronic Data Capture) hosted at the hospital\u2019s server.Patients were selected consecutively, during outpatient care or by visits to the inpatient unit of our institution. Individuals who met the eligibility criteria were invited to answer the FSAS questionnaire. At the same time, demographic, clinical and ICD data were collected by using electronic case report forms developed in REDCap25Sample size determination for psychometric studies is usually calculated based on the number of items of the instrument. Some studies have demonstrated that a ratio of 20:1 or greater, that is, 20 participants per item would be ideal. However, ratios of 10:1 are sufficient to allow for adequate analysis. Thus, a minimum sample number of 150 patients was established.Detailed descriptive analysis was performed, using measures of central tendency . The Kolmogorov-Smirnov (KS) test was used to test the normality of each item in the questionnaire, whereas the Mardia test was employed to assess multivariate normality.All analyses were performed using SPSS 23 statistical package software and Factor 10.8.01, adopting a level of significance of 5%.In this study, we conducted an exploratory factor analysis (EFA) and a confirmation factor analysis (CFA) to verify the dimensionality of the FSAS in its Portuguese version.28The robustness of the test was determined from the association of a bootstrap with sample extrapolation to 5,000. Factor extraction was done initially with Robust Unweighted Least Squares (RULS), which reduces the matrix of residuals.29The dimensionality testing was performed using Robust Parallel Analysis (RPA) through the Optimal implementation of Parallel Analysis (PA) with minimum rank factor analysis (MRFA), which minimizes the common variance of residuals.Item discrimination index was used (a), which measures the association strength between the item and the latent variable, and whose interpretation is similar to factor loading in the exploratory factor analysis.To adequate the items and the models, the following criteria were taken into account: the explained variance of the model (60 to 70%), factor loading values (> 0.50), communalities (> 0.40) and item discrimination, and collinearity and multicollinearity problems (factor loads ranging from 0.80 to 0.85).31The model adjustment indices and their respective expected values were: Robust Mean-Scaled Chi Square/df NNFI (Non-Normed Fit Index > 0.93), CFI (Comparative Fit Index > 0.94), GFI (Goodness Fit Index > 0.95), AGFI (Adjusted Goodness Fit Index > 0.93), RMSEA (Root Mean Square Error of Approximation < 0.07) and RMSR .Three indicators were adopted to assess the reliability of the Brazilian version of the FSAS questionnaire: Coefficient Alpha (\u201cCronbach\u2019s Alpha\u201d), Omega and the Greatest Lower Bound (GLB).The stages of the translation and cross-cultural adaptation resulted in similar versions of the FSAS instrument. The synthesis of the translations was quite concise and combined the most coherent elements of each translation. The back-translations confirmed the good quality of the translations and the synthesis process carried out in the initial stages.A total of 20 ICD patients, with a mean of age 55.6 \u00b1 6.8 years, took part in the pretest. Of these, 50% were female, 50% were white and 30% had studied up to High School. All participants reported that the items were relevant, easy to understand and that the response options were clear. No modifications in the instrument were required. In this stage of the study, 151 ICD patients, with a mean of 55.7\u00b114.1 years , were included. There was a male sex predominance, which corresponded to 64% of the cases. Most patients were white (85.4%) and 49% had attended Middle School .Among the cardiac diseases, there was a predominance of Chagas disease, which was present in 30.5% of the cases, followed by ischemic cardiomyopathy in 25.2%. Brugada syndrome and congenital long-QT syndrome (LQTS) were identified in 4.6 and 3.3% of patients, respectively.Baseline assessment showed that most patients were in the New York Heart Association (NYHA) functional classes I (37.1%) and II (47.7%). Left ventricular function was determined by bidimensional transthoracic echocardiography and ranged from 18 to 77%, with a median of 35%.Only 29.1% of the patients did not present any associated comorbidities. Dyslipidemia and arterial hypertension were the most frequent comorbidities, being present in 51.4% and 49.5% of patients, respectively. Atrial fibrillation was present in 27.1% of the individuals studied .As expected, 80.1% of the indications for ICD were r secondary prophylaxis of sudden cardiac death. In Brazil, due to lack of resources, ICD implantation is still underused for primary prophylaxis of sudden cardiac death.Through descriptive analysis of the instrument items, it was possible to identify that normality of distribution was violated, indicating, therefore, the need for polychoric correlation, instead of Pearson\u2019s correlation coefficient.The means of the instrument items ranged from 1.5 to 2.9. The FSAS average score was 22.8 \u00b1 11.1, with a median of 20 points and variation of 10 to 50 points. There was no impact of extreme values on the mean .2= 565.5, df= 45; p<0.001) and the matrix determinant (0.0206 (p<0.0001)) revealed a significant correlation between the items, which confirmed the adequacy of the EFA.The values obtained from the Kaiser-Meyer-Olkin index (KMO= 0.88), the Bartlett\u2019s sphericity test .2/ df (35) = 40.40; p < 0.243; NNFI= 0.997; CFI= 0,997; GFI= 0.986; AGFI= 0.982. The residual indicators were at good levels , showing little difference between the original matrix and the matrix generated from factor loadings.31The CFA revealed good adjustment to the unidimensional model, with values similar to those recommended by the literature: Robust Mean and Variance-Adjusted Chi Square XSatisfactory reliability evidence was provided by the FSAS-Br scale, with a Cronbach\u2019s alpha coefficient of 0.92, a McDonald\u2019s Omega coefficient of 0.92 and GLB of 0.98.25 The final translation of the FSAS into Brazilian Portuguese (FSAS-Br) presented conceptual, semantic, cultural and measurement equivalences compared to the original items in English.16In the present study, we described the translation and cross-cultural adaptation process of a brief scale designed to provide a quantitative measure of ICD shock-related anxiety, folllowing international methodological standards.Efforts were made to include patients with different sociodemographic profiles and various types of underlying heart diseases to ensure heterogeneous representation, aiming at providing the best calibration of the items. Thus, patients with different ICD types were included, as well as patients with indications for primary or secondary prophylaxis of sudden cardiac death. Notwithstanding, the most common kinds of heart disease among these patients\u2019 profiles have also been contemplated, with expressive prevalence of Chagas Disease, ischemic and hypertrophic cardiomyopathy.22 Thus, it is important to highlight that the FSAS was not designed to assess relevant aspects of adaptation to the device and its real impact on quality of life, which makes it necessary to use other instruments to complement the assessment of these patients. In this sense, the authors who had created the FSAS developed another instrument, the Florida Patient Acceptance Survey (FPAS),32 which aims at assessing the psychosocial adjustment of ICD patients. The results of the cross-cultural adaptation and validation process of the FPAS into Portuguese will be published in due course.In the international scenario, the FSAS scale has been widely used in different scenarios, since it presents good sensitivity to identify the level of ICD shock-related anxiety and requires reduced time for completion.31 In the original publication of the FSAS, the authors claim that the instrument was bidimensional, presenting two dimensions: Consequence (composed of 7 items) and Trigger (composed of 3 items).15This model was not reproducible to the Brazilian version, because all analyses performed in this study supported the FSAS-Br scale unidimensionality. Revisiting the study by Kuhl et at.,15 it is important to highlight that the sample was constituted by only 72 participants, which may have had an impact on the results of the psychometric analyses.Evidence of validity of an instrument has been recommended by the scientific community as a way to check whether the instrument actually and accurately measures the latent variable of interest. In addition, it is important to analyze whether the instrument factor structure is adequately represented by its dimensionality, that is, the number of dimensions that make up the instrument of assessment.16 The CFA showed that the two previously identified dimensions were highly related to a second-order factor (\u201cShock anxiety\u201d). In other words, the two dimensions identified previously could have been better explained by their association to a common factor, namely the \u201cshock-related anxiety\u201d dimension. Due to these results, the authors recommended that the total scale score may be more clinically useful, instead of subdividing it into the two dimensios described before. These results corroborate the factor structure identified in our study.Afterwards, the psychometric properties of the FSAS were evaluated, with a sample of 443 participants.33Reliability assessment of the FSAS-Br scale revealed the accuracy of the Brazilian version, which was confirmed by adequate values of Cronbach\u2019s alpha, McDonald\u2019s Omega and GLB. The adoption of these three indications aimed to increase the accuracy of interpretation, since the Cronbach\u2019s coefficient alpha is affected by the nature of data distribution and by sample size. Besides, its values may be increased by extensive scales, parallel and/or redundant elements or limited coverage of the construct under analysis, decreasing the reliability of the measurement.In general, the results observed in the present study showed that the instrument is reliable and valid for application in Brazil, meeting the quality requirements for patient-reported outcome measurements.Although the population studied is larger than the samples of several other studies which have used the FSAS, further studies with more robust samples are crucial for the consolidation of its validity and for attesting its stability in the various possible scenarios and profiles of ICD patients.Further studies, evaluating the association of the FSAS-Br scores with the occurrence of ICD shock therapies and other clinic parameters will be useful to identify factors which may be associated with increased anxiety levels and, therefore, allow for the establishment of specific and personalized interventions for these patients.The FSAS-Br instrument presented consistent validity and reliability evidence and, therefore, its use can be recommended for the ICD population in Brazil, both in clinical practice and in scientific research."} +{"text": "Pacientes com HIV t\u00eam maior probabilidade de apresentar doen\u00e7as cardiovasculares quando comparados \u00e0 popula\u00e7\u00e3o em geral. Este foi um estudo de caso-controle que teve como objetivo avaliar quais fatores estavam associados a uma redu\u00e7\u00e3o na espessura m\u00e9dio-intimal da car\u00f3tida (IMT) da car\u00f3tida e ao aumento na dilata\u00e7\u00e3o mediada por fluxo (DMF) da art\u00e9ria braquial em pacientes com HIV que receberam atorvastatina + aspirina por um per\u00edodo de 6 meses. Foi realizada uma an\u00e1lise secund\u00e1ria de um ensaio cl\u00ednico, que incluiu pessoas vivendo com HIV e baixo risco cardiovascular. Um total de 38 pacientes alocados para o bra\u00e7o de interven\u00e7\u00e3o e tratados por 6 meses com uma combina\u00e7\u00e3o de atorvastatina + aspirina foram inclu\u00eddos. Todos os participantes foram submetidos a ultrassonografia da car\u00f3tida e da art\u00e9ria braquial, tanto no in\u00edcio quanto no final do estudo. Os casos que responderam com aumento >10% da dilata\u00e7\u00e3o braquial (DMF) e redu\u00e7\u00e3o da espessura m\u00e9dio-intimal da car\u00f3tida (IMT) foram considerados casos, e aqueles que n\u00e3o responderam foram considerados controles. Avaliamos os fatores associados \u00e0s respostas positivas obtidas atrav\u00e9s da IMT e DMF. A redu\u00e7\u00e3o do IMT n\u00e3o se associou significativamente a nenhum dos fatores de risco avaliados: idade , sexo , tabagismo ou tempo de diagn\u00f3stico do HIV . Um aumento na DMF foi significativamente associado com a idade entre aqueles na faixa et\u00e1ria de 40-59 anos, p = 0,015 . Os indiv\u00edduos mais velhos foram mais propensos a apresentar um aumento na DMF ap\u00f3s 6 meses de tratamento com atorvastatina + aspirina. Isso se deve ao grande sucesso da terapia antirretroviral.8Um marcador precoce da aterosclerose \u00e9 a disfun\u00e7\u00e3o endotelial e prevenir essa disfun\u00e7\u00e3o pode ser uma alternativa para evitar futuros eventos cardiovasculares. A aspirina e, mais recentemente, as estatinas t\u00eam demonstrado efeitos pleiotr\u00f3picos, tais como: efeitos imunomoduladores, antitrombog\u00eanicos e anti-inflamat\u00f3rios. Tais medicamentos podem ser uma alternativa para a preven\u00e7\u00e3o prim\u00e1ria e secund\u00e1ria desses eventos em pessoas vivendo com HIV.intima-medial thickening ) e relataram resultados conflitantes.12 Para contribuir com essa discuss\u00e3o, nosso estudo tem como objetivo avaliar os fatores associados \u00e0 melhora da fun\u00e7\u00e3o endotelial e espessura da car\u00f3tida medida por DMF e IMT em indiv\u00edduos com HIV, com carga viral sob controle, que foram tratados com uma combina\u00e7\u00e3o de atorvastatina + aspirina por um per\u00edodo de 6 meses.Estudos observacionais e de interven\u00e7\u00e3o avaliaram os efeitos das estatinas na melhora da fun\u00e7\u00e3o endotelial e na progress\u00e3o da espessura da car\u00f3tida em indiv\u00edduos com e sem HIV. Esses estudos utilizaram t\u00e9cnicas de ultrassom n\u00e3o-invasivas, como a dilata\u00e7\u00e3o mediada por fluxo (DMF), que mede o fluxo mediado da art\u00e9ria braquial, e a espessura m\u00e9dio-intimal da car\u00f3tida e carga viral indetect\u00e1vel.Esta foi uma an\u00e1lise secund\u00e1ria de um ensaio cl\u00ednico ainda n\u00e3o publicado,O estudo foi planejado para 6 meses, utilizando regimes de tratamento com 2 inibidores de transcriptase reversa de nucleos\u00eddeos e 1 inibidor n\u00e3o nucleos\u00eddeo, que foram randomizados em grupos de interven\u00e7\u00e3o e placebo. Trinta e oito participantes foram alocados para o grupo de interven\u00e7\u00e3o e tratados por 6 meses com uma combina\u00e7\u00e3o de 20mg de atorvastatina + 100mg de aspirina, e 42 receberam placebo. O estudo avaliou a efic\u00e1cia da combina\u00e7\u00e3o de drogas atrav\u00e9s de medidas ultrassonogr\u00e1ficas do aumento da dilata\u00e7\u00e3o da art\u00e9ria braquial (DMF), espessura carot\u00eddea (IMT) reduzido e marcadores inflamat\u00f3rios e nenhuma diferen\u00e7a foi encontrada entre o grupo de interven\u00e7\u00e3o e o grupo placebo.No estudo de caso-controle apresentado aqui, foram inclu\u00eddos 38 indiv\u00edduos do grupo interven\u00e7\u00e3o do referido ensaio cl\u00ednico. O objetivo foi avaliar subgrupos que poderiam se beneficiar do uso de atorvastatina 20mg e aspirina 100mg na redu\u00e7\u00e3o da aterosclerose subcl\u00ednica e das doen\u00e7as cardiovasculares.14 e 14 pacientes considerados controles, por n\u00e3o terem apresentado resposta na DMF.Na primeira parte do estudo de caso-controle, um total de 38 indiv\u00edduos foram divididos em 24 casos, que foram aqueles que tiveram uma resposta favor\u00e1vel na DMF e conduzido no Ambulat\u00f3rio de Doen\u00e7as Infecciosas / Parasit\u00e1rias do Hospital das Cl\u00ednicas da Universidade Federal de Pernambuco / Recife, Brasil.Todos os indiv\u00edduos assinaram o termo de consentimento livre e esclarecido. O estudo foi aprovado pelo Comit\u00ea de \u00c9tica em Pesquisa da Universidade Federal de Pernambuco, sob n\u00famero 13097213.2.0000.5208. O ensaio cl\u00ednico foi registrado na \u00ae(GE) LOGIQe BT12 DICOM 3.0 AUTO IMT, com transdutor GE 9-L RS Linear, trabalhando na frequ\u00eancia de 7-10 MHz. As medidas foram realizadas de acordo com t\u00e9cnicas padronizadas.16Foi utilizado um aparelho de ultrassom General Electric\u00ae colocado no bra\u00e7o foi inflado a 30 mmHg acima da press\u00e3o sist\u00f3lica por 5 minutos e depois liberado. Um minuto ap\u00f3s a libera\u00e7\u00e3o do clampeamento, o di\u00e2metro da art\u00e9ria foi medido novamente. A dilata\u00e7\u00e3o normal foi considerada >10% - DMF: o di\u00e2metro da art\u00e9ria braquial foi medido em repouso e ap\u00f3s est\u00edmulo. Para estimular a art\u00e9ria braquial, um esfigmoman\u00f4metro Becton Dickinson17 ou mediana e intervalo interquartil (IIQ) para as vari\u00e1veis num\u00e9ricas e frequ\u00eancias absolutas e percentuais para as vari\u00e1veis categ\u00f3ricas e foram analisados inferencialmente atrav\u00e9s de testes estat\u00edsticos. Na compara\u00e7\u00e3o entre duas categorias, foram utilizados os seguintes testes: teste Os dados foram digitados em planilha EXCEL e o programa IBM-SPSS, vers\u00e3o 23, foi utilizado para a realiza\u00e7\u00e3o dos c\u00e1lculos estat\u00edsticos.As caracter\u00edsticas dos 38 sujeitos inclu\u00eddos no estudo est\u00e3o descritas na OR = 4,37, IC 95%: 1,07 - 17,79), em compara\u00e7\u00e3o com a obtida no grupo de 21 a 39 anos.Uma diferen\u00e7a estatisticamente significativa foi obtida para a m\u00e9dia de idade . Quando avaliadas as faixas et\u00e1rias (21-39 anos e 40-59 anos), a signific\u00e2ncia foi mantida . Ao avaliar a faixa et\u00e1ria mais avan\u00e7ada, observou-se que houve excelente resposta \u00e0 dilata\u00e7\u00e3o da art\u00e9ria braquial para o sexo feminino. Os demais fatores de risco avaliados n\u00e3o apresentaram signific\u00e2ncia estat\u00edstica: hipertens\u00e3o arterial sist\u00eamica ; diabetes mellitus ; tabagismo , como mostrado na Quando avaliamos o desfecho em rela\u00e7\u00e3o ao sexo, obteve-se um resultado lim\u00edtrofe ; sexo , HAS e DM ; tabagismo , IMC , como mostrado na Nosso estudo avaliou pacientes vivendo com HIV, recebendo terapia antirretroviral e com baixo risco cardiovascular, que tomaram a combina\u00e7\u00e3o de atorvastatina + aspirina por um per\u00edodo de 6 meses. Realizou-se uma an\u00e1lise explorat\u00f3ria para avaliar os fatores associados \u00e0 resposta positiva ao tratamento avaliado atrav\u00e9s das t\u00e9cnicas vasculares de DMF e IMT.18 Por outro lado, quanto maior o controle viral, melhor \u00e9 a fun\u00e7\u00e3o endotelial.19 Outra hip\u00f3tese seria de que indiv\u00edduos em uma faixa et\u00e1ria mais avan\u00e7ada estariam mais sujeitos \u00e0s consequ\u00eancias do processo ateroscler\u00f3tico relacionado \u00e0 idade e mais sens\u00edveis aos efeitos delet\u00e9rios do HIV sobre o endot\u00e9lio. Por sua vez, nossos achados podem sugerir que esses indiv\u00edduos mais idosos seriam mais responsivos \u00e0s a\u00e7\u00f5es pleiotr\u00f3picas e anti-inflamat\u00f3rias da combina\u00e7\u00e3o de atorvastatina + aspirina. Nossos achados sugerem que h\u00e1 benef\u00edcio do uso de estatinas + aspirina como profilaxia prim\u00e1ria para doen\u00e7as cardiovasculares em indiv\u00edduos com HIV, que deve ser avaliado de forma diferente em indiv\u00edduos de acordo com sua faixa et\u00e1ria, principalmente indiv\u00edduos com 40 anos ou mais.21Os resultados demonstraram que indiv\u00edduos pertencentes \u00e0 faixa et\u00e1ria mais avan\u00e7ada (entre 40 e 59 anos) responderam positivamente \u00e0 combina\u00e7\u00e3o atorvastatina + aspirina, ou seja, com aumento da DMF ao final do estudo. Pode-se inferir que indiv\u00edduos mais velhos foram expostos por mais tempo \u00e0 inflama\u00e7\u00e3o decorrente do HIV. Sabe-se que existem n\u00edveis mais elevados de inflama\u00e7\u00e3o nas pessoas com HIV do que nas n\u00e3o infectadas, mesmo aquelas sob controle virol\u00f3gico, e essa exposi\u00e7\u00e3o \u00e9 um fator importante na g\u00eanese da disfun\u00e7\u00e3o endotelial. Esses achados s\u00e3o semelhantes aos obtidos por outros autores, que verificaram que um alto n\u00edvel de replica\u00e7\u00e3o do v\u00edrus resulta em piora na dilata\u00e7\u00e3o da art\u00e9ria braquial.OR para o grupo feminino foi igual a 3,5. Embora n\u00e3o tenha apresentado signific\u00e2ncia estat\u00edstica, essa resposta chamou nossa aten\u00e7\u00e3o, pois sugere que as mulheres podem responder melhor ao tratamento com atorvastatina + aspirina do que os homens. Estudos t\u00eam sugerido que, entre as pessoas que vivem com HIV, as mulheres apresentam n\u00edveis mais elevados de ativa\u00e7\u00e3o imunol\u00f3gica e inflama\u00e7\u00e3o do que os homens.22 Considerando que os medicamentos atualmente em uso t\u00eam um efeito importante na redu\u00e7\u00e3o da inflama\u00e7\u00e3o, um mecanismo intrinsecamente relacionado \u00e0 progress\u00e3o da aterosclerose, pode-se inferir que esta poderia ser a poss\u00edvel raz\u00e3o para uma resposta mais evidente nas mulheres do que nos homens. Nosso estudo, entretanto, n\u00e3o foi capaz de confirmar essa associa\u00e7\u00e3o, mas outros que avaliaram um n\u00famero maior de indiv\u00edduos podem ter poder suficiente para alcan\u00e7ar signific\u00e2ncia estat\u00edstica. Estudos que associam o sexo com resposta \u00e0 fun\u00e7\u00e3o endotelial seriam necess\u00e1rios.Quando avaliamos a resposta relacionada ao sexo, obtivemos um resultado lim\u00edtrofe, no qual o 23 Dube et al.,24 em estudo transversal comparando indiv\u00edduos com ou sem uso de IP, n\u00e3o observaram diferen\u00e7a na resposta \u00e0 DMF. No entanto, v\u00e1rios outros autores descobriram maior espessura carot\u00eddea medido pelo IMT naqueles em uso de IP em compara\u00e7\u00e3o com aqueles que n\u00e3o receberam IP.26Os regimes antirretrovirais utilizados n\u00e3o foram significativamente associados \u00e0s respostas de DMF e IMT; entretanto, eles inclu\u00edram apenas inibidores de transcriptase reversa an\u00e1logos de nucleos\u00eddeos (ITRNs) e n\u00e3o-an\u00e1logos. Pacientes em uso de inibidores de protease (IP) ou inibidores da integrase (INI) n\u00e3o foram inclu\u00eddos. Sabe-se que entre os medicamentos utilizados atualmente, os IP causam mais dist\u00farbios metab\u00f3licos do que os demais e consequentemente predisp\u00f5em a um maior risco cardiovascular.O uso de esquemas de tratamento com grupos restritos de antirretrovirais tem como objetivo homogeneizar os grupos de compara\u00e7\u00e3o e evitar que os medicamentos se tornem fatores de confus\u00e3o quanto \u00e0 resposta ao uso de atorvastatina + aspirina.27 o que, como mencionado anteriormente, resulta em maior risco cardiovascular. Um estudo de coorte relacionou o tabagismo \u00e0 piora da progress\u00e3o do espessura carot\u00eddea.28 Estudos com maior n\u00famero de pacientes s\u00e3o necess\u00e1rios para determinar o papel dessa interven\u00e7\u00e3o em fumantes.O tabagismo n\u00e3o foi associado \u00e0s respostas de DMF ou IMT. Deve-se enfatizar que a baixa preval\u00eancia do tabagismo pode ter dificultado a avalia\u00e7\u00e3o do papel que o mesmo desempenhou. No entanto, deve-se ressaltar que, na avalia\u00e7\u00e3o do IMT, os n\u00e3o fumantes apresentaram chance 4,3 vezes maior de obter redu\u00e7\u00e3o do IMT com atorvastatina + aspirina. Contudo, provavelmente devido ao pequeno n\u00famero de casos, o intervalo de confian\u00e7a foi alto e n\u00e3o houve signific\u00e2ncia estat\u00edstica. Um estudo recente demonstrou que o tabagismo resulta em controle viral e resposta imunol\u00f3gica ruim,29 Dados relataram uma rela\u00e7\u00e3o entre lipodistrofia e fun\u00e7\u00e3o endotelial deficiente30 e aumento da espessura carot\u00eddea, principalmente em indiv\u00edduos com obesidade visceral.31 Em nosso estudo, n\u00e3o diagnosticamos lipodistrofia. Avaliamos apenas a composi\u00e7\u00e3o corporal com o \u00edndice de massa corporal (IMC) e classificamos os indiv\u00edduos de acordo com a presen\u00e7a de baixo peso, eutrofia, sobrepeso ou obesidade. Entretanto, como existe uma alta preval\u00eancia de lipodistrofia em pacientes com HIV e o IMC n\u00e3o \u00e9 um \u00edndice que possa fornecer uma correla\u00e7\u00e3o com esse dist\u00farbio, tal fator de risco deve ser avaliado nesses indiv\u00edduos.Nossos achados n\u00e3o revelaram uma associa\u00e7\u00e3o entre obesidade e resposta da fun\u00e7\u00e3o endotelial medida pela DMF, ou progress\u00e3o do espessura carot\u00eddea (IMT) ap\u00f3s receber atorvastatina + aspirina. Um estudo de coorte que monitorou pacientes obesos com HIV e os comparou com obesos n\u00e3o infectados pelo HIV demonstrou uma maior incid\u00eancia de dist\u00farbios do metabolismo da glicose e inflama\u00e7\u00e3o entre aqueles com HIV, embora DMF e IMT n\u00e3o tenham diferido entre os dois grupos.32 H\u00e1 dados que demonstram que os dois m\u00e9todos s\u00e3o \u00fanicos e independentes e n\u00e3o se correlacionam entre si, embora sejam considerados v\u00e1lidos para a detec\u00e7\u00e3o da aterosclerose subcl\u00ednica. Eles provavelmente refletem diferentes aspectos e est\u00e1gios da aterosclerose inicial.33 Portanto, a diverg\u00eancia dos resultados em nosso estudo est\u00e1 de acordo com a literatura, e demonstra que a DMF mostrou ser capaz de identificar o benef\u00edcio da utiliza\u00e7\u00e3o da combina\u00e7\u00e3o de atorvastatina + aspirina em indiv\u00edduos HIV-positivos com idade entre 40-59 anos quando comparados com pacientes mais jovens.A associa\u00e7\u00e3o da idade com a resposta positiva ao tratamento foi diferente quando comparada aos m\u00e9todos utilizados para sua avalia\u00e7\u00e3o: enquanto a DMF apresentou melhora com o tratamento nos pacientes mais velhos, a avalia\u00e7\u00e3o do IMT n\u00e3o demonstrou essa diferen\u00e7a entre os grupos. A DMF e o IMT s\u00e3o frequentemente utilizados como medidas substitutas para aterosclerose subcl\u00ednica. Enquanto o IMT identifica anormalidades estruturais precoces, a DMF, considerada um bioensaio endotelial, avalia a integridade funcional do vaso.O estudo cl\u00ednico original demonstrou uma redu\u00e7\u00e3o percentual dos n\u00edveis de LDL nos indiv\u00edduos do grupo caso , mas sem melhora da fun\u00e7\u00e3o endotelial. Consideramos algumas limita\u00e7\u00f5es nesse estudo, destacando-se o tempo de uso das estatinas, que foi planejado e realizado por um per\u00edodo de 6 meses. Estudos que demonstraram resultados encorajadores utilizaram as estatinas por per\u00edodos muito mais longos do que o nosso, sugerindo um caminho a ser seguido. Outra quest\u00e3o diz respeito ao perfil dos pacientes envolvidos em nosso estudo. Todos apresentavam poucos fatores tradicionais de risco cardiovascular, a carga do HIV estava sob controle e eles estavam em tratamento antirretroviral h\u00e1 v\u00e1rios anos. Essa sele\u00e7\u00e3o resultou em um grupo de indiv\u00edduos com pouca ou nenhuma inflama\u00e7\u00e3o, como demonstrado pelos baixos n\u00edveis de marcadores inflamat\u00f3rios, revelando assim uma popula\u00e7\u00e3o para a qual o uso de estatinas em curto prazo associado \u00e0 aspirina provavelmente n\u00e3o forneceria resultados eficazes.Os pontos fortes destacados pelo presente estudo seriam a sele\u00e7\u00e3o de indiv\u00edduos com baixo risco cardiovascular e o uso de medicamentos antirretrovirais com baixo potencial para causar dist\u00farbios metab\u00f3licos. Essas caracter\u00edsticas permitem investiga\u00e7\u00f5es sobre os poss\u00edveis efeitos dos medicamentos e os fatores associados a um melhor desfecho no est\u00e1gio inicial da doen\u00e7a ateroscler\u00f3tica, ou seja, o per\u00edodo em que ocorrem altera\u00e7\u00f5es no endot\u00e9lio vascular, sendo, portanto, um processo que pode ser revertido. Uma poss\u00edvel fragilidade, por\u00e9m, que deve ser destacada, foi o fato de o estudo envolver um n\u00famero pequeno de indiv\u00edduos. Tal amostra pode ter sido insuficiente para detectar poss\u00edveis associa\u00e7\u00f5es a fatores que possivelmente poderiam ser observados em uma maior amostra de indiv\u00edduos.O estudo revelou que o fator idade influencia a melhora da fun\u00e7\u00e3o endotelial em indiv\u00edduos com HIV e baixo risco cardiovascular tratados com a combina\u00e7\u00e3o de atorvastatina + aspirina. Tamb\u00e9m mostrou que a DMF \u00e9 um m\u00e9todo capaz de revelar esse efeito. Estudos semelhantes, envolvendo um maior n\u00famero de indiv\u00edduos, s\u00e3o necess\u00e1rios para confirmar nossa hip\u00f3tese e apoiar o uso precoce da combina\u00e7\u00e3o atorvastatina + aspirina em indiv\u00edduos de 40 a 59 anos, em tratamento antirretroviral e com baixo risco cardiovascular para preven\u00e7\u00e3o de doen\u00e7as cardiovasculares. 1 Living with the virus has now become a chronic condition, which imposes the challenge of maintaining viral suppression coupled with the management of age-related comorbidities.2 A substantial increase in non-AIDS-related deaths, such as those related to cardiovascular diseases, has been reported,3 and are more prevalent in these individuals, when compared to the general population.5Life expectation and quality of life among people infected with HIV has increased significantly over recent decades. This is due to the great success of antiretroviral therapy.8An early marker for atherosclerosis is endothelial dysfunction and preventing this dysfunction may be an alternative for preventing future cardiovascular events. Aspirin and, more recently, statins have demonstrated pleiotropic effects, such as: immunomodulatory, and antithrombogenic and anti-inflammatory effects. Such medications may be an alternative for the primary and secondary prevention of these events among people living with HIV.12 To contribute to this discussion, our study aims to assess the factors associated with endothelial function improvement and carotid thickness measured by FMD and IMT in subjects with HIV, with a viral load under control, who were treated with a combination of atorvastatin + aspirin for a period of 6 months.Observational and interventional studies have evaluated the effects of statins in improving endothelial function, and the progression of carotid thickening in individuals both with and without HIV. These studies have used non-invasive ultrasound techniques, such as FMD, which measures the mediated flow of the brachial artery, and IMT, which measures carotid intima-media thickening, and have reported conflicting results.13 in which 80 participants who presented with low cardiovascular risk, as measured by the Framingham Risk Score (FRS), and an undetectable viral load were assessed.This was a secondary analysis of a clinical trial not yet publishedThe study was planned for 6 months, using 2 nucleoside reverse transcriptase inhibitors and 1 non-nucleoside inhibitor regimens, which were randomized into intervention and placebo groups. Thirty-eight participants were allocated to the intervention group and treated for 6 months with a combination of 20mg atorvastatin + 100mg aspirin, and 42 received placebo. The study assessed the efficacy of the drug combination through ultrasound measurements of the increased brachial artery dilation (FMD), reduced carotid thickening (IMT), and inflammatory markers and no difference was found between the intervention group and the placebo group.In the case-control study presented herein, 38 individuals from the intervention group of the aforementioned clinical trial were included. The aim was to assess subgroups that could benefit from the use of atorvastatin 20mg and aspirin 100mg in reducing subclinical atherosclerosis and cardiovascular disease.14 and 14 patients who were considered controls, as they did not show response in FMD.In the first part of the case-control study, a total of 38 individuals were divided into 24 cases, which were those who had a favorable response in FMD and conducted at the Infectious/Parasitic Diseases Outpatient Clinic at Hospital das Cl\u00ednicas, Universidade Federal de Pernambuco/Recife, Brazil.16A General Electric\u2122 (GE) LOGIQe BT12 DICOM 3.0 AUTO IMT ultrasound device was used, with a GE 9-L RS Linear transducer, working at a frequency of 7-10 MHz. The measurements were performed according to standardized techniques.FMD: The brachial artery diameter was measured at rest and after stimulation. To stimulate the brachial artery, a Becton Dickinson\u2122 sphygmomanometer placed on the arm was inflated to 30mmHg above the systolic pressure for 5 minutes, and then released. One minute after releasing the clamp, the diameter of the artery was measured once again. Normal dilation was considered > 10% - 17 or median and interquartile range (IQR) for numerical variables and absolute and percentage frequencies for categorical variables and were analyzed inferentially through statistical tests. In the comparison between two categories, the following tests were used: unpaired Student\u2019s t-test with equal variances or Mann-Whitney test for the numerical variables and Pearson\u2019s Chi-square test or Fisher\u2019s Exact for the categorical variables. Student\u2019s t-test was used with variables with normal distribution and Mann-Whitney\u2019s test with variables with a non-normal distribution. Fisher\u2019s Exact test was used in cases where the condition for using the Chi-square test was not verified. The verification of data normality was performed by the Shapiro-Wilk\u2019s test and the hypothesis of equality of variances through the Levene F-test. The level of statistical significance adopted was 5% and the confidence intervals were 95.0%The data were entered into the EXCEL spreadsheet and the IMB-SPSS program, version 23, was used to perform the statistical calculations.The characteristics of the 38 subjects included in the study are described in A statistically significant difference was obtained for the mean age (p = 0.015). When age ranges were assessed (21-39 years and 40-59 years), the significance was maintained (p = 0.034). When assessing the older age group, it was observed that there was an excellent response to brachial artery dilation , compared to that obtained in the 21- 39 year-old group.When we assessed the outcome regarding sex, a borderline result was obtained for female subjects. The other risk factors assessed did not show any statistical significance: systemic arterial hypertension ; diabetes mellitus ; smoking (p = 0.383) in No statistically significant differences were observed for any of the variables assessed in relation to a reduction in the carotid intima-media thickness: age (p = 0.706); gender (p = 0.260), SAH and DM (p = 1.00); smoking (p = 0.131), BMI (p = 0.945), as shown in Our study assessed patients living with HIV, receiving antiretroviral therapy and with a low cardiovascular risk, who took a combination of atorvastatin + aspirin during a period of 6 months. An exploratory analysis was performed in order to evaluate the factors associated with a positive response to the treatment assessed through FMD and IMT vascular techniques.18 Conversely, the higher the viral control, the better the endothelial function.19 Another hypothesis would be that individuals at an older age range would be more prone to the consequences of the age-related atherosclerotic process and more sensitive to the deleterious effects of HIV on the endothelium. In turn, our findings may suggest that these older individuals would be more responsive to the pleiotropic and anti-inflammatory actions of the combination of atorvastatin + aspirin. Our findings suggest that there is a benefit of the use of statins + aspirin as a primary prophylaxis for cardiovascular disease in individuals with HIV, which should be assessed differently in individuals according to their age group, particularly individuals aged 40 years or older.21The results have demonstrated that individuals belonging to the older age group (between 40 and 59 years) responded positively to a combination of atorvastatin + aspirin, i.e., with increased FMD by the end of the study. It may be inferred that older individuals have been exposed for a longer period to the inflammation resulting from the HIV. It is known that there are higher levels of inflammation in people with HIV than in non-infected people, even those under virological control, and this exposure is an important factor in the genesis of endothelial dysfunction. These findings are similar to those obtained by other authors, who have verified that a high level of virus replication results in a brachial artery dilation worsening.22 Considering that the currently used medications have an important effect in reducing inflammation, a mechanism intrinsically related to the progression of atherosclerosis, one could infer that this may be the possible reason for a more evident response in women than in men. Our study, however, was unable to confirm this association, but others that have assessed a larger number of individuals may have sufficient power to obtain statistical significance. Studies that associate gender with response to endothelial function would be necessary.When we assessed the response related to gender, we obtained a borderline result, in which the OR for the female group was equal to 3.5. Although there was no statistical significance, this response nonetheless attracted our attention, since it suggests that females may respond better to treatment with atorvastatin + aspirin than males. Studies have suggested that amongst people living with HIV, women show higher levels of immune activation and inflammation than men.23 Dube et al.,24 in a cross-sectional study comparing individuals with or without the use of PIs, observed no difference regarding the response to FMD. However, several other authors have discovered greater carotid thickening measured by IMT in those receiving PIs when compared to those not receiving them.26The antiretroviral regimens used were not significantly associated with FMD and IMT responses; however, they only included analogue and non-analogue nucleoside NRTIs. Patients receiving protease inhibitors (PIs) or integrase inhibitors (INI) were not included. It is known that amongst the currently used medications, the PIs cause more metabolic disorders than the others and, consequently, they predispose to a higher cardiovascular risk.The use of regimens with restricted groups of antiretroviral drugs have aimed to homogenize the comparison groups and prevent medications from becoming confounding factors regarding the response to atorvastatin + aspirin.27 which, as previously mentioned, results in a higher cardiovascular risk. One cohort study related smoking to a worsening progression of carotid thickening.28 Studies with a higher number of patients are necessary in order to determine the role of this intervention in smokers.Smoking was not associated with FMD or IMT responses. It should be emphasized that the low prevalence of smoking may have made it difficult to assess the role it played. However, it should be noted that in the IMT assessment, non-smokers showed a 4.3-fold higher chance of obtaining an IMT reduction with atorvastatin + aspirin. However, probably due to the small number of cases, the confidence interval was high (0.70 - 27.01) and there was no statistical significance. One recent study demonstrated that smoking results in poor viral control and immune response,29 Data have reported a relationship between lipodystrophy and poor endothelial function30 and increased carotid thickening, especially among individuals with visceral obesity.31 In our study, we did not diagnose lipodystrophy. We only assessed body composition with the body mass index (BMI) and classified individuals according to low weight, normal weight, overweight or obesity. However, because there is a high prevalence of lipodystrophy among HIV patients, and BMI is not an index that may provide us with a correlation with this disorder, this risk factor should be evaluated in these individuals.Our findings revealed no association between obesity and an endothelial function response measured by FMD, or carotid thickness (IMT) progression after receiving atorvastatin + aspirin. A cohort study that monitored obese patients with HIV and compared them with non-HIV-infected obese individuals, demonstrated a higher incidence of glucose metabolism disorders and inflammation amongst those with HIV, although FMD and IMT did not differ between the two groups.32 There are data demonstrating that the two methods are unique and independent and do not correlate with one other, although they are considered valid for detecting subclinical atherosclerosis. They probably reflect different aspects and stages of early atherosclerosis.33 Therefore, the divergence of the results in our study is consistent with the literature and demonstrates that FMD has shown to be able to identify the benefit of using the combination of atorvastatin + aspirin in HIV-positive individuals aged 40-59 years when compared to younger patients.The association of age with a positive response to treatment was different when compared to the methods used for its assessment: while FMD displayed an improvement with treatment in the older patients, the IMT assessment did not demonstrate this difference between the groups. FMD and IMT are frequently used as surrogate measures for subclinical atherosclerosis. While IMT identifies early structural abnormalities, FMD, considered an endothelial bioassay, assesses the functional integrity of the vessel.The original clinical study demonstrated a percentage reduction in LDL levels in individuals in the case group , but without improving endothelial function. We considered some limitations in that study, and we would highlight the time receiving statins, which was planned and conducted for a duration of 6 months. Studies that have demonstrated encouraging results used statins for much longer periods than ours, thereby suggesting a path to be followed. One further question concerns the profile of the patients involved in our study. They all presented with few traditional factors of cardiovascular risk, the HIV viral load was well under control and they had been on antiretroviral treatment for several years. This selection resulted in a group of individuals with little or no inflammation, as shown by the low levels of inflammatory markers, thus revealing a population for which the short-term use of statins associated with aspirin would probably not provide any effective results.The strong points highlighted by the present study would be the selection of individuals with low cardiovascular risk and the use of antiretroviral drugs with a low potential for causing metabolic disorders. These characteristics enable investigations into the possible effects of the drugs and the factors associated with a better outcome in an early stage of atherosclerotic disease, i.e., the period in which changes in the vascular endothelium occur, being therefore a process that can be reversed. One possible weak point, however, which should be highlighted, was the fact that the study involved a small number of individuals. The present sample may have been insufficient to detect possible associations to factors that could possibly be observed in a larger sample of individuals.The study has shown that the age factor influences endothelial function improvement in subjects with HIV and low cardiovascular risk receiving a combination of atorvastatin + aspirin. It has also shown that FMD is a method capable of disclosing this effect. Similar studies involving a greater number of individuals are needed to confirm our hypothesis and to support the early use of the combination of atorvastatin + aspirin in subjects aged 40 to 59 years, undergoing antiretroviral treatment and with low cardiovascular risk for the prevention of cardiovascular disease."} +{"text": "A \u00faltima Diretriz de Insufici\u00eancia Card\u00edaca do Departamento de Insufici\u00eancia Card\u00edaca da Sociedade Brasileira de Cardiologia (DEIC/SBC) foi finalizada em mar\u00e7o de 2018. A partir de ent\u00e3o, houve um importante n\u00famero de interven\u00e7\u00f5es terap\u00eauticas e abordagens diagn\u00f3sticas que surgiram ou se consolidaram na pr\u00e1tica cl\u00ednica internacional e na pesquisa cl\u00ednica. Ao lado disso, a pandemia da COVID-19 trouxe-nos conhecimento sobre o modelo fisiopatol\u00f3gico da agress\u00e3o mioc\u00e1rdica e muitas d\u00favidas acerca da continuidade e da seguran\u00e7a dos medicamentos nos pacientes com IC cr\u00f4nica que apresentaram quadro agudo dessa complexa e nova entidade cl\u00ednica.Nos \u00faltimos 6 meses, trabalhamos de forma r\u00e1pida e colaborativa utilizando pela primeira vez em 20 anos do DEIC as plataformas digitais para discutir, deliberar e redigir esse importante documento, optando por realizar uma revis\u00e3o focada em vez de uma ampla atualiza\u00e7\u00e3o da Diretriz ainda muito recente.I Heart Failure Summit Brazil 2020 , com cerca de 900 participantes, muitos destes associados do DEIC.Inspiramo-nos no modelo de atualiza\u00e7\u00e3o da Diretriz Canadense de Insufici\u00eancia Card\u00edaca de 2020,A lideran\u00e7a da Diretoria Cient\u00edfica foi fundamental para a organiza\u00e7\u00e3o de diferentes grupos de trabalhos e elabora\u00e7\u00e3o de uma forma pr\u00e1tica e segura de discuss\u00e3o e vota\u00e7\u00e3o. Garantindo o distanciamento social e empregando a tecnologia digital, o encontro permitiu amplos debates sobre os diferentes pontos de vista alicer\u00e7ados nas melhores evid\u00eancias cient\u00edficas.No presente documento, o DEIC/SBC apresenta uma revis\u00e3o e uma atualiza\u00e7\u00e3o detalhadas de sua Diretriz de Insufici\u00eancia Card\u00edaca Cr\u00f4nica. Os trabalhos tiveram in\u00edcio no m\u00eas de julho de 2020, com a defini\u00e7\u00e3o do Comit\u00ea Editor, que estabeleceu prioridades, dividiu os 52 participantes em grupos de trabalho e definiu o cronograma das atividades. Estes grupos, compostos por cinco a sete participantes cada, deram in\u00edcio a intensas discuss\u00f5es virtuais que culminaram com a reda\u00e7\u00e3o de tabelas preliminares, sendo posteriormente amplamente divulgadas e revisadas pelo Comit\u00ea Revisor composto por 11 membros. As discuss\u00f5es finais foram realizadas em plen\u00e1ria virtual em 4 de dezembro de 2020, com a participa\u00e7\u00e3o de todos os colaboradores, nas quais as principais recomenda\u00e7\u00f5es foram votadas. As decis\u00f5es quanto \u00e0s classes de recomenda\u00e7\u00e3o foram definidas com a concord\u00e2ncia de mais de 75% dos participantes.As defini\u00e7\u00f5es de Classes de Recomenda\u00e7\u00e3o e N\u00edvel de Evid\u00eancia respeitam as mesmas normas da \u00faltima diretriz, conforme preconiza o SBC/CONDir para elabora\u00e7\u00e3o de diretrizes e s\u00e3o assim descritas:As recomenda\u00e7\u00f5es terap\u00eauticas propostas no presente documento embasam-se nas evid\u00eancias cient\u00edficas mais atuais, considerando n\u00e3o apenas os aspectos de efic\u00e1cia cl\u00ednica demonstrados em grandes ensaios cl\u00ednicos. Buscamos sumarizar as principais recomenda\u00e7\u00f5es em fluxogramas e algoritmos de f\u00e1cil entendimento e grande aplicabilidade cl\u00ednica, propondo abordagens para o diagn\u00f3stico e o tratamento da insufici\u00eancia card\u00edaca.Nosso compromisso com a comunidade cient\u00edfica, ligado \u00e0 pesquisa e \u00e0 assist\u00eancia aos pacientes com insufici\u00eancia card\u00edaca, gestores p\u00fablicos e privados e tamb\u00e9m formuladores de pol\u00edticas p\u00fablicas, certamente contar\u00e1 com um documento que buscou apresentar as interven\u00e7\u00f5es cient\u00edficas de forma did\u00e1tica e, assim, facilitar sua implanta\u00e7\u00e3o nas diferentes esferas de atendimento do paciente com insufici\u00eancia card\u00edaca.Dr. Evandro Tinoco Mesquita2FPEF (esquerda) quanto a HFA-PEFF (direita) podem ser utilizadas deve ser efetuada com dados cl\u00ednicos, eletrocardiogr\u00e1ficos, ecocardiogr\u00e1fico e laboratorial. A seguir, na ilizadas e 1.2. Nilizadas .\u2013At\u00e9 o momento, ainda n\u00e3o h\u00e1 interven\u00e7\u00e3o espec\u00edfica que reduza eventos cardiovasculares de pacientes com ICFEp. Os ensaios cl\u00ednicos que avaliaram o uso de inibidores da enzima de convers\u00e3o de angiotensina II (iECA), bloqueadores dos receptores de angiotensina II (BRA), inibidores da neprilisina e antagonistas dos receptores de angiotensina II (INRA) e espironolactona foram neutros quanto \u00e0 redu\u00e7\u00e3o do risco de eventos comparado ao placebo para pacientes com ICFEp.\u2013Assistimos recentemente a grandes avan\u00e7os no conhecimento da amiloidose card\u00edaca, o que acarretou na profunda transforma\u00e7\u00e3o do seu significado cl\u00ednico, epidemiol\u00f3gico e no surgimento de tratamentos espec\u00edficos. V\u00e1rias evid\u00eancias sugerem que a amiloidose card\u00edaca n\u00e3o seja uma doen\u00e7a rara, mas uma condi\u00e7\u00e3o amplamente subdiagnosticada, considerada hoje uma causa relativamente comum e trat\u00e1vel de insufici\u00eancia card\u00edaca com fra\u00e7\u00e3o de eje\u00e7\u00e3o preservada (ICFEp), particularmente a amiloidose card\u00edaca ligada \u00e0 transtirretina (ATTR) na sua forma selvagem ou sist\u00eamica senil (ATTR-wt), cujo diagn\u00f3stico tem aumentado de forma expressiva.Trata-se de uma doen\u00e7a multisist\u00eamica causada pela deposi\u00e7\u00e3o tecidual de prote\u00ednas fibrilares insol\u00faveis que perdem a sua conforma\u00e7\u00e3o, o que leva \u00e0 disfun\u00e7\u00e3o org\u00e2nica, inclusive do cora\u00e7\u00e3o. Mais de 30 tipos de prote\u00ednas amiloidog\u00eanicas s\u00e3o descritas,A AL apresenta incid\u00eancia de 6 a 10 milh\u00f5es de pessoas por ano e era considerada a principal causa de amiloidose card\u00edaca.Tendo em vista que a ATTR, particularmente a ATTRwt, \u00e9 uma condi\u00e7\u00e3o mais prevalente do que se antecipava, \u00e9 importante suspeitar dessa condi\u00e7\u00e3o na presen\u00e7a de pistas cl\u00ednicas para posterior investiga\u00e7\u00e3o diagn\u00f3stica . Por se Al\u00e9m disso, certas manifesta\u00e7\u00f5es multisist\u00eamicas podem levantar suspeita de ATTR: s\u00edndrome de t\u00fanel do carpo bilateral, ruptura do tend\u00e3o do b\u00edceps, hipotens\u00e3o ortost\u00e1tica, estenose do canal vertebral, altera\u00e7\u00f5es digestivas e intoler\u00e2ncia a medica\u00e7\u00f5es anti-hipertensivas.Diante da suspeita da doen\u00e7a, o primeiro passo \u00e9 investigar a presen\u00e7a de cadeias leves de imunoglobulinas para o diagn\u00f3stico da AL, uma vez que essa forma da amiloidose card\u00edaca exige tratamento espec\u00edfico com quimioter\u00e1picos e o progn\u00f3stico piora muito com o retardo no in\u00edcio do tratamento. A confirma\u00e7\u00e3o da AL depende da detec\u00e7\u00e3o da prote\u00edna amiloide em tecidos envolvidos (biopsia), mas a forma ATTR pode ser confirmada n\u00e3o invasivamente, mediante emprego de cintilografia card\u00edaca com radiotra\u00e7adores \u00f3sseos. No Brasil, \u00e9 usado o Tc-99m-pirofosfato.A baixa voltagem no complexo QRS \u00e9 achado comum na AL, sendo menos prevalente na ATTR (aproximadamente 30% dos casos), sendo mais comum a discrep\u00e2ncia entre a magnitude da hipertrofia ao ecocardiograma e a amplitude dos complexos QRS. Fibrila\u00e7\u00e3o atrial e o padr\u00e3o de \u201cpseudoinfarto\u201d tamb\u00e9m podem ser encontrados.strain sist\u00f3lico longitudinal pode mostrar a preserva\u00e7\u00e3o da contratilidade do \u00e1pice do ventr\u00edculo esquerdo com rela\u00e7\u00e3o aos demais segmentos .\u00c9 um dos principais exames para levantar a suspeita. Entre os achados sugestivos se destacam o espessamento da parede ventricular esquerda maior que 12 mm, especialmente na aus\u00eancia de hipertens\u00e3o arterial, aumento bi-atrial e desproporcional ao tamanho dos ventr\u00edculos, espessamento das valvas atrioventriculares e do septo interatrial, e o aumento da ecogenicidade do mioc\u00e1rdio com apar\u00eancia granular. O \u00edndice de deforma\u00e7\u00e3o sist\u00f3lica longitudinal do mioc\u00e1rdio ou Cintilografia card\u00edaca com radiotra\u00e7adores \u00f3sseos, como Tc99m-pirofosfato usado no Brasil, pode ser utilizada para o diagn\u00f3stico diferencial entre a amiloidose AL e ATTR, esta \u00faltima mostrando capta\u00e7\u00e3o mioc\u00e1rdica an\u00f4mala com intensidade maior ou equivalente \u00e0 \u00f3ssea. No entanto, a capta\u00e7\u00e3o card\u00edaca pode ocorrer, ainda que mais discreta, em at\u00e9 30% dos casos de AL. A capta\u00e7\u00e3o card\u00edaca intensa (grau 2 ou 3), em conjunto com aus\u00eancia de cadeias leves nos exames bioqu\u00edmicos, tem especificidade de 100% para ATTR, podendo dispensar a biopsia card\u00edaca para o diagn\u00f3stico da doen\u00e7a.A Resson\u00e2ncia Magn\u00e9tica Card\u00edaca possui alta sensibilidade e especificidade para o diagn\u00f3stico, sendo \u00fatil tamb\u00e9m para diferenciar a amiloidose card\u00edaca de outras miocardiopatias. A deposi\u00e7\u00e3o amiloide no mioc\u00e1rdio causa aumento no volume de distribui\u00e7\u00e3o do contraste paramagn\u00e9tico nas regi\u00f5es do mioc\u00e1rdio em que os cardiomi\u00f3citos s\u00e3o substitu\u00eddos ou deslocados por fibrose ou inflama\u00e7\u00e3o, cursando com padr\u00e3o de realce tardio (RT) mais comumente subendoc\u00e1rdico difuso e circunferencial do ventr\u00edculo esquerdo, ainda que realces tardios transmural e difuso tamb\u00e9m possam ser encontrados.Frente \u00e0 import\u00e2ncia cl\u00ednica e epidemiol\u00f3gica, al\u00e9m de novas terapias emergentes para esta doen\u00e7a, um Posicionamento sobre Diagn\u00f3stico e Tratamento da Amiloidose Card\u00edaca ser\u00e1 publicado em breve, abordando de forma mais ampla os diferentes aspectos da doen\u00e7a.A hist\u00f3ria natural da IC caracteriza-se pela piora progressiva da fun\u00e7\u00e3o card\u00edaca e dos sintomas de IC. Apesar dos avan\u00e7os no tratamento farmacol\u00f3gico e do impacto progn\u00f3stico dos dispositivos implant\u00e1veis como terapia de ressincroniza\u00e7\u00e3o card\u00edaca, pacientes com insufici\u00eancia card\u00edaca podem progredir para uma condi\u00e7\u00e3o cl\u00ednica denominada IC avan\u00e7ada, em que o tratamento tradicional n\u00e3o \u00e9 efetivo e as terapias avan\u00e7adas, tais como transplante card\u00edaco, suporte com dispositivo de assist\u00eancia circulat\u00f3ria mec\u00e2nica (DACM) ou cuidados paliativos s\u00e3o necess\u00e1rias.\u2013,\u2013Embora o termo IC avan\u00e7ada j\u00e1 venha sendo usado desde 2007, atualiza\u00e7\u00f5es recentes foram descritas visando incluir situa\u00e7\u00f5es cl\u00ednicas que tamb\u00e9m podem requerer terapias avan\u00e7adas e que n\u00e3o foram contempladas na primeira classifica\u00e7\u00e3o, como pacientes com ICFEp com quadro restritivo grave e n\u00e3o limitando apenas aos pacientes com IC com fra\u00e7\u00e3o de eje\u00e7\u00e3o gravemente reduzida.Estes crit\u00e9rios variam de acordo com as diferentes sociedades de cardiologia, por\u00e9m a presen\u00e7a de sintomas graves persistentes; a capacidade reduzida ao exerc\u00edcio; e epis\u00f3dios recorrentes de congest\u00e3o pulmonar ou sist\u00eamica que necessitem hospitaliza\u00e7\u00f5es est\u00e3o presentes em todas elas como descrito na \u00c9 importante ressaltar que o reconhecimento precoce de um paciente com IC avan\u00e7ada \u00e9 fator determinante para o seu progn\u00f3stico, uma vez que isto permitir\u00e1 o encaminhamento para um centro especializado que disponha das terapias avan\u00e7adas necess\u00e1rias para a condu\u00e7\u00e3o do caso.I-NEED-HELP, que integra a hist\u00f3ria cl\u00ednica, as hospitaliza\u00e7\u00f5es e a intoler\u00e2ncia medicamentosa, al\u00e9m de sintomas e a disfun\u00e7\u00e3o de \u00f3rg\u00e3o-alvo. para atender potenciais cadidatos ao transplante card\u00edaco (e posterior seguimento) e pacientes em choque cardiog\u00eanico (CC). Ele assume a lideran\u00e7a na coordena\u00e7\u00e3o dos trabalhos envolvendo o time de choque e, portanto, est\u00e1 familiarizado com as diversas (e crescentes) op\u00e7\u00f5es de uso de assist\u00eancia circulat\u00f3ria. Por fim, este profissional deve ser capaz de compreender o momento e as implica\u00e7\u00f5es de discutir cuidados paliativos e diretivas antecipadas de vontade para pacientes n\u00e3o eleg\u00edveis para transplante card\u00edaco, bem como o uso de dispositivos de longa perman\u00eancia.Society for Cardiovascular Angiography and Interventions (SCAI) prop\u00f4s uma nova classifica\u00e7\u00e3o para o CC, visando facilitar a identifica\u00e7\u00e3o das diferentes fases de deteriora\u00e7\u00e3o cl\u00ednica e a necessidade de intensifica\u00e7\u00e3o do tratamento., .O est\u00e1gio A inclui pacientes sob risco de choque cardiog\u00eanico, enquanto os est\u00e1gios B a E descrevem fases progressivas do choque cardiog\u00eanico convencional. A diferen\u00e7a entre os est\u00e1gios B e C \u00e9 a presen\u00e7a de hipoperfus\u00e3o, que est\u00e1 presente nos est\u00e1gios C e superiores. O est\u00e1gio D indica que as medidas de manejo inicial do choque cardiog\u00eanico n\u00e3o foram suficientes para restaurar a estabilidade hemodin\u00e2mica ou a perfus\u00e3o tecidual ap\u00f3s pelo menos 30 minutos de observa\u00e7\u00e3o, enquanto o est\u00e1gio E caracteriza casos extremos, no qual os pacientes se apresentam hemodinamicamente inst\u00e1veis e frequentemente em colapso circulat\u00f3rio. Pacientes em est\u00e1gios SCAI D e E apresentam maior mortalidade e podem se beneficiar da transfer\u00eancia precoce para centros especializados, capazes de oferecer modalidades avan\u00e7adas de suporte circulat\u00f3rio. The latest Heart Failure Guidelines by the Department of Heart Failure of the Brazilian Society of Cardiology (DEIC/SBC) were finalized on March 2018. Since then, a significant number of therapeutic interventions and diagnostic approaches has arisen or consolidated their position in international clinical practice and in clinical research. In addition, the COVID-19 pandemic has taught us much about the pathophysiological model of myocardial damage and raised many questions about the continuity and safety of medication use in patients with chronic HF suffering from an acute manifestation of this new and complex clinical entity.In the last few months, we have been working quickly and collaboratively, and for the first time in 20 years, DEIC used digital platforms to discuss, deliberate, and draft this important document, opting for a focused update instead of a full-text guideline.We were inspired by the 2020 Canadian Heart Failure Guidelines,The leadership of the Science Board was key in organizing the various working groups and developing a secure and practical method for discussions and votes. With social distancing and the use of digital technology, the conference enabled wide-ranging debates from various perspectives, based on the best available scientific evidence.In this document, DEIC/SBC provides reviews and detailed updates to its Chronic Heart Failure Guidelines. The work started in July 2020, with the choice of the Editorial Board, which established priorities, divided the 52 participants into working groups, and developed a schedule of activities. These working groups, each consisting of five to seven participants, began intense online discussions that led to the elaboration of preliminary tables, widely circulated before their subsequent review by the 11-member Review Board. The final discussions took place during a virtual plenary session on December 4, 2020, with all collaborators, who had the opportunity to vote on the main recommendations. Decisions regarding classes of recommendation required a three-quarters supermajority vote.Class of Recommendation and Level of Evidence follow the same definitions used in the last guideline, as established by SBC/CONDir. See below.The therapeutic recommendations proposed in this document are based on the latest available scientific evidence, considering not only the aspects of clinical efficacy from large clinical trials. We have sought to summarize the primary recommendations in flowcharts and algorithms that are easy to understand and to apply in clinical practice, proposing approaches for the diagnosis and treatment of heart failure.Our commitment to the scientific community, linked to research and assistance to heart failure patients, public and private managers, and policy-makers, will certainly have the benefit of a document that sought to present scientific interventions in an accessible format, facilitating its implementation in the various spheres where heart failure patients receive care.Dr. Evandro Tinoco Mesquita2FPEF should be based on clinical, electrocardiography, echocardiography, and laboratory data. Next, two scoring systems have been developed to check that diagnosis; both the H2FPEF and the 2FPEF scores m2FPEF .To date, there is no specific intervention to reduce cardiovascular events in patients with HFpEF. Clinical trials assessing the use of angiotensin-converting enzyme II inhibitors (ACEIs), angiotensin II receptor blockers (ARBs), angiotensin II receptor-neprilysin inhibitors (ARNIs), and spironolactone were neutral in terms of risk reduction compared to placebo for patients with HFpEF.We have recently seen great advances in cardiac amyloidosis, with a profound transformation of its clinical and epidemiological significance and the development of specific treatments. Evidence suggests that cardiac amyloidosis is not a rare disease, but rather a largely underdiagnosed condition, now considered a relatively common and treatable cause of HFpEF, particularly transthyretin amyloidosis (ATTR) in its wild type (ATTR-wt), of which diagnosis has increased expressively.It is a multisystemic disease caused by the tissue deposition of insoluble fibrillary proteins that lose their conformation, leading to organ dysfunction, including the heart. Over 30 types of amyloidogenic proteins have been described,AL incidence ranges from 6 to 10 million people/year and, until recently, was considered the primary cause of cardiac amyloidosis.Considering that ATTR, particularly ATTRwt, is more prevalent than previously expected, it is important to suspect it in the presence of clinical clues for further diagnostic investigation . ATTR coWhen suspected, the first step in investigating cardiac amyloidosis is the search for the presence of immunoglobulin light chains for the diagnosis of AL, which requires specific treatment with chemotherapeutic agents and has a worse prognosis with delayed treatment initiation. Confirmation of AL depends on the detection of amyloid protein in the tissues involved (biopsy), but the ATTR form can be confirmed noninvasively, using cardiac scintigraphy with bone-avid radiotracers. In Brazil, Tc-99m pyrophosphate is used in the examination.A low-amplitude QRS complex is a frequent finding in AL, but less prevalent in ATTR (around 30% of cases), that more commonly presents discrepancy between the magnitude of the hypertrophy on the echocardiogram and the amplitude of QRS complexes is more frequent. Atrial fibrillation and a \u201cpseudo-infarction\u201d pattern may also be found.Echocardiogram is the most important exam to raise the suspicion of CA. Suggestive findings include left ventricular wall thickening greater than 12 mm, especially in the absence of hypertension, bi-atrial enlargement disproportionate to ventricle size, atrioventricular valve and interatrial septum thickening, and increased myocardial echogenicity with a granular aspect. Myocardial longitudinal systolic strain rates may show the preservation of left ventricular apical contractility as compared to the remaining segments as compared to the reduced contractility in the remaining segments.Cardiac scintigraphy with bone-avid radiotracers, such as Tc-99m pyrophosphate as used in Brazil, can be used for the differential diagnosis between amyloidosis AL and ATTR, with the latter showing anomalous myocardial uptake, higher than or equivalent to bone uptake. However, cardiac uptake may occur, albeit with milder intensity, in up to 30% of AL cases. The combination of intense cardiac uptake (grades 2 or 3) and the absence of light chains in biochemical exams presents 100% specificity for ATTR, and can obviate a cardiac biopsy for the diagnosis of the disease.Cardiac magnetic resonance imaging has high sensitivity and specificity for the diagnosis and discrimination between cardiac amyloidosis and other cardiomyopathies. Amyloid deposits in the myocardium cause an increase in the distribution volume of paramagnetic contrast agent in myocardial regions where cardiomyocytes are replaced or displaced by inflammation or fibrosis, originating a diffuse subendocardial and circumferential late enhancement pattern of the left ventricle; a diffuse transmural pattern can also be found.Given its clinical and epidemiological importance, in addition to new emerging therapies for the condition, a Position Paper on Diagnosis and Treatment of Cardiac Amyloidosis will be published shortly, and should review the different aspects of the disease more broadly.The natural history of HF is characterized by a progressive deterioration of cardiac function and HF symptoms. Despite advances in pharmacological treatment and the prognostic impact of implantable devices such as cardiac resynchronization therapy, HF patients may progress to a clinical condition known as advanced HF, where traditional treatment is not effective and advanced therapies are required, such as heart transplantation, mechanical circulatory support device (MCSD) or palliative care are required.\u2013,\u2013Although the expression advanced HF has been used since 2007, recent updates were described to include clinical situation that may also require advanced therapies such as HFpEF patients with severe restrictive condition, rather than limiting it to patients with HF with severely reduced ejection fraction.Different societies of cardiology adopt different criteria for the condition, but all of them include the presence of persistent severe symptoms, exercise intolerance, and recurrent episodes of systemic or pulmonary congestion requiring hospitalization, as described in Early recognition is decisive for the prognosis of patients with advanced HF, since it allows timely referral to a specialized center able to provide the necessary advanced therapies to manage such cases.A particularly useful mnemonic that may help identify patients requiring referral to a HF specialist is I-NEED-HELP, which combines clinical history, hospitalizations and intolerance to medications, as well as symptoms and end-organ dysfunction. As the specific profile of patients fitting the current definition of advanced HF becomes increasingly clear, there is also a need to define the importance of the specialist in advanced HF in specialized centers. These professionals must be familiar (and trained) in the care of potential heart transplant candidates and their subsequent follow-up, as well as in patients with CS. They should coordinate the work of the shock team and therefore must be familiar with the diverse and growing options for circulatory support. Finally, the advanced HF specialist should be able to understand the timing and implications of discussing palliative care and advanced directives for patients who are not eligible for heart transplantation, as well as the use of long-term devices.,In 2019, the Society for Cardiovascular Angiography and Interventions (SCAI) proposed a new classification for cardiogenic shock (CS) in order to make it easier to identify the various stages of clinical deterioration as well as the need for more intensive treatment.Stage A includes patients at risk of cardiogenic shock, while stages B through E describe progressive stages of conventional cardiogenic shock. The difference between stages B and C is the presence of hypoperfusion, present in stages C and above. Stage D indicates initial cardiogenic shock management measures were not enough to restore hemodynamic stability or tissue perfusion within at least 30 minutes of observation, while stage E characterizes extreme cases, where patients present as hemodynamically unstable and frequently in circulatory collapse. Patients in SCAI stages D and E have higher mortality rates and may benefit from early referral to specialized centers, where more advanced modes of circulatory support may be available."} +{"text": "Uma metodologia para identifica\u00e7\u00e3o de pacientes portadores de aneurisma de aorta ascendente (AAAs) sob alto risco de remodelamento a\u00f3rtico n\u00e3o est\u00e1 completamente definida. Esta pesquisa objetiva caracterizar numericamente o fluxo sangu\u00edneo a\u00f3rtico, relacionando a distribui\u00e7\u00e3o do estresse mec\u00e2nico resultante com o crescimento de AAAs. Estudo anal\u00edtico, observacional, unic\u00eantrico, em que um protocolo de fluidodin\u00e2mica computacional foi aplicado a imagens de angiotomografia computadorizada (ATC) de aorta de pacientes portadores de AAAs. Duas ATC de aorta com pelo menos um ano de intervalo foram obtidas. Dados cl\u00ednicos dos pacientes foram registrados e, a partir das imagens de ATC, foram gerados modelos tridimensionais. Foram realizados estudos do campo de velocidade e estruturas coerentes (v\u00f3rtices) com o objetivo de relacion\u00e1-los ao crescimento ou n\u00e3o do aneurisma e, posteriormente, compar\u00e1-los com os dados cl\u00ednicos dos pacientes. O teste de Kolmogorov-Smirnov foi utilizado para avaliar a normalidade da amostra e o teste n\u00e3o-param\u00e9trico Wilcoxon signed-rank foi aplicado para compara\u00e7\u00f5es de dados pareados entre os \u00e2ngulos a\u00f3rticos. A signific\u00e2ncia estat\u00edstica foi fixada em 5%. Para o grupo que apresentou crescimento do aneurisma, a incid\u00eancia do jato na parede a\u00f3rtica gerou \u00e1reas de recircula\u00e7\u00e3o posterior ao jato, induzindo \u00e0 forma\u00e7\u00e3o de v\u00f3rtices complexos, ocasionando um incremento na press\u00e3o m\u00e9dia no endot\u00e9lio a\u00f3rtico. O grupo sem crescimento do aneurisma apresentou diminui\u00e7\u00e3o na press\u00e3o m\u00e9dia. Este estudo piloto mostrou que a CFD baseada em ATC pode, em um futuro pr\u00f3ximo, ser uma ferramenta auxiliar na identifica\u00e7\u00e3o dos padr\u00f5es de fluxo associados ao processo de remodelamento de AAAs. As vari\u00e1veis relacionadas ao crescimento do AAAs ainda n\u00e3o s\u00e3o perfeitamente compreendidas.4Aneurisma de aorta ascendente (AAAs) \u00e9 geralmente assintom\u00e1tico e sua evolu\u00e7\u00e3o \u00e9 impercept\u00edvel.5 Segundo Hope et al.,4 mudan\u00e7as no comportamento do fluxo ao longo da aorta ascendente s\u00e3o relacionadas ao processo de remodelamento e, portanto, podem influenciar no crescimento aneurism\u00e1tico.A forma\u00e7\u00e3o do AAAs \u00e9 um processo degenerativo multifatorial, resultante de fatores hemodin\u00e2micos e processos biol\u00f3gicos.7 Como destacado por Morris et al.,7 modelos de CFD aplicados em escala populacional t\u00eam o potencial de reduzir riscos, custos e tempo associados aos estudos cl\u00ednicos.Cada vez mais, a fluidodin\u00e2mica computacional (CFD) tem se tornado uma ferramenta complementar para promover o melhor entendimento da patog\u00eanese e progress\u00e3o de doen\u00e7as cardiovasculares, al\u00e9m de se mostrar adequada para avalia\u00e7\u00f5es minimamente invasivas.8 sugerindo uma associa\u00e7\u00e3o dessas grandezas com a dilata\u00e7\u00e3o aneurism\u00e1tica.9 Essas observa\u00e7\u00f5es foram discutidas por G\u00fclan et al.,10 que encontraram fluxos rotacionais formados na fase sist\u00f3lica, mostrando similaridade qualitativa com a forma\u00e7\u00e3o de anel de v\u00f3rtice, isto \u00e9, um jato central cercado por dois grandes v\u00f3rtices. Sob o ponto de vista da mec\u00e2nica dos fluidos, o fluxo atrav\u00e9s de uma expans\u00e3o abrupta, como ocorre entre o anel a\u00f3rtico e a por\u00e7\u00e3o dilatada da aorta ascendente, leva \u00e0 separa\u00e7\u00e3o da camada limite e \u00e0 forma\u00e7\u00e3o de uma zona de separa\u00e7\u00e3o. Portanto, \u00e9 poss\u00edvel que haja uma correla\u00e7\u00e3o entre separa\u00e7\u00e3o do fluxo, turbul\u00eancia, varia\u00e7\u00e3o de press\u00e3o e crescimento do aneurisma.10Atrav\u00e9s de estudos de CFD do fluxo a\u00f3rtico, \u00e9 poss\u00edvel identificar regi\u00f5es da parede a\u00f3rtica com altos valores de tens\u00e3o de cisalhamento e de press\u00e3o,11 A A presen\u00e7a de estruturas coerentes dentro da aorta tamb\u00e9m pode explicar a forma\u00e7\u00e3o de regi\u00f5es com altas tens\u00f5es cisalhantes e altas press\u00f5es, o que pode contribuir com o processo de remodelamento a\u00f3rtico.O presente estudo \u00e9 anal\u00edtico, observacional, unic\u00eantrico, no qual um protocolo de CFD foi aplicado a imagens de angiotomografia computadorizada (ATC) de aorta. A pesquisa foi registrada na Comiss\u00e3o Nacional de \u00c9tica em Pesquisa do Minist\u00e9rio da Sa\u00fade e aprovada pelo comit\u00ea de \u00e9tica em pesquisa local . Foi obtido consentimento informado de todos os participantes, de acordo com a resolu\u00e7\u00e3o 466/2012 do Conselho Nacional de Sa\u00fade.12Para realiza\u00e7\u00e3o do estudo, nove pacientes com AAAs foram investigados, tendo sido selecionados a partir de avalia\u00e7\u00f5es de 100 pacientes consecutivos do ambulat\u00f3rio de aortopatias de um centro terci\u00e1rio de sa\u00fade. Pacientes com diagn\u00f3stico de AAAs foram inclu\u00eddos e aqueles com doen\u00e7as do col\u00e1geno, cirurgia card\u00edaca pr\u00e9via ou de aorta foram exclu\u00eddos. Para todos os pacientes, duas imagens de ATC, com um intervalo m\u00ednimo de um ano entre os exames, encontravam-se dispon\u00edveis. As imagens foram obtidas por indica\u00e7\u00f5es cl\u00ednicas de acompanhamento, n\u00e3o especificamente para o presente estudo. Os pacientes estavam sob tratamento cl\u00ednico, de acordo com as diretrizes recentes sobre AAAs.13A geometria de cada aorta foi constru\u00edda com base em uma varredura de ATC obtida por um scanner de 64 canais SOMATOM Sensation\u00ae 64 . Os cortes selecionados de ATC foram medidos desde o anel a\u00f3rtico at\u00e9 a aorta descendente. Informa\u00e7\u00f5es como tamanho de pixel e dist\u00e2ncia dos planos de aquisi\u00e7\u00e3o das imagens foram adquiridos a fim de ajustar o modelo a\u00f3rtico tridimensional (3D) ao seu tamanho real. A segmenta\u00e7\u00e3o da imagem foi realizada atrav\u00e9s do programa FIJI de processamento de imagem de c\u00f3digo aberto baseado em ImageJ, focado em an\u00e1lise de imagens biol\u00f3gicas.Dois modelos a\u00f3rticos 3D de cada paciente foram sobrepostos visando obter refer\u00eancia espacial e possibilitar a compara\u00e7\u00e3o entre os exames, atrav\u00e9s da superposi\u00e7\u00e3o do in\u00edcio do tronco braquiocef\u00e1lico e das art\u00e9rias coron\u00e1rias direitas . A aort\u03b8I e \u03b8II ) foram definidos entre o plano de entrada e a sa\u00edda do tronco braquiocef\u00e1lico, com o objetivo de relacion\u00e1-los com o padr\u00e3o de fluxo que pode induzir o crescimento de um aneurisma .21 Em condi\u00e7\u00f5es normais, o sangue pode ser considerado um fluido incompress\u00edvel,22 tendo a densidade sido fixada em \u03c1 = 1054 kg/m3 A viscosidade din\u00e2mica foi definida como \u03bc = 7,2 cP para permitir uma compara\u00e7\u00e3o direta com os dados experimentais in vitro .23 Adicionalmente, foram realizados alguns testes e os resultados obtidos com viscosidades iguais a 3,5 cP e 7,2 cP forneceram resultados equivalentes, com diferen\u00e7as de press\u00e3o inferiores a 0,01 mmHg na regi\u00e3o de interesse da aorta ascendente. Esses testes corroboram os achados de Becsek et al.,24 que avaliaram viscosidades iguais a 4 cP, 6 cP e 8 cP e mostraram efeito desprez\u00edvel no campo de escoamento m\u00e9dio e efeito muito pequeno com rela\u00e7\u00e3o a transi\u00e7\u00e3o turbulenta.Uma vez que s\u00e3o encontradas altas taxas de cisalhamento15 conforme mostrado na Para todos os casos, as mesmas distribui\u00e7\u00f5es de vaz\u00f5es nas sa\u00eddas (porcentagens baseadas no fluxo de entrada) foram impostas com base em valores m\u00e9dios do corpo humano, em acordo com Alastruey et al.,Fluent v18.1, que resolve as equa\u00e7\u00f5es de conserva\u00e7\u00e3o discretizadas com base no m\u00e9todo de volumes finitos.25 As simula\u00e7\u00f5es foram p\u00f3s-processadas com a ferramenta ANSYS CFD- Post tool26 e com o programa de c\u00f3digo aberto Paraview.27A presente an\u00e1lise foi realizada utilizando-se o programa ANSYS power-law \u201d,25 o acoplamento press\u00e3o-velocidade foi tratado pelo algoritmo SIMPLE25 e o erro residual m\u00e1ximo de converg\u00eancia foi definido em 10-6 para todas as equa\u00e7\u00f5es.As equa\u00e7\u00f5es discretizadas foram obtidas com o esquema \u201c Para todos os casos estudados, empregaram-se 400.000 n\u00f3s, os quais foram definidos ap\u00f3s a realiza\u00e7\u00e3o de um teste de independ\u00eancia da solu\u00e7\u00e3o na malha. Garantiram-se varia\u00e7\u00f5es inferiores a 0,3% na queda de press\u00e3o na \u00e1rea principal de interesse (aorta ascendente), ao dobrar a malha.\u03b8I e \u03b8II ; moderada ; bom ; excelente .A forma anat\u00f4mica da aorta de cada paciente pode ser inferida pelos \u00e2ngulos de refer\u00eancia \u03b8II , que fosigned-rank , foi aplicado para compara\u00e7\u00f5es de dados pareados (amostras dependentes) entre as medi\u00e7\u00f5es de \u03b8I e \u03b8II . A signific\u00e2ncia estat\u00edstica foi estabelecida em 5%. Todas as an\u00e1lises estat\u00edsticas foram conduzidas utilizando-se o software da IBM\u00ae SPSS Statistics\u00ae vers\u00e3o 26.29O teste de normalidade Kolmogorov-Smirnov foi usado para examinar se as vari\u00e1veis apresentavam distribui\u00e7\u00e3o normal. Devido \u00e0 distribui\u00e7\u00e3o n\u00e3o normal, o teste n\u00e3o param\u00e9trico Wilcoxon 30 Considerou-se que o aneurisma cresceu, quando se observou um aumento de 10% ou mais no volume da regi\u00e3o de interesse, ao comparar as duas ATC de cada paciente. A Os pacientes foram divididos em dois grupos de acordo com o volume da regi\u00e3o de interesse.Conforme mostrado na \u03b8I ( \u03b8I\u2265 94o) e pequeno \u03b8II (51o\u2264 \u03b8II\u2264 68o). Entretanto, ao examinar os \u00e2ngulos correspondentes aos pacientes sem crescimento do aneurisma, n\u00e3o se observou uma tend\u00eancia n\u00edtida. Existem pacientes com grandes e pequenos \u03b8I e \u03b8II . Os pacientes 6 e 7 apresentaram claramente uma faixa diferente de \u00e2ngulos, contudo, o mesmo n\u00e3o \u00e9 verdade para os pacientes 8 e 9. A medi\u00e7\u00e3o de \u03b8I em pacientes com crescimento de aneurisma foi de 127,47 \u00b0, naqueles sem crescimento do aneurisma foi de 119,22 \u00b0 e na popula\u00e7\u00e3o total foi de 121,53 \u00b0. As medi\u00e7\u00f5es de \u03b8II em pacientes com crescimento de aneurisma foi de 51,46 \u00b0, naqueles sem crescimento de aneurisma foi de 68,5 \u00b0 e da popula\u00e7\u00e3o total foi 57,94 \u00b0.As medidas correspondentes ao primeiro e ao segundo ano est\u00e3o listadas na w ) igual a 50% da velocidade axial de entrada ( win ) atingindo a parede distal, em ambos os anos. A curvatura angular do paciente 1 pode ter causado a forma\u00e7\u00e3o do v\u00f3rtice descolado. O paciente 2 (menor dilata\u00e7\u00e3o do grupo) apresentou um v\u00f3rtice toroidal pr\u00f3ximo \u00e0 raiz da aorta, com um v\u00f3rtice longitudinal ao longo da parede distal. O mesmo fen\u00f4meno foi observado para os pacientes 3 e 5. Examinando a iso-superf\u00edcie Q para o paciente 4, pode-se observar uma estrutura semelhante a um v\u00f3rtice hairpin , como visto para o paciente 1. Nesses pacientes, nota-se que o fluxo permaneceu pr\u00f3ximo \u00e0 parede a\u00f3rtica durante o per\u00edodo sist\u00f3lico.Analisando a \u03b8I ) apresenta uma estrutura toroidal pr\u00f3xima \u00e0 raiz da aorta e uma estrutura grande e compacta aproximando-se da parede distal, que aumentou em tamanho no segundo ano.Para todos os pacientes cujo aneurisma n\u00e3o cresceu , estrutExaminando a helicidade normalizada nas Comparando as estruturas coerentes e helicoidais de ambos os grupos, n\u00e3o h\u00e1 uma diferen\u00e7a clara entre eles, embora seja poss\u00edvel observar estruturas mais complexas no escoamento do grupo em que o aneurisma cresceu. Para esse grupo, as estruturas verticais s\u00e3o alongadas, terminando por formar um la\u00e7o, com forte v\u00f3rtice contra-rotativo, enquanto para o segundo grupo, o v\u00f3rtice toroidal permanece pr\u00f3ximo \u00e0 raiz a\u00f3rtica, girando predominantemente em uma dire\u00e7\u00e3o. O v\u00f3rtice mais intenso pode levar a diferentes n\u00edveis de estresse mec\u00e2nico no endot\u00e9lio a\u00f3rtico.Pm\u00e1x e \u00afPwall ), bem como a porcentagem da \u00e1rea da parede a\u00f3rtica ascendente com alta press\u00e3o ( %Acr ). Analisando essas figuras, pode-se associar ao jato de entrada, mostrado nas As Notavelmente, h\u00e1 uma tend\u00eancia para \u00e1reas maiores de press\u00f5es aumentadas no grupo de aneurisma com crescimento, com press\u00f5es m\u00e1ximas elevadas acima de 200 Pa, em compara\u00e7\u00e3o com o grupo de aneurisma sem crescimento, que demonstrou press\u00f5es m\u00e1ximas mais baixas (abaixo de 200 Pa) . Al\u00e9m disso, houve um aumento na press\u00e3o m\u00e9dia da parede para o grupo de aneurisma em crescimento e uma redu\u00e7\u00e3o para o outro grupo, exceto para o paciente 7, cuja press\u00e3o m\u00e1xima \u00e9 bastante baixa, o que tamb\u00e9m poderia explicar porque seu aneurisma n\u00e3o cresceu.O perfil cl\u00ednico da popula\u00e7\u00e3o estudada \u00e9 mostrado na 15 com poss\u00edvel aumento de aplica\u00e7\u00f5es cl\u00ednicas em um futuro pr\u00f3ximo. No presente estudo, pacientes foram classificados como portadores de AAAs com crescimento e sem crescimento de acordo com o volume da regi\u00e3o de interesse. Raghavan et al.30 demonstraram uma melhor correla\u00e7\u00e3o da poss\u00edvel ruptura do aneurisma com o volume da aorta ascendente e alto estresse na parede, do que apenas com o di\u00e2metro da aorta ascendente.A simula\u00e7\u00e3o computacional hemodin\u00e2mica tem se mostrado uma promissora \u00e1rea da pesquisa cient\u00edfica translacional\u03b8I e pequenos \u03b8II podem indicar a possibilidade de um processo de remodelamento do aneurisma da aorta.Biasetti et al.39Uma hip\u00f3tese para explicar esses achados \u00e9 a transfer\u00eancia de estresse \u00e0 advent\u00edcia por perda de deposi\u00e7\u00e3o de elastina e col\u00e1geno, promovendo cont\u00ednua adapta\u00e7\u00e3o por causa da distribui\u00e7\u00e3o da tens\u00e3o e, consequentemente, mudan\u00e7as na forma do AAAs.Existem algumas limita\u00e7\u00f5es a esta pesquisa. Primeiramente, neste estudo, a varredura por ATC foi realizada por diferentes equipes m\u00e9dicas. Como resultado, a qualidade dos exames n\u00e3o foi a mesma, o que pode ter afetado a acur\u00e1cia da compara\u00e7\u00e3o das imagens e a defini\u00e7\u00e3o da forma da aorta. Em segundo lugar, a pequena popula\u00e7\u00e3o deste estudo limita qualquer correla\u00e7\u00e3o entre padr\u00f5es cl\u00ednicos e crescimento do AAAs. Em terceiro lugar, dados cl\u00ednicos de dois pacientes n\u00e3o estavam dispon\u00edveis, assim, eles n\u00e3o foram inclu\u00eddos na Claramente, h\u00e1 muitos fatores que precisam ser considerados e que interferem no progn\u00f3stico dos pacientes em rela\u00e7\u00e3o ao crescimento do aneurisma da aorta, como sua idade, comorbidades e uso de medicamentos. O \u00e2ngulo entre a entrada da aorta e o tronco braquiocef\u00e1lico pode ser uma importante vari\u00e1vel a ser considerada, assim como a estrutura do escoamento e a distribui\u00e7\u00e3o da press\u00e3o na parede anterior da aorta, que pode ser numericamente determinada.A partir da distribui\u00e7\u00e3o de press\u00e3o na parede da aorta, \u00e9 poss\u00edvel observar que a press\u00e3o m\u00e9dia aumentou mais de 13% para todos os pacientes que tiveram crescimento do aneurisma, enquanto, para aqueles sem crescimento patol\u00f3gico, a press\u00e3o m\u00e9dia decresceu mais de 18% para todos os pacientes, com exce\u00e7\u00e3o de um. Esse resultado indica que essa vari\u00e1vel pode influenciar no crescimento do AAAs.Estudos futuros com maior n\u00famero de pacientes e maior tempo de acompanhamento s\u00e3o necess\u00e1rios para definir a rela\u00e7\u00e3o entre padr\u00e3o de fluxo sangu\u00edneo e processo de remodelamento a\u00f3rtico. A simula\u00e7\u00e3o num\u00e9rica \u00e9 uma ferramenta n\u00e3o invasiva que pode ser utilizada juntamente com a varredura de ATC para estimar a tend\u00eancia de crescimento do AAAs. A identifica\u00e7\u00e3o de diferentes perfis de risco entre pacientes com AAAs pode abrir caminho para avalia\u00e7\u00e3o m\u00e9dica diferenciada para esses pacientes. Todo o exposto pode levar a um avan\u00e7o na medicina personalizada e ajudar a decifrar a fisiopatologia dessa grave doen\u00e7a.Considerando que algumas caracter\u00edsticas do escoamento podem estar associadas ao crescimento do aneurisma, o estudo de padr\u00f5es de fluxo na aorta ascendente pode ajudar a predizer o processo de remodelamento de AAAs.Este estudo piloto mostrou que a CFD aplicada em ATC pode, em um futuro pr\u00f3ximo, ser uma ferramenta para ajudar a identificar padr\u00f5es de comportamento do escoamento associados com o processo de remodelamento de AAAs. V\u00f3rtices foram formados na regi\u00e3o posterior do jato incidente na parede a\u00f3rtica, gerando estruturas complexas no grupo com crescimento do AAAs. Como consequ\u00eancia, houve um aumento da press\u00e3o m\u00e9dia na parede anterior da aorta no grupo que apresentou crescimento do aneurisma, enquanto, para os casos sem crescimento do aneurisma, essa vari\u00e1vel decresceu. 2 However, its complications, such as rupture and dissection, are catastrophic events.3 Some aneurysms do not grow, while others increase significantly in size in a short period. The variables related to the AsAA growth are not known precisely.4Ascending aortic aneurysm (AsAA) is usually asymptomatic and progresses imperceptibly.5 According to Hope et al.,4 changes in the flow pattern along the ascending aorta are related to the remodeling process, and, thus, may influence aneurysm growth.The AsAA formation is a multifactorial degenerative process, resulting from hemodynamic factors and biological processes.7 As highlighted by Morris et al.7 CFD combined with population-scale numerical models have the potential to reduce the risks, costs, and time associated with clinical trials.Computational Fluid Dynamics (CFD) has gained great interest as a complementary tool to improve the understanding of cardiovascular disease pathogenesis and progression, in addition to proving itself suitable for minimally invasive assessments.8 suggesting an association of these variables with aneurysmal dilatation.9 These observations were discussed by G\u00fclan et al.,10 who found large rotational regions formed during the systolic phase, showing qualitative similarity with the vortex ring formation, i.e., a central jet surrounded by two large vortices. From the fluid mechanics viewpoint, the flow through an abrupt expansion, such as that between the aortic annulus and the dilated ascending portion, leads to separation of the boundary layer and formation of a separation zone. Hence, there might be a correlation between flow separation, turbulence, pressure loss, and aneurysm growth.10Through CFD studies of aortic flow, regions with high rates of wall shear stress and pressure can be identified,11In addition, the presence of coherent structures inside the aorta can be applied to explain the formation of high shear stress and pressure regions, which can contribute to the aortic remodeling process.The present work is an analytical, observational, single-center study, in which a CFD protocol was applied to aortic computed tomography angiogram (CTA) images. This study was registered in the National Research Ethics Committee of the Health Ministry and approved by the local Research Ethics Committee . All participants provided written informed consent in accordance with Resolution 466/2012 of the National Health Board.12This study assessed nine patients with AsAA selected from 100 consecutive patients from the aortic disease outpatient clinic of a tertiary health center. Patients with AsAA diagnosis were included, and all of them had stable disease. Patients with collagen pathologies and previous aortic and cardiac surgery were excluded. All patients had two available images of CTA scan with a minimum interval of one-year between the exams. The images were obtained due to clinical indications for follow-up, not specifically for the present study. Patients were under medical treatment according to recent guidelines on AsAA.13The aorta geometry was built based on a CTA scan obtained with a 64-slice scanner SOMATOM Sensation 64 . The selected CTA slices were spanned from the aortic annulus to the thoracic aorta. Information, such as pixel size and distance slice, was obtained to further adjust the three-dimensional (3D) aortic model to its actual size. The image segmentation was performed with the software FIJI, an open-source image processing software based on ImageJ, focused on biological-image analysis.Two 3D aortic models for each patient were super-imposed to provide a spatial reference for comparison between the exams, aiming to overlap the images of the beginning of the brachiocephalic trunk and the right coronary arteries . The ao\u03b8I and \u03b8II ) were defined between the inflow plane and the brachiocephalic trunk outlet, aiming to relate it with the flow pattern that can induce aneurysm growth .21 In addition, under normal conditions, blood can be considered an incompressible fluid.22 Density was set at \u03c1 = 1054 kg/m3The dynamic viscosity was defined as \u03bc = 7.2 cP to allow a direct comparison with in vitro experimental data.23 Additionally, a few tests were performed and the results obtained with viscosities equal to 3.5 cP and 7.2 cP provided equivalent results, with pressure differences inferior to 0.01 mm Hg in the region of interest of the ascending aorta. These tests corroborate the findings by Becsek et al.,24 who evaluated viscosities equal to 4 cP, 6 cP, and 8 cP and showed negligible effect on the mean flow field and a very small effect on the location of the turbulent breakdown.Blood was modeled as a Newtonian fluid since high shear rates15 as shown in For all cases, the same outlet flow-rate distributions (inflow percentage) were imposed based on average values in the human body, according to Alastruey et al.,25 Simulations were post-processed with the ANSYS CFD-Post tool26 and with the open-source Paraview.27The present analyses were performed using ANSYS Fluent v18.1 software, which solves the conservation equations discretized based on the finite volume method.25 The pressure-velocity coupling was handled with the SIMPLE algorithm,25 and the maximum convergence residual error was set as 10-6for all equations.The discretized equations were obtained with the \u201cPower Law scheme\u201d.A mesh with 400 000 nodes was applied to all cases studied, which were determined based on a mesh independent test, by guaranteeing a pressure drop variation smaller than 0.3% at the main area of interest (ascending aorta), when the mesh size was doubled.\u03b8I and \u03b8II ( \u03b8I and \u03b8II measurements. The following criteria for reliability28 was applied: poor (ICC \u2264 0.50); moderate (0.50 < ICC \u2264 0.75); good (0.75 < ICC \u2264 0.90); excellent (ICC > 0.90).The anatomic shape of each patient\u2019s aorta can be inferred by the reference angles \u03b8II , which \u03b8I and \u03b8II measurements. Statistical significance was set at 5%. All statistical analyses were conducted using IBM SPSS software, version 26.29The Kolmogorov-Smirnov normality test was used to examine if variables had a normal distribution. Due to non-normal distribution, the Wilcoxon signed-rank test, a non-parametric test, was applied for comparison of paired data (dependent samples) between the 30 The aneurysm was considered as having grown when the difference between the volume of the area of interest increased by 10% or more, comparing the two CTAs for each patient. Patients were divided into two groups based on the area of interest volume.As shown in \u03b8I\u2265 94o) and small Angle II (51o\u2264 \u03b8II\u2264 68o). However, when examining the angles corresponding to the patients without aneurysm growth, a clear tendency was not observed. There were patients with large and small \u03b8I and \u03b8II . Patients 6 and 7 clearly presented a different range of angles, but the same was not true for patients 8 and 9. The \u03b8I measurement in patients with aneurysm growth was 127.47 [116.64 \u2013 135.79]\u00b0; without aneurysm growth, 119.22 [91.26 \u2013 128.97]\u00b0; and in total population, 121.53 [94.9 \u2013 135.79]\u00b0. The \u03b8II measurement in patients with aneurysm growth was 51.46 [46.07 \u2013 56.61]\u00b0; without aneurysm growth, 68.5 [61.53 \u2013 75.41]\u00b0; and in the total population, 57.94 [51.34 \u2013 67.86]\u00b0.w ) equal to 50% of inlet axial velocity ( win ) was drawn were also included, as well as the percent area of the ascending aorta under high pressure ( %Acr ). Analyzing these figures, one can associate the inlet jet, shown in Notably, there is a tendency towards larger areas of increased pressures in the growing aneurysm group, with elevated maximum pressures above 200 Pa, compared to the non-growing aneurysm group, which demonstrated lower maximum pressures (below 200 Pa), except for patient 9 in the non-growing aneurysm group, who presented a significant reduction in the average pressure, which might explain the lack of aneurysm growth. Further, there was an increase in the average wall pressure in the growing aneurysm group and a reduction in the other group, except for patient 7, whose maximum pressure was quite low, which could also explain the lack of aneurysm growth.The clinical profile of the population studied is shown in 15 for possible clinical applications in the near future. In this study, patients were classified as AsAA with growth or without growth according to the volume of the area of interest. Raghavan et al.30 have shown a better relationship of possible aneurysm rupture with the ascending aorta volume and high wall stresses, rather than the ascending aorta diameter.Hemodynamics simulation has been a promising area of translational scientific research\u03b8I and small \u03b8II may indicate the possibility of an aortic aneurysm remodeling process.Biasetti et al.39One hypothesis to explain these findings is the transfer of stress to adventitia due to loss of elastin and collagen deposition, promoting continuous adaptation because of stress distribution and, consequently, changes in the shape of AsAA.There are some limitations to this research. Firstly, the CTA scans were performed by different physicians. As a result, the quality of the exams was not the same. This fact may have affected the accuracy of measurements and definition of the aortic shape. Secondly, the small size of the population studied limits any correlation between clinical patterns and AsAA growth. Thirdly, clinical data of two patients were not available, so they were not included in Clearly, there are several factors that must be considered to improve the prognosis of patients with regards to aortic aneurysm growth, such as their age, previous diseases, and medicine intake. The angle between the aortic inlet and the brachiocephalic trunk could be an important feature to be considered, as well as the flow structure and the pressure distribution on the anterior wall, which can be numerically determined.By the pressure distribution in the aortic wall, the mean pressure was observed to increase over 13% in all patients with aneurysm growth, while for those without pathological growth, the mean pressure decreased over 18% for all patients, except for one. This result indicates that this variable might influence the growth of the AsAA.Future studies with a larger number of patients and longer follow-up are needed to precisely define the relationship between blood flow pattern and aortic remodeling process. The CFD could be a completely non-invasive tool to be used from CTA scans to estimate the growth trend of AsAA. The identification of different risk profiles among patients with AsAA could open the way for different medical care for these patients. All the above can lead to a step forward in personalized medicine and help to decipher the pathophysiology of this severe disease.Considering that some flow characteristics could be associated with aneurysm growth, the study of the flow pattern in the ascending aorta could help predict the remodeling process of AsAA.This pilot study showed that CFD based on CTA may in the near future be a tool to help identify patterns of flow behavior associated with the AsAA remodeling process. Vortices were formed in the posterior region of the incident jet in the aortic wall, generating more complex structures for the aneurysm growth group. Consequently, there was an increase in the mean pressure in the anterior aortic wall between the CTA exams for the patients with aneurysm growth, while for those without aneurysm growth, this variable decreased."} +{"text": "Nos \u00faltimos anos, foram consolidados avan\u00e7os expressivos no conhecimento de amiloidose card\u00edaca (AC), trazendo uma profunda reformula\u00e7\u00e3o do seu significado cl\u00ednico. Al\u00e9m de haver evid\u00eancias convincentes de que AC seja uma causa relativamente comum de insufici\u00eancia card\u00edaca com fra\u00e7\u00e3o de eje\u00e7\u00e3o preservada (ICFEP), assistimos o surgimento de terapias espec\u00edficas modificadoras do curso natural da doen\u00e7a, capazes de prolongar a sobrevida dos pacientes acometidos.Em paralelo, relevantes progressos nas t\u00e9cnicas de imagem cardiovascular t\u00eam contribu\u00eddo grandemente para o reconhecimento mais acurado e precoce da doen\u00e7a. Particularmente, o emprego da cintilografia card\u00edaca com radiotra\u00e7adores \u00f3sseos tornou poss\u00edvel o diagn\u00f3stico n\u00e3o invasivo da AC ligada \u00e0 transtirretina (TTR), prescindindo da bi\u00f3psia endomioc\u00e1rdica, o que simplificou muito o fluxo diagn\u00f3stico.Assim, o presente posicionamento tem por objetivo apresentar as recomenda\u00e7\u00f5es mais atuais para o diagn\u00f3stico, estadiamento progn\u00f3stico e tratamento da AC, com base na revis\u00e3o cr\u00edtica das evid\u00eancias cient\u00edficas atuais.Neste posicionamento, as tabelas de classes de recomenda\u00e7\u00e3o e n\u00edveis de evid\u00eancia foram realizadas conforme a padroniza\u00e7\u00e3o a seguir.>wild type (ATTRwt) quanto heredit\u00e1ria ou variante (ATTRv).\u2013A amiloidose sist\u00eamica \u00e9 uma doen\u00e7a causada pela deposi\u00e7\u00e3o tecidual de agregados proteicos fibrilares e insol\u00faveis em diferentes \u00f3rg\u00e3os, incluindo o cora\u00e7\u00e3o, levando \u00e0 disfun\u00e7\u00e3o org\u00e2nica.A transtirretina (TTR) \u00e9 uma prote\u00edna composta por quatro mon\u00f4meros, que circulam como um tetr\u00e2mero.Na ATTRwt, a sequ\u00eancia de amino\u00e1cidos \u00e9 normal e n\u00e3o est\u00e1 completamente esclarecido o processo pelo qual a prote\u00edna selvagem se torna inst\u00e1vel e se agrega em fibrilas amiloides. No entanto, o envelhecimento parece estar envolvido na fisiopatologia da doen\u00e7a.Na forma AL, a cadeia leve amiloidog\u00eanica origina-se de plasm\u00f3citos ou, menos frequentemente, linf\u00f3citos B an\u00f4malos, configurando, portanto, uma doen\u00e7a hematol\u00f3gica clonal e neopl\u00e1sica. No cora\u00e7\u00e3o, o dep\u00f3sito de fibrilas amiloides causa dano estrutural ao aumentar a rigidez card\u00edaca e vascular, prejudicando a contra\u00e7\u00e3o e o relaxamento card\u00edaco e gerando dist\u00farbios de condu\u00e7\u00e3o. Em paralelo, as cadeias leves circulantes tamb\u00e9m apresentam toxicidade direta ao mioc\u00e1rdio, atrav\u00e9s de disfun\u00e7\u00e3o lisossomal, autofagia defeituosa, produ\u00e7\u00e3o de esp\u00e9cies reativas de oxig\u00eanio, disfun\u00e7\u00e3o celular e mitocondrial, altera\u00e7\u00f5es na homeostase do c\u00e1lcio do cardiomi\u00f3cito e, por fim, morte celular.A \u2013Diferentes subtipos de amiloidose podem originar manifesta\u00e7\u00f5es cl\u00ednicas sobrepostas e, uma vez diagnosticada a amiloidose, \u00e9 imprescind\u00edvel caracterizar corretamente a prote\u00edna precursora para institui\u00e7\u00e3o do tratamento espec\u00edfico.A depender dos \u00f3rg\u00e3os acometidos e do grau de disfun\u00e7\u00e3o ocasionada, um amplo espectro de manifesta\u00e7\u00f5es cl\u00ednicas pode ser observado, com evolu\u00e7\u00e3o progressiva e potencialmente fatal. Os principais \u00f3rg\u00e3os que podem ser acometidos na amiloidose sist\u00eamica s\u00e3o cora\u00e7\u00e3o, rins, olhos, sistema nervoso central e perif\u00e9rico e f\u00edgado. Manifesta\u00e7\u00f5es cl\u00ednicas inespec\u00edficas s\u00e3o frequentemente observadas e incluem fadiga, perda ponderal, edema perif\u00e9rico e hipotens\u00e3o ortost\u00e1tica. Por esse motivo, \u00e9 comum o diagn\u00f3stico tardio, de modo que s\u00e3o necess\u00e1rios conhecimento sobre a doen\u00e7a e elevado grau de suspei\u00e7\u00e3o cl\u00ednica para conclus\u00e3o diagn\u00f3stica.t, a cardiopatia \u00e9 a principal manifesta\u00e7\u00e3o cl\u00ednica, ocorrendo principalmente em homens idosos que desenvolvem ICFEP sem fatores de risco previamente conhecidos.Na ATTRv, a depender da muta\u00e7\u00e3o, o quadro cl\u00ednico \u00e9 dominado por neuropatia ou cardiopatia. Na ATTRwAlgumas altera\u00e7\u00f5es extracard\u00edacas podem anteceder, emanos, o desenvolvimento da AC, destacando-se a s\u00edndrome do t\u00fanel do carpo bilateral e a ruptura espont\u00e2nea do tend\u00e3o do b\u00edceps. O reconhecimento de tais sinais como parte do quadro cl\u00ednico de amiloidose \u00e9 fundamental, podendo levar a diagn\u00f3stico mais precoce e evitar a progress\u00e3o da cardiopatia pela institui\u00e7\u00e3o de tratamento espec\u00edfico.A ATTRwt tem incid\u00eancia aumentada em pacientes mais idosos, usualmente acima de 70 anos de idade; contudo, as manifesta\u00e7\u00f5es cl\u00ednicas da ATTRv tamb\u00e9m costumam ocorrer em idosos, fazendo com que a idade n\u00e3o deva ser levada em considera\u00e7\u00e3o para diferencia\u00e7\u00e3o entre as duas formas de ATTR. No que diz respeito ao sexo, h\u00e1 uma forte predomin\u00e2ncia no sexo masculino, em 80% a 90%, apenas na ATTRwt.,,,Com rela\u00e7\u00e3o \u00e0 ATTRv, a muta\u00e7\u00e3o V30M \u00e9 a mais disseminada pelo mundo, sendo end\u00eamica em Portugal, Su\u00e9cia e Jap\u00e3o e, provavelmente, a mais comum no Brasil. Outra muta\u00e7\u00e3o bastante frequente \u00e9 a V122I, que est\u00e1 presente em 3,4% dos afro-americanos e relacionada ao desenvolvimento de cardiopatia em pacientes acima dos 60 anos de idade. t.A A amiloidose card\u00edaca por cadeias leves (AL) apresenta incid\u00eancia de 6 a 10/milh\u00e3o de pessoas/ano e era considerada a principal causa de AC.Muito mais que uma doen\u00e7a rara, na verdade, a AC \u00e9 uma condi\u00e7\u00e3o subdiagnosticada. Dados recentes nos EUA registram o progressivo aumento na preval\u00eancia, aumentando de 18 para 55,2 (100.000 pessoas-ano),wild type \u00e9 de 3,6 anos e, na ATTRv, o progn\u00f3stico depende da muta\u00e7\u00e3o. Nos casos de fen\u00f3tipo neurol\u00f3gico, a progress\u00e3o da neuropatia leva \u00e0 incapacidade sensitivo-motora, mas a mortalidade \u00e9 mais relacionada a comprometimento card\u00edaco.Com rela\u00e7\u00e3o ao progn\u00f3stico, a amiloidose AL causa envolvimento multiorg\u00e2nico, de car\u00e1ter mais agressivo em rela\u00e7\u00e3o aos demais subtipos. O diagn\u00f3stico tardio ainda faz com que a mortalidade precoce nos primeiros 6 a 12 meses seja elevada em decorr\u00eancia de complica\u00e7\u00f5es da cardiopatia avan\u00e7ada.Muta\u00e7\u00f5es no gene TTR est\u00e3o associadas a uma grande variedade de manifesta\u00e7\u00f5es cl\u00ednicas, que refletem o dep\u00f3sito da prote\u00edna variante (TTRv) em diferentes tipos de tecidos, sendo o envolvimento card\u00edaco e o do sistema nervoso perif\u00e9rico os mais frequentes; o primeiro particularmente causado pela muta\u00e7\u00e3o V122I e o segundo, pela muta\u00e7\u00e3o V30M.Neste cap\u00edtulo, descreveremos as principais manifesta\u00e7\u00f5es neurol\u00f3gicas que deveriam levantar a possibilidade de ATTRv.As manifesta\u00e7\u00f5es neurol\u00f3gicas na ATTRv podem se dividir em: neuropatia perif\u00e9rica, manifesta\u00e7\u00f5es tardias de acometimento do sistema nervoso central (SNC) ligadas \u00e0 angiopatia amiloide e manifesta\u00e7\u00f5es do SNC associadas \u00e0 infiltra\u00e7\u00e3o oculomen\u00edngea.A forma cl\u00e1ssica de acometimento dos nervos perif\u00e9ricos na ATTRv \u00e9 a polineuropatia axonal, auton\u00f4mica-sensitiva-motora de evolu\u00e7\u00e3o comprimento dependente, ou seja: afeta inicialmente segmentos mais distais dos membros, sobretudo os inferiores, e evolui para o acometimento de segmentos proximais e para os membros superiores.Na forma de in\u00edcio precoce (< 50 anos), em geral associada \u00e0 muta\u00e7\u00e3o ATTRv V30M (V50M), as fibras finas, pouco ou n\u00e3o mielinizadas s\u00e3o as acometidas inicialmente, seguindo-se, \u00e0 medida que a doen\u00e7a progride, comprometimento das fibras grossas, muito mielinizadas, respons\u00e1veis pelas sensibilidades vibrat\u00f3ria, cin\u00e9tico-postural e pela motricidade. Assim, os primeiros sintomas s\u00e3o disfun\u00e7\u00e3o er\u00e9til, saciedade precoce, n\u00e1useas, v\u00f4mitos, diarreia, constipa\u00e7\u00e3o, altern\u00e2ncia de diarreia com constipa\u00e7\u00e3o, hipotens\u00e3o ortost\u00e1tica, s\u00edncopes, arritmias, altera\u00e7\u00e3o da condu\u00e7\u00e3o atrioventricular, olho seco, reten\u00e7\u00e3o ou incontin\u00eancia urin\u00e1ria, dor neurop\u00e1tica, perda das sensibilidades ao calor e ao frio e uma importante perda ponderal. J\u00e1 nessa fase inicial, podem surgir les\u00f5es indolores, mal perfurante plantar e suas repercuss\u00f5es, tais como infec\u00e7\u00f5es localizadas, celulite, osteomielite e at\u00e9 septicemia. Ap\u00f3s alguns anos, surgem instabilidade \u00e0 marcha e fraqueza por atrofia muscular, sempre evoluindo de distal para proximal.J\u00e1 nas formas tardias, a neuropatia, desde o in\u00edcio, compromete todos os tipos de fibras, e a disautonomia n\u00e3o \u00e9 t\u00e3o importante, pelo menos na fase inicial da doen\u00e7a. Estas formas tanto podem estar associadas \u00e0 muta\u00e7\u00e3o TTRv V30M quanto a v\u00e1rias outras muta\u00e7\u00f5es, e a evolu\u00e7\u00e3o costuma ser mais agressiva. Em estudo brasileiro, 26 % dos pacientes com ATTRv V30M tiveram in\u00edcio tardio.A s\u00edndrome do t\u00fanel do carpo bilateral \u00e9 uma manifesta\u00e7\u00e3o frequente na ATTRv, podendo ser a manifesta\u00e7\u00e3o inicial. Est\u00e1 associada a qualquer muta\u00e7\u00e3o, mas \u00e9 particularmente importante em algumas delas, incluindo a TTR V122I, que aparenta ser particularmente frequente no Brasil, associada \u00e0 doen\u00e7a card\u00edaca, a quem pode preceder por v\u00e1rios anos.stroke-like, TIA-like, ou aura-like, quanto do tipo irritativo epil\u00e9tico. Nos casos mais graves, pode se instalar isquemia ou mesmo hemorragia intracraniana. Dentre outras manifesta\u00e7\u00f5es neurol\u00f3gicas, destacam-se: comprometimento auditivo; migr\u00e2nia; dem\u00eancia; s\u00edndrome cerebelar; mielopatia e radiculopatia.O prolongamento da sobrevida, inicialmente associada ao transplante hep\u00e1tico e agora pelos novos medicamentos, tem possibilitado o aparecimento de manifesta\u00e7\u00e3o que antes eram incomuns. A produ\u00e7\u00e3o de TTR pelo plexo coroide , a longo prazo, tanto est\u00e1 associada a uma angiopatia amiloide como \u00e0 infiltra\u00e7\u00e3o men\u00edngea. A angiopatia amiloide se manifesta como epis\u00f3dios focais tanto do tipo deficit\u00e1rio Algumas muta\u00e7\u00f5es raras t\u00eam predile\u00e7\u00e3o pelo acometimento oculocerebral e constituem quadros de amiloidose oculoleptomen\u00edngea .Est\u00e1gios da neuropatia: Os est\u00e1gios de Coutinho para ATTRv s\u00e3o demarcados pela polineuropatia. O est\u00e1gio 1 \u00e9 aquele em que a polineuropatia sensitivo-motora afeta a marcha, mas o paciente n\u00e3o necessita de apoio para deambular. No est\u00e1gio 2, um ou mais apoios s\u00e3o necess\u00e1rios para a deambula\u00e7\u00e3o. No est\u00e1gio 3, o paciente est\u00e1 restrito \u00e0 cadeira de rodas ou ao leito.A ATTRv \u00e9 uma doen\u00e7a de heran\u00e7a autoss\u00f4mica dominante, mas que apresenta penetr\u00e2ncia vari\u00e1vel, que \u00e9 ao mesmo tempo muta\u00e7\u00e3o dependente, idade dependente e tamb\u00e9m sofre influ\u00eancia regional.Pelo menos 140 diferentes muta\u00e7\u00f5es foram descritas at\u00e9 o momento, mas nem todas s\u00e3o patog\u00eanicas. Algumas s\u00e3o polimorfismos j\u00e1 bem determinados, enquanto outras ainda t\u00eam significado indeterminado.Um cap\u00edtulo especial dentre os aspectos gen\u00e9ticos \u00e9 o teste pr\u00e9-sintom\u00e1tico, ou seja, o teste de parentes de pessoas sabidamente afetadas. Diferentemente do teste diagn\u00f3stico, ele deve ser feito por pessoas treinadas e deve haver uma equipe de suporte, incluindo psic\u00f3logo. Deve incluir obrigatoriamente uma fase de prepara\u00e7\u00e3o, pr\u00e9-diagn\u00f3stica, o teste gen\u00e9tico e uma fase de suporte, p\u00f3s-resultado. N\u00e3o deve ser realizado em crian\u00e7as, e somente deve ser aplicado quando o candidato manifestar expressamente que esta \u00e9 sua vontade e for considerado psicologicamente preparado.\u2013A AC cursa com infiltra\u00e7\u00e3o da matriz extracelular card\u00edaca por fibrilas amiloides, resultando em um aumento progressivo da espessura da parede ventricular e aumento acentuado na rigidez da c\u00e2mara, acarretando em comprometimento da fun\u00e7\u00e3o diast\u00f3lica, levando \u00e0 insufici\u00eancia card\u00edaca com fisiologia restritiva.Dessa forma, a manifesta\u00e7\u00e3o cl\u00ednica mais frequente \u00e9 a s\u00edndrome de insufici\u00eancia card\u00edaca, mais comumente com fra\u00e7\u00e3o de eje\u00e7\u00e3o preservada (ICFEP), mas que pode evoluir com queda da fra\u00e7\u00e3o de eje\u00e7\u00e3o nas fases mais avan\u00e7adas da doen\u00e7a. A s\u00edndrome cl\u00ednica pode se apresentar com sintomas predominantes de IC esquerda, com congest\u00e3o pulmonar , ou sintomas de IC direita ou ambos os conjuntos de sintomas. A amiloidose card\u00edaca deve ser considerada no diagn\u00f3stico diferencial da etiologia da ICFEP em homens idosos,S\u00edncope e hipotens\u00e3o ortost\u00e1tica s\u00e3o sintomas comuns e indicam a presen\u00e7a de disautonomia. Um aspecto cl\u00ednico caracter\u00edstico que pode levantar a suspeita de amiloidose \u00e9 a necessidade de reduzir a dose ou descontinuar medicamentos anti-hipertensivos em pacientes com diagn\u00f3stico pr\u00e9vio de HAS, principalmente betabloqueadores e inibidores da enzima conversora da angiotensina/bloqueadores dos receptores de angiotensina.Pode ainda ocorrer infiltra\u00e7\u00e3o amiloide causando doen\u00e7a do sistema de condu\u00e7\u00e3o card\u00edaco desde as fases precoces da doen\u00e7a, com graus vari\u00e1veis de bloqueio atrioventricular, resultando, em alguns casos, em bradicardia de alto risco, com necessidade de implante de marca-passo. Outra altera\u00e7\u00e3o importante \u00e9 decorrente do endurecimento das paredes atriais, com taxas elevadas de arritmias atriais, incluindo a fibrila\u00e7\u00e3o atrial, e presen\u00e7a de trombos atriais, sendo o acidente vascular encef\u00e1lico cardioemb\u00f3lico uma manifesta\u00e7\u00e3o cl\u00ednica comum, mesmo em indiv\u00edduos com ritmo sinusal. Arritmias ventriculares complexas parecem ser frequentes nas fases avan\u00e7adas da doen\u00e7a, aspecto mais bem documentado na amiloidose AL.A amiloidose card\u00edaca, particularmente na forma ATTR, \u00e9 frequentemente subdiagnosticada por raz\u00f5es associadas \u00e0 avalia\u00e7\u00e3o m\u00e9dica e por caracter\u00edsticas da pr\u00f3pria doen\u00e7a, incluindo: conhecimento fragmentado entre diferentes especialidades e subespecialidades, escassez de centros e especialistas dedicados ao manejo dessa doen\u00e7a, conhecimento equivocado acreditando-se ser uma doen\u00e7a rara e incur\u00e1vel, heterogeneidade fenot\u00edpica e genot\u00edpica nas formas ATTR.Dessa forma, o reconhecimento de \u201csinais de alerta\u201d pode auxiliar na presun\u00e7\u00e3o do diagn\u00f3stico da amiloidose card\u00edaca nos pacientes com IC,S\u00edndrome do t\u00fanel do carpo bilateral \u00e9 muitas vezes um dos primeiros indicadores de ATTR, \u00e9 a manifesta\u00e7\u00e3o n\u00e3o card\u00edaca mais comum e pode preceder os sintomas de insufici\u00eancia card\u00edaca em v\u00e1rios anos. Um estudo recente observou que aproximadamente 50% dos indiv\u00edduos com ATTRwt apresentava s\u00edndrome do t\u00fanel do carpo 5 a 7 anos antes do diagn\u00f3stico.Outros sinais de alerta para a doen\u00e7a podem emergir de altera\u00e7\u00f5es t\u00edpicas nos exames complementares cardiol\u00f3gicos de rotina, e s\u00e3o abordadas em t\u00f3picos espec\u00edficos deste documento.Vale salientar, ainda, que a amiloidose card\u00edaca pode muitas vezes apresentar-se simulando outras cardiopatias. A amiloidose deve ser considerada como uma das poss\u00edveis etiologias de pacientes que apresentam fen\u00f3tipo de cardiomiopatia hipertr\u00f3fica, particularmente se iniciada em fases tardias da vida (> 60 anos). O padr\u00e3o assim\u00e9trico de hipertrofia mioc\u00e1rdica (HVE) nos pacientes com amiloidose ATTR difere dos pacientes com forma AL, geralmente sim\u00e9trico. Em estudo de 263 pacientes com AC por ATTR confirmada e comparados com 50 pacientes com a forma AL, observou-se na forma ATTR presen\u00e7a de hipertrofia assim\u00e9trica em 79% dos casos, sim\u00e9trica em 18% e 3% sem HVE.Pacientes idosos portadores de estenose a\u00f3rtica grave de baixo fluxo e baixo gradiente podem exibir AC em at\u00e9 10% a 15% dos casos, com progn\u00f3stico desfavor\u00e1vel.O eletrocardiograma (ECG) \u00e9 um exame essencial na avalia\u00e7\u00e3o diagn\u00f3stica e no planejamento terap\u00eautico dos pacientes, sendo importante sua interpreta\u00e7\u00e3o em conjunto com as informa\u00e7\u00f5es cl\u00ednicas e ecocardiogr\u00e1ficas. Embora o achado de sinais de baixa voltagem ao ECG tenha grande especificidade no diagn\u00f3stico de infiltra\u00e7\u00e3o mioc\u00e1rdica secund\u00e1ria \u00e0 AC, este n\u00e3o \u00e9 o achado mais prevalente na doen\u00e7a. A aus\u00eancia de progress\u00e3o de ondas R em deriva\u00e7\u00f5es precordiais, simulando uma zona el\u00e9trica inativa anterosseptal (padr\u00e3o de pseudoinfarto) \u00e9 achado muito mais frequente, chegando \u00e0 preval\u00eancia de 60% a 70% dos casos com diagn\u00f3stico confirmado, independentemente do tipo de amiloidose .Nos casos de AC por ATTR, menos de 40% dos pacientes com diagn\u00f3stico confirmado por bi\u00f3psia apresentam sinais de baixa voltagem ao ECG.Assim, a aus\u00eancia de crit\u00e9rios de baixa voltagem ao ECG, ou mesmo a presen\u00e7a de sinais de sobrecarga de VE, n\u00e3o deve afastar a suspeita diagn\u00f3stica de AC, especialmente a ATTR. A despropor\u00e7\u00e3o da voltagem em rela\u00e7\u00e3o \u00e0 espessura mioc\u00e1rdica tamb\u00e9m \u00e9 sinal de alerta importante e alcan\u00e7a preval\u00eancia de 73% a 80% em pacientes com AC, independentemente de seu tipo.Dentre as altera\u00e7\u00f5es do ritmo card\u00edaco, a fibrila\u00e7\u00e3o atrial \u00e9 mais prevalente em pacientes com ATTR, assim como os bloqueios atrioventriculares.granular sparkling) n\u00e3o s\u00e3o aumentadas, os volumes s\u00e3o normais ou reduzidos e h\u00e1 aumento da espessura de paredes ventriculares. O aumento de dimens\u00f5es atriais \u00e9 comum, refletindo uma disfun\u00e7\u00e3o diast\u00f3lica precoce e progressiva, com aumento das press\u00f5es de enchimento. As valvas atrioventriculares podem estar espessadas e as regurgita\u00e7\u00f5es valvares s\u00e3o funcionais. Pode, ainda, haver sinais sugestivos de infiltra\u00e7\u00e3o do septo interatrial e tamb\u00e9m eleva\u00e7\u00e3o da press\u00e3o sist\u00f3lica da art\u00e9ria pulmonar. O ventr\u00edculo direito tamb\u00e9m pode estar acometido. \u00c9 muito comum a presen\u00e7a de derrames pleurais e peric\u00e1rdicos e, em casos em que h\u00e1 intensa infiltra\u00e7\u00e3o tecidual, \u00e9 poss\u00edvel observar o cl\u00e1ssico, embora subjetivo, aspecto granular da imagem de paredes mioc\u00e1rdicas (arkling) 35,54Na AC, a fra\u00e7\u00e3o de eje\u00e7\u00e3o do VE (FEVE) permanece normalmente preservada at\u00e9 os est\u00e1gios mais avan\u00e7ados da doen\u00e7a, mas a fun\u00e7\u00e3o contr\u00e1til longitudinal \u00e9 reduzida precocemente.A avalia\u00e7\u00e3o do padr\u00e3o de enchimento diast\u00f3lico dos ventr\u00edculos \u00e9 essencial e frequentemente demonstra algum grau de disfun\u00e7\u00e3o diast\u00f3lica. Nas fases iniciais, \u00e9 poss\u00edvel observar altera\u00e7\u00f5es compat\u00edveis com disfun\u00e7\u00e3o diast\u00f3lica tipo I .Com a progress\u00e3o da doen\u00e7a, pode se instalar padr\u00e3o pseudonormal de disfun\u00e7\u00e3o diast\u00f3lica por consequ\u00eancia da eleva\u00e7\u00e3o da press\u00e3o no \u00e1trio esquerdo.Na fase mais adiantada da doen\u00e7a, observa-se o padr\u00e3o restritivo do enchimento ventricular .\u2013apical sparing,bulls eye disp\u00f5e de t\u00e9cnicas que permitem avaliar de forma precisa as altera\u00e7\u00f5es teciduais mioc\u00e1rdicas que ocorrem na AC.O aumento global de \u00e1gua tecidual mioc\u00e1rdica leva a aumento dos tempos m\u00e9dios de relaxamento do hidrog\u00eanio, sejam eles T1 ou T2 .\u2013Assim, o conjunto do dep\u00f3sito de miofibrilas e fibrose intersticial pode ser facilmente detectado de forma muito precisa, e at\u00e9 mesmo quantificado pelas t\u00e9cnicas de realce tardio (RT)O contraste utilizado na RMC baseia-se em gadol\u00ednio que, por sua vez, \u00e9 ligado a um quelante macromolecular que n\u00e3o permite sua passagem pela membrana celular \u00edntegra, distribuindo-se assim exclusivamente no VEC do mioc\u00e1rdio. A imagem de RT permite levantar a suspeita de AC pelo seu padr\u00e3o de distribui\u00e7\u00e3o .A AC pode modificar a apar\u00eancia de todas as c\u00e2maras card\u00edacas.strain).A AC tamb\u00e9m \u00e9 habitualmente associada \u00e0 presen\u00e7a de aumento da espessura mioc\u00e1rdica, que \u00e9, na maior parte das vezes, mais expressiva do que nos casos secund\u00e1rios \u00e0 hipertens\u00e3o, e a espessura do m\u00fasculo card\u00edaco geralmente \u00e9 maior nos casos de ATTR do que nos casos AL.Em consequ\u00eancia da disfun\u00e7\u00e3o diast\u00f3lica, n\u00e3o \u00e9 raro observar derrame peric\u00e1rdico ou pleural.A t\u00e9cnica de RT ap\u00f3s inje\u00e7\u00e3o do contraste gadol\u00ednio tem sido amplamente reconhecida como um dos pilares no diagn\u00f3stico por imagem da AC.Em s\u00e9rie com 250 pacientes com amiloidose de distintas formas, a presen\u00e7a de RT com padr\u00e3o transmural esteve associada a um risco de morte 5,4 vezes maior .,,,,,,,Diferentes grupos t\u00eam investigado a utilidade de mapas de T1 derivados da RMC para melhorar o desempenho diagn\u00f3stico e progn\u00f3stico na AC.\u2013Os radiotra\u00e7adores marcados com tecn\u00e9cio-99m derivados do bifosfonato, originalmente desenvolvidos para imagens \u00f3sseas, encontraram um novo papel como ferramenta diagn\u00f3stica n\u00e3o invasiva da AC por ATTR.99mTc utilizados para o diagn\u00f3stico da AC por ATTR s\u00e3o o 99mTc-pirofosfato, o 99mTc-DPD e o 99mTc-HMDP (hidroximetilenodifosfonato marcado com 99mTc).\u201399mTc-pirofosfato \u00e9 o \u00fanico dispon\u00edvel no Brasil. \u00c9 importante ressaltar que o 99mTc-MDP (metilenodifosfonato marcado com 99mTc), apesar de comprovadamente eficiente para a realiza\u00e7\u00e3o da cintilografia \u00f3ssea, apresenta baixa sensibilidade para o diagn\u00f3stico da AC por ATTR e n\u00e3o deve ser usado para esta finalidade.Os principais radiotra\u00e7adores \u00f3sseos marcados com 99mTc-pirofosfato se liga no cora\u00e7\u00e3o n\u00e3o seja conhecido, um mecanismo de capta\u00e7\u00e3o c\u00e1lcio-dependente \u00e9 amplamente aceito.99mTc-pirofosfato no mioc\u00e1rdio esteja relacionado \u00e0 presen\u00e7a de microcalcifica\u00e7\u00f5es.99mTc-pirofosfato, enquanto a amiloidose card\u00edaca AL tem m\u00ednima ou nenhuma avidez pelos radiotra\u00e7adores \u00f3sseos; al\u00e9m disso, a AC por ATTR apresenta evolu\u00e7\u00e3o mais indolente, o que propicia mais microcalcifica\u00e7\u00f5es e, consequentemente, maior ac\u00famulo do radiotra\u00e7ador.Embora o componente estrutural do dep\u00f3sito amiloide ao qual o 99mTc-pirofosfato: um m\u00e9todo simples, de f\u00e1cil execu\u00e7\u00e3o, amplamente dispon\u00edvel, com baixa dosimetria, capaz de diferenciar de forma n\u00e3o invasiva e com alta especificidade a AC por ATTR da AL, orientando assim a conduta. Esta diferencia\u00e7\u00e3o \u00e9 \u00fatil, pois as formas AL e ATTR t\u00eam implica\u00e7\u00f5es progn\u00f3sticas e terap\u00eauticas completamente diversas.Embora os achados do ecocardiograma e da RMC possam ser indicativos de AC, eles n\u00e3o s\u00e3o capazes de diferenciar a forma ATTR da forma AL. Esta \u00e9 a principal vantagem da cintilografia card\u00edaca com expertise em AC: em an\u00e1lise cumulativa de 1.217 pacientes, sendo 867 com amiloidose confirmada por bi\u00f3psia e 360 portadores de cardiomiopatia n\u00e3o amiloide, a cintilografia foi altamente sens\u00edvel (99%) e espec\u00edfica (86%) para ATTR.99mTc-pirofosfato apresentou sensibilidade de 88% e especificidade de 88% para ATTR quando se utilizou apenas a avalia\u00e7\u00e3o visual (escore \u2265 2) 99mTc-pirofosfato foi preditiva de mortalidade geral e hospitaliza\u00e7\u00e3o por IC. Nesses estudos, a combina\u00e7\u00e3o da intensidade de capta\u00e7\u00e3o do radiotra\u00e7ador no mioc\u00e1rdio com vari\u00e1veis anat\u00f4micas, funcionais e biomarcadores melhorou a estratifica\u00e7\u00e3o de risco.O papel dos radiotra\u00e7adores \u00f3sseos na ATTR foi recentemente reavaliado por um grupo internacional de v\u00e1rios centros com 99mTc-pirofosfato . Imagens planas e tomogr\u00e1ficas do tipo SPECT (single photon emission computed tomography) do t\u00f3rax s\u00e3o realizadas 1 e 3h ap\u00f3s a administra\u00e7\u00e3o do radiof\u00e1rmaco.N\u00e3o \u00e9 necess\u00e1rio preparo. As imagens s\u00e3o obtidas ap\u00f3s a administra\u00e7\u00e3o venosa de 10 a 25 mCi (370 a 925 Mbq) de Imagens planas do t\u00f3rax: podem ser obtidas nas proje\u00e7\u00f5es anterior, obl\u00edqua anterior esquerda e lateral esquerda, utilizando o fotopico do 99mTc , colimador de baixa energia/alta resolu\u00e7\u00e3o e matriz de 256\u00d7256, 500\u2013750 mil contagens.Imagens SPECT do t\u00f3rax: obtidas com matriz de 128\u00d7128 (64\u00d764 \u00e9 aceit\u00e1vel), rota\u00e7\u00e3o de 180 graus de obl\u00edqua anterior direita a obl\u00edqua posterior esquerda (360 graus \u00e9 aceit\u00e1vel), 1 imagem a cada 3 a 6 graus. Se dispon\u00edveis, imagens SPECT/CT (SPECT associado \u00e0 tomografia computadorizada) proporcionam maior seguran\u00e7a na interpreta\u00e7\u00e3o das imagens.Imagens m\u00ednimas recomendadas: SPECT de 1h e imagens planas de 1 e 3h na proje\u00e7\u00e3o anterior. Recomenda-se a utiliza\u00e7\u00e3o de imagens SPECT, para diferenciar capta\u00e7\u00e3o difusa no mioc\u00e1rdio de persist\u00eancia do radiof\u00e1rmaco no pool sangu\u00edneo, capta\u00e7\u00e3o focal por infarto do mioc\u00e1rdio e sobreposi\u00e7\u00e3o \u00f3ssea. Al\u00e9m disso, imagens planas de 1 e 3h s\u00e3o \u00fateis para a quantifica\u00e7\u00e3o e para acompanhar o whashout do pool sangu\u00edneo, que \u00e9 vari\u00e1vel, sendo, por exemplo, bem mais lento em pacientes com insufici\u00eancia renal.An\u00e1lise semiquantitativa (imagem plana de 1h): a an\u00e1lise semiquantitativa foi definida para o radiof\u00e1rmaco 99mTc-pirofosfato como sendo a raz\u00e3o entre a capta\u00e7\u00e3o na proje\u00e7\u00e3o card\u00edaca e a capta\u00e7\u00e3o no hemit\u00f3rax contralateral, medidas na imagem plana de 1h na incid\u00eancia anterior. Para isso, \u00e9 desenhada uma \u00e1rea de interesse (AI) circular (ou el\u00edptica) sobre a proje\u00e7\u00e3o card\u00edaca, sem incluir o esterno, evitando a inclus\u00e3o tanto do pulm\u00e3o adjacente como de \u00e1reas de hipercapta\u00e7\u00e3o focal em arcos costais. Uma AI id\u00eantica (\u201cem espelho\u201d) \u00e9 colocada no hemit\u00f3rax contralateral, com os mesmos cuidados. As contagens no interior da AI card\u00edaca dividem-se pelas contagens da AI contralateral, obtendo-se, assim, a rela\u00e7\u00e3o C/CL . A C/CL \u2265 1,5 \u00e0 1h identifica ATTR com elevada acur\u00e1cia, caso a amiloidose AL sist\u00eamica tenha sido exclu\u00eddaGradua\u00e7\u00e3o visual (imagem de 3h): a gradua\u00e7\u00e3o visual \u00e9 feita comparando-se a capta\u00e7\u00e3o card\u00edaca com a capta\u00e7\u00e3o fisiol\u00f3gica nos arcos costais adjacentes, e pode ser realizada tanto nas imagens planas na proje\u00e7\u00e3o anterior como nas imagens SPECT ou mesmo nas imagens de corpo inteiro, obtidas 3h ap\u00f3s a inje\u00e7\u00e3o do radiof\u00e1rmaco. A intensidade da capta\u00e7\u00e3o \u00e9 numericamente definida como grau 0 (sem capta\u00e7\u00e3o mioc\u00e1rdica), 1 (capta\u00e7\u00e3o mioc\u00e1rdica inferior \u00e0 dos arcos costais adjacentes), 2 (capta\u00e7\u00e3o semelhante \u00e0 dos arcos costais) e 3 (superior \u00e0 dos arcos costais). Intensidades de capta\u00e7\u00e3o graus 2 ou 3 s\u00e3o fortemente sugestivas de ATTR, se gamopatia monoclonal tiver sido exclu\u00edda implicam risco de diagn\u00f3sticos imprecisos. A causa mais comum de diagn\u00f3stico err\u00f4neo de ATTR \u00e9 a amiloidose AL. Estudos recentes apontam que at\u00e9 22% dos pacientes com AC da forma AL podem apresentar capta\u00e7\u00e3o grau 2 ou 3 na cintilografia com angu\u00edneo . A TabelFinalmente, \u00e9 importante considerar que o estudo cintilogr\u00e1fico n\u00e3o \u00e9 recomendado para o seguimento dos pacientes, visto que, at\u00e9 o momento, n\u00e3o se evidenciou correla\u00e7\u00e3o entre a altera\u00e7\u00e3o no padr\u00e3o evolutivo das imagens e progress\u00e3o da doen\u00e7a ou a resposta ao tratamento.A N\u00e3o existe um marcador laboratorial espec\u00edfico para o diagn\u00f3stico de AC.A troponina e os pept\u00eddios natriur\u00e9ticos t\u00eam se mostrado \u00fateis para avaliar acometimento card\u00edaco pela amiloidose, permitindo aux\u00edlio diagn\u00f3stico n\u00e3o invasivo, acess\u00edvel e relativamente de baixo custo.An\u00e1lise de dados do registro THAOS (Transthyretin Amyloidosis Outcomes Survey), com pacientes acometidos por ATTR, demonstrou que a dosagem dos pept\u00eddios natriur\u00e9ticos pode ser utilizada para aux\u00edlio diagn\u00f3stico, e foram observados valores mais elevados nos pacientes com muta\u00e7\u00f5es de acometimento preferencialmente card\u00edaco, como a V122I e V30M-tardia, do que nas formas de predom\u00ednio neurol\u00f3gico, como a V30M-precoce. Tamb\u00e9m foram observados maiores n\u00edveis dos biomarcadores na ATTRwt quando comparada com as formas heredit\u00e1rias de predom\u00ednio neurol\u00f3gico. Por\u00e9m, observou-se que, mesmo nos pacientes com fen\u00f3tipo predominantemente neurol\u00f3gico, 45% a 90% apresentavam altera\u00e7\u00f5es desses biomarcadores, indicando algum grau de acometimento mioc\u00e1rdico subcl\u00ednico.Observam-se maiores valores de NT-proBNP na forma AL, em compara\u00e7\u00e3o \u00e0 ATTR. Isso se deve ao fato de as cadeias leves amiloidog\u00eanicas modularem a prote\u00edna cinase ativada por mit\u00f3geno p38 (MAPK), que diretamente promove a express\u00e3o do NT-proBNP, de forma que, para um mesmo grau de altera\u00e7\u00f5es hemodin\u00e2micas nas duas formas de amiloidose, os n\u00edveis s\u00e9ricos da NT-proBNP podem ser maiores na forma AL.Vale ressaltar que os pacientes com ICFEP causada por ATTR apresentam valores de NT-ProBNP desproporcionalmente elevados em rela\u00e7\u00e3o \u00e0 gravidade da s\u00edndrome de IC, quando comparados aos pacientes com ICFEP n\u00e3o amiloide.Eleva\u00e7\u00e3o discreta e persistente dos n\u00edveis de troponina \u00e9 frequentemente observada e sugere dano mioc\u00e1rdico subcl\u00ednico em v\u00e1rias cardiomiopatias n\u00e3o isqu\u00eamicas.V\u00e1rios mecanismos para justificar a eleva\u00e7\u00e3o das troponinas nesses pacientes t\u00eam sido postulados: isquemia mioc\u00e1rdica, estresse de parede aumentado, les\u00e3o direta do mi\u00f3cito por citocinas inflamat\u00f3rias e/ou estresse oxidativo, ativa\u00e7\u00e3o neuro-hormonal e disfun\u00e7\u00e3o coronariana microvascular na insufici\u00eancia card\u00edaca. A disfun\u00e7\u00e3o microvascular na amiloidose \u00e9 causada, presumidamente, por dep\u00f3sito intersticial e na regi\u00e3o perivascular, press\u00e3o de enchimento ventricular aumentada e disfun\u00e7\u00e3o endotelial por toxicidade induzida pelas imunoglobulinas, na forma AL. Al\u00e9m disso, tamb\u00e9m \u00e9 descrito efeito cardiot\u00f3xico direto das cadeias leves, independentemente do dep\u00f3sito extracelular de fibrilas, podendo justificar os n\u00edveis mais elevados de troponina em pacientes com forma AL do que em pacientes com ATTR, nos quais geralmente se observa eleva\u00e7\u00e3o discreta e persistente.Vale ressaltar que, entre os pacientes com ATTR, muitos apresentam comorbidades, como miocardiopatia isqu\u00eamica, o que pode causar valores alterados de troponina. Sendo assim, a dosagem de troponina n\u00e3o deve ser utilizada para afastar ou confirmar acometimento card\u00edaco, mas sim como um potencial sinal de alerta para a doen\u00e7a, que ser\u00e1 mais bem avaliada com exames mais espec\u00edficos.Uma s\u00e9rie de outros biomarcadores tem sido estudada, alguns com alta especificidade para subtipos de amiloidose, como a dosagem de prote\u00edna carreadora de retinol 4 para a amiloidose heredit\u00e1ria pela muta\u00e7\u00e3o Val142Ile. No entanto, ainda mais estudos s\u00e3o necess\u00e1rios, assim como a disponibiliza\u00e7\u00e3o comercial dos testes para an\u00e1lises de rotina.Evid\u00eancias recentes indicam que a AC, particularmente na forma ATTRwt, \u00e9 uma condi\u00e7\u00e3o mais prevalente do que anteriormente se estimava, e que, em parte, isso decorre de um amplo subdiagn\u00f3stico e do fato de esta doen\u00e7a mimetizar outras cardiopatias, tais como a miocardiopatia hipertr\u00f3fica, a ICFEP n\u00e3o amiloide e a estenose a\u00f3rtica de baixo fluxo e baixo gradiente.etapa preliminar e fundamental \u00e9 o estabelecimento da suspeita cl\u00ednica, baseada na hist\u00f3ria cl\u00ednica, exame f\u00edsico , com detec\u00e7\u00e3o de uma rela\u00e7\u00e3o anormal entre as cadeias kappa/lambda , aumenta a sensibilidade de detec\u00e7\u00e3o para acima de 99%.A eletroforese de prote\u00edna n\u00e3o \u00e9 um teste de rota hematol\u00f3gica no fluxograma, e a realiza\u00e7\u00e3o de bi\u00f3psia tecidual, procedimento fundamental para confirmar o dep\u00f3sito de prote\u00edna amiloide e a elabora\u00e7\u00e3o da estrat\u00e9gia terap\u00eautica.A detec\u00e7\u00e3o de cadeias leves monoclonais torna necess\u00e1rio o encaminhamento do paciente para avalia\u00e7\u00e3o com hematologista, seguindo-se a O diagn\u00f3stico de amiloidose AL dever ser confirmado atrav\u00e9s de bi\u00f3psia.Deve-se dar prefer\u00eancia, inicialmente, \u00e0 bi\u00f3psia de gordura abdominal, um m\u00e9todo simples e seguro.Em casos de amiloidose sist\u00eamica, em que h\u00e1 acometimento de v\u00e1rios \u00f3rg\u00e3os ou tecidos, a bi\u00f3psia de gordura abdominal com essas colora\u00e7\u00f5es espec\u00edficas apresenta sensibilidade de 60% a 80% e especificidade de 90% a 100% no diagn\u00f3stico de amiloidose,Vale ressaltar que a colora\u00e7\u00e3o vermelho-congo permite a confirma\u00e7\u00e3o da infiltra\u00e7\u00e3o amiloide nos tecidos, por\u00e9m n\u00e3o identifica o tipo da prote\u00edna precursora. Al\u00e9m disso, 40% dos pacientes com ATTR podem ter uma gamopatia monoclonal de significado indeterminado (GMSI), apresentando investiga\u00e7\u00e3o positiva para cadeias leves monoclonais.laser do material amiloide da bi\u00f3psia com realiza\u00e7\u00e3o de espectrometria de massa. A imuno-histoqu\u00edmica permanece o m\u00e9todo mais amplamente dispon\u00edvel para identifica\u00e7\u00e3o do tipo de dep\u00f3sito; contudo, quando o tipo de amiloidose \u00e9 o de cadeias leves, os resultados n\u00e3o s\u00e3o sempre conclusivos, frequentemente com coexist\u00eancia de positividade para mais de um tipo de antissoro, geralmente TTR e cadeia kappa ou lambda. A espectrometria de massa tornou-se, portanto, o novo \u201cpadr\u00e3o-ouro\u201d para a identifica\u00e7\u00e3o do tipo do dep\u00f3sito amiloide.,,Considerando esses aspectos, para identifica\u00e7\u00e3o do tipo do dep\u00f3sito amiloide, aspecto fundamental para o tratamento apropriado, torna-se necess\u00e1ria a realiza\u00e7\u00e3o de imuno-histoqu\u00edmica ou, preferencialmente, de microdissec\u00e7\u00e3o a resultados negativos para cadeias leves monoclonais, situa\u00e7\u00e3o em que o diagn\u00f3stico de ATTR \u00e9 mais prov\u00e1vel, ainda que outras formas mais raras de AC tamb\u00e9m possam ser diagnosticadas, a investiga\u00e7\u00e3o deve seguir pela rota cardiol\u00f3gica, tomando-se duas sub-rotas de acordo com a disponibilidade da cintilografia card\u00edaca com marcadores \u00f3sseos.Perante cen\u00e1rio de disponibilidade da cintilografia com marcadores \u00f3sseos (sub-rota 2A), desde que as cadeias leves monoclonais estejam ausentes, o achado de cintilografia card\u00edaca mostrando capta\u00e7\u00e3o acentuada, graus 2 ou 3 da classifica\u00e7\u00e3o de Perugini e rela\u00e7\u00e3o de capta\u00e7\u00e3o na \u00e1rea card\u00edaca em rela\u00e7\u00e3o \u00e0 \u00e1rea contralateral do t\u00f3rax \u2265 1,5, confirmando-se que a capta\u00e7\u00e3o aumentada encontra-se nas paredes ventriculares por imagens tomogr\u00e1ficas de emiss\u00e3o (SPECT), a AC por ATTR est\u00e1 confirmada sem a necessidade de realiza\u00e7\u00e3o de bi\u00f3psia endomioc\u00e1rdica. A seguir, o sequenciamento gen\u00e9tico da TTR deve ser realizado para defini\u00e7\u00e3o do caso se ATTR heredit\u00e1ria ou selvagem.No wild-type tem implica\u00e7\u00f5es progn\u00f3sticas e terap\u00eauticas, sendo tamb\u00e9m importante para potencial screening familiar e aconselhamento gen\u00e9tico.A diferencia\u00e7\u00e3o entre ATTR heredit\u00e1ria ou Na vig\u00eancia de cintilografia card\u00edaca com marcadores \u00f3sseos negativa e associada \u00e0 aus\u00eancia de cadeias leves monoclonais, o diagn\u00f3stico de AC \u00e9 improv\u00e1vel. Contudo, quando a suspeita cl\u00ednica persiste, baseada principalmente nos resultados de outros m\u00e9todos de imagem muito sugestivos de amiloidose, a bi\u00f3psia endomioc\u00e1rdica tem um papel diagn\u00f3stico relevante e deve ser solicitada. Nesse caso, podemos estar diante de ATTRv com algumas muta\u00e7\u00f5es com dep\u00f3sitos amiloides que n\u00e3o captam marcador \u00f3sseo, como a V30M de in\u00edcio precoce e a P64I, al\u00e9m de outros tipos n\u00e3o usuais de amiloidose.indisponibilidade da cintilografia com marcadores \u00f3sseos (sub-rota 2B), a bi\u00f3psia endomioc\u00e1rdica tamb\u00e9m est\u00e1 indicada para elucidar o diagn\u00f3stico.No cen\u00e1rio de Portadores de AC forma AL exibem dep\u00f3sitos amiloides mais t\u00f3xicos e de forma\u00e7\u00e3o mais acelerada se comparados aos casos pela forma ATTR. Como consequ\u00eancia, aumentos mensais da espessura mioc\u00e1rdica da monta de 1,45 a 2,16 mm podem ser observados associados a eleva\u00e7\u00f5es mais acentuadas de biomarcadores como troponina e BNP,A somat\u00f3ria desses achados \u00e0s informa\u00e7\u00f5es obtidas gra\u00e7as aos recentes avan\u00e7os tecnol\u00f3gicos e laboratoriais permitiu estabelecer como principal determinante do progn\u00f3stico da amiloidose AL a extens\u00e3o do envolvimento card\u00edaco pelo material amiloide,strain global longitudinal do VE combinados aos biomarcadores BNP e troponina e status hematol\u00f3gico s\u00e3o preditores de maior mortalidade.Adicionalmente, achados ecocardiogr\u00e1ficos, tais como aumento acentuado da espessura das paredes, disfun\u00e7\u00e3o diast\u00f3lica, disfun\u00e7\u00e3o do VE, espessamento das valvas e redu\u00e7\u00e3o do ,\u2013\u2013extracellular volume) calculado na fase de equil\u00edbrio do contraste superiores a 0,45 associam-se a aumento da mortalidade cardiovascular de 5,84 e 3,48 vezes, respectivamente.A RMC tamb\u00e9m fornece elementos que se correlacionam com a sobrevida em pacientes com AC.Os biomarcadores apresentam n\u00e3o apenas um papel no estadiamento desses doentes, mas tamb\u00e9m possibilitam avalia\u00e7\u00e3o da resposta ao tratamento, que na pr\u00e1tica cl\u00ednica \u00e9 sempre um desafio, principalmente para a forma AL, uma vez que a quimioterapia pode ser um fator adicional para les\u00e3o mioc\u00e1rdica. O que se tem bem estabelecido na literatura \u00e9 a concord\u00e2ncia entre redu\u00e7\u00e3o dos n\u00edveis de NT-proBNP e resposta hematol\u00f3gica ao tratamento, assim como melhora de classe funcional NYHA. O inverso tamb\u00e9m \u00e9 verdadeiro, como o aumento dos n\u00edveis de NT-proBNP, troponina e redu\u00e7\u00e3o da fra\u00e7\u00e3o de eje\u00e7\u00e3o. e a RMC, fornecem dados complementares que auxiliam na estratifica\u00e7\u00e3o progn\u00f3stica.,,,\u2013Achados em exames de imagem, como o ecocardiograma, a cintilografia card\u00edaca com Tecn\u00e9cio-pirofosfato . Com isso, a base do tratamento espec\u00edfico consiste em eliminar a sua produ\u00e7\u00e3o por meio da erradica\u00e7\u00e3o do clone de plasm\u00f3citos na medula \u00f3ssea. A redu\u00e7\u00e3o r\u00e1pida e, idealmente, normaliza\u00e7\u00e3o dos n\u00edveis de cadeias leves s\u00e3o alguns dos principais objetivos no tratamento da amiloidose AL, sendo denominada resposta hematol\u00f3gica. J\u00e1 a revers\u00e3o da les\u00e3o nos tecidos e \u00f3rg\u00e3os acometidos pelo dep\u00f3sito amiloide \u00e9 chamada de resposta org\u00e2nica, e compreende o segundo maior objetivo do tratamento. Outros objetivos do tratamento s\u00e3o melhorar a qualidade de vida e a sobrevida geral.A amiloidose AL \u00e9 uma doen\u00e7a causada pela produ\u00e7\u00e3o aberrante da cadeia leve da imunoglobulina aut\u00f3logo quanto naqueles tratados sem esse procedimento. A sobrevida mediana segundo essa estratifica\u00e7\u00e3o progn\u00f3stica foi de 55, 19, 12 e 5 meses em pacientes que tiveram ao diagn\u00f3stico est\u00e1gios I, II, III e IV, respectivamente, e que n\u00e3o receberam ou tiveram falha no TCTH e 97, 58 e 22 meses, para os est\u00e1gios II, III e IV, respectivamente, naqueles submetidos ao TCTH.Pacientes avaliados como baixo risco s\u00e3o candidatos \u00e0 terapia com quimioterapia (QT) em altas doses seguidas de TCTH aut\u00f3logo . AtualmeAp\u00f3s 3 meses do t\u00e9rmino do tratamento, as respostas hematol\u00f3gica e org\u00e2nica devem ser avaliadas com exames espec\u00edficos relacionados \u00e0 detec\u00e7\u00e3o de gamopatia monoclonal e comprometimento de cada \u00f3rg\u00e3o acometido no diagn\u00f3stico. Os crit\u00e9rios de resposta hematol\u00f3gica e org\u00e2nica est\u00e3o resumidos nas Tabelas Mesmo alcan\u00e7ando resposta hematol\u00f3gica completa, a melhora das disfun\u00e7\u00f5es se d\u00e1 mais lentamente, podendo ocorrer at\u00e9 meses a anos ap\u00f3s a normaliza\u00e7\u00e3o das cadeias leves.,,Apenas 20% dos pacientes diagnosticados com amiloidose AL apresentam condi\u00e7\u00f5es cl\u00ednicas para o TCTH, e o n\u00famero de pacientes eleg\u00edveis pode aumentar se houver resposta org\u00e2nica ap\u00f3s esquemas de indu\u00e7\u00e3o contendo terapias antiplasmocit\u00e1rias eficazes .A avalia\u00e7\u00e3o de elegibilidade ao TCTH \u00e9 ponto-chave para se obter sucesso terap\u00eautico com essa estrat\u00e9gia. Um \u00fanico estudo prospectivo randomizado comparou o TCTH com QT convencional, n\u00e3o havendo benef\u00edcio de sobrevida global deste em rela\u00e7\u00e3o \u00e0 estrat\u00e9gia menos intensiva. Neste estudo, por\u00e9m, a alta taxa de mortalidade associada ao TCTH (24%) foi relacionada aos crit\u00e9rios de inclus\u00e3o, inexperi\u00eancia do centro transplantador e uso de dose subterap\u00eautica de melfalano.New York Heart Association I ou II e press\u00e3o arterial sist\u00f3lica > 90 mmHg. A adequada estratifica\u00e7\u00e3o de risco pr\u00e9-tratamento concomitante ao desenvolvimento de melhores condi\u00e7\u00f5es de suporte tais como antibioticoterapia e terapia intensiva tem levado \u00e0 redu\u00e7\u00e3o da mortalidade associada ao TCTH nas \u00faltimas d\u00e9cadas, com taxas de 2,4% a 3,4% em an\u00e1lises retrospectivas.\u2013Uma vez que n\u00e3o h\u00e1 crit\u00e9rios precisos bem estabelecidos e validados prospectivamente na literatura, cada centro estabelece seus pr\u00f3prios crit\u00e9rios de elegibilidade ao TCTH. Embora a avalia\u00e7\u00e3o subjetiva seja relevante, alguns fatores podem guiar a indica\u00e7\u00e3o do TCTH: \u201cidade fisiol\u00f3gica\u201d \u2264 70 anos, ClCr > 30 mL/min (exceto aqueles em di\u00e1lise cr\u00f4nica), troponina T < 0,06 ng/mL, escala de performance ECOG (Eastern Cooperative Oncology Group) \u2264 2, classe funcional da performance status, aumento da sobrevida e qualidade de vida. Como exemplo, em uma s\u00e9rie de 672 pacientes com amiloidose AL submetidos a TCTH, 84% apresentaram resposta hematol\u00f3gica, sendo 39% de RC, e a sobrevida alcan\u00e7ada foi > 50% em 15 anos.,,Dessa forma, o TCTH deve ser recomendado como terapia de primeira linha nos indiv\u00edduos classificados como eleg\u00edveis . Essa reupfront) em pacientes com < 10% de plasm\u00f3citos clonais na medula \u00f3ssea ou ser precedido de terapia de indu\u00e7\u00e3o com esquemas contendo bortezomibe e/ou anticorpo monoclonal anti-CD38. Esse \u00faltimo cen\u00e1rio deve ser considerado quando h\u00e1 associa\u00e7\u00e3o com mieloma m\u00faltiplo, atraso previs\u00edvel para realiza\u00e7\u00e3o do TCTH, baixa performance que pode melhorar com a terapia de indu\u00e7\u00e3o e presen\u00e7a de > 10% de infiltra\u00e7\u00e3o por plasm\u00f3citos clonais na medula \u00f3ssea, a qual est\u00e1 associada a pior progn\u00f3stico.,O TCTH pode ser realizado logo ap\u00f3s diagn\u00f3stico e daratumumabe, bortezomibe, ciclofosfamida e dexametasona (Dara-CyBorD).Dado o novo cen\u00e1rio de surgimento de novas terapias no tratamento da amiloidose AL e a escassez de estudos randomizados consolidando o papel do transplante, ser\u00e3o necess\u00e1rios novos estudos para entender se o TCTH permanecer\u00e1 sendo a terapia mais eficaz nas pr\u00f3ximas d\u00e9cadas.A maior parte dos pacientes diagnosticados com amiloidose AL n\u00e3o \u00e9 eleg\u00edvel \u00e0 terapia de maior intensidade com TCTH aut\u00f3logo devido a comorbidades, como doen\u00e7a card\u00edaca avan\u00e7ada, insufici\u00eancia renal, envolvimento de mais de dois \u00f3rg\u00e3os ou idade avan\u00e7ada. Dessa forma, nesse grupo de pacientes, terapias quimioter\u00e1picas com medica\u00e7\u00f5es antiplasmocit\u00e1rias s\u00e3o a base do tratamento . Os esqu,\u2013O esquema combinando ciclofosfamida, bortezomibe e dexametasona (CyBorD) demonstrou-se altamente eficaz (resposta hematol\u00f3gica 81% a 94%), bem tolerado e capaz de gerar resposta hematol\u00f3gica r\u00e1pida (em at\u00e9 3 meses) em dois estudos retrospectivos com pequeno n\u00famero de pacientes (n = 17 e 43). A sobrevida global em 2 anos chegou a 92% nesses estudos.vs. 52%), resposta org\u00e2nica card\u00edaca (38% vs. 28%) e melhora de sobrevida global, com redu\u00e7\u00e3o de 2 vezes na taxa de mortalidade.versus 77% no grupo-controle que recebeu CyBorD sem daratumumabe, com RC de 53% versus 18%, e mediana de 2 meses para atingir esta resposta. Melhora de fun\u00e7\u00e3o org\u00e2nica foi tamb\u00e9m observada no grupo que recebeu o anticorpo monoclonal, com melhora da sobrevida livre de progress\u00e3o. \u00c9 importante ressaltar que pacientes com cardiopatia avan\u00e7ada (classificados como estadiamento IIIb) foram exclu\u00eddos de ambos os estudos randomizados citados, sendo que o tratamento para este grupo permanece um desafio na pr\u00e1tica cl\u00ednica.Recentemente, dois estudos randomizados institu\u00edram combina\u00e7\u00f5es de QT como poss\u00edveis novos tratamentos de escolha em pacientes com amiloidose AL n\u00e3o eleg\u00edveis a TCTH. O primeiro comparou o esquema bortezomibe, melfalano e dexametasona (BMD) a MD, demonstrando benef\u00edcio da adi\u00e7\u00e3o do bortezomibe a MD com maior taxa de resposta hematol\u00f3gica global ap\u00f3s 3 meses e, assim, inibindo efetivamente a cascata que resulta na forma\u00e7\u00e3o das fibrilas amiloides.2. Os pacientes foram randomizados para receber tafamidis na dose de 80 mg/dia, tafamidis 20 mg/dia ou placebo nas propor\u00e7\u00f5es de 2:1:2. O desfecho prim\u00e1rio do estudo avaliou de forma hier\u00e1rquica mortalidade por qualquer causa, seguida pela frequ\u00eancia de hospitaliza\u00e7\u00e3o cardiovascular ao longo de 30 meses de seguimento. Os desfechos secund\u00e1rios principais foram a mudan\u00e7a da dist\u00e2ncia percorrida no TC6m e do escore de qualidade de vida obtido no Kansas City Cardiomyopathy Questionnaire (KCCQ). Para an\u00e1lise estat\u00edstica, agruparam-se os resultados dos grupos de pacientes recebendo as doses de tafamidis de 80mg e 20mg. Os resultados reportados mostraram que os pacientes recebendo tafamidis (n=264) exibiram redu\u00e7\u00e3o de 30% no risco relativo de mortalidade por qualquer causa quando comparados aos pacientes recebendo placebo (n = 177) , p = 0,0374.Com base nessas evid\u00eancias, recomenda-se o uso de tafamidis na dose de 80mg/dia para tratamento de pacientes com AC por ATTRv ou ATTRwt, para pacientes com insufici\u00eancia card\u00edaca, em CF I a III da NYHA, sem disfun\u00e7\u00e3o renal grave, devendo-se iniciar a terapia nas fases mais precoces da doen\u00e7a . No BrasAG10 \u00e9 um estabilizador seletivo dos tetr\u00e2meros da TTR, desenhado para mimetizar a influ\u00eancia estrutural da muta\u00e7\u00e3o protetora superestabilizadora T119M, que reduz significativamente a taxa de dissocia\u00e7\u00e3o dos tetr\u00e2meros.anti-sense (Inotersen). Ambos os f\u00e1rmacos foram testados em estudos multic\u00eantricos fase 3 em pacientes com polineuropatia por ATTRv e se mostraram efetivos em reduzir a progress\u00e3o das manifesta\u00e7\u00f5es neurol\u00f3gicas.,post hoc de subgrupos de pacientes com AC inclu\u00eddos nesses estudos sugerem efeitos positivos sobre a progress\u00e3o da cardiomiopatia. Ambas as classes de silenciadores de express\u00e3o g\u00eanica est\u00e3o atualmente sendo testadas em estudos multic\u00eantricos fase 3 em pacientes com diagn\u00f3stico de AC-ATTR .Terapias baseadas no silenciamento da express\u00e3o dos genes que codificam a produ\u00e7\u00e3o hep\u00e1tica de TTR s\u00e3o muito promissoras, incluindo estrat\u00e9gias com RNA de interfer\u00eancia (Patisiran) e oligonucleot\u00eddeos in vitro e experimentais para promover a desagrega\u00e7\u00e3o das fibras amiloides depositadas nos tecidos, permitindo a reabsor\u00e7\u00e3o dos seus dep\u00f3sitos pelo sistema macrof\u00e1gico.Alguns compostos com base em mol\u00e9culas hidrof\u00f3bicas mostraram-se efetivos em estudos ,Al\u00e9m da terapia espec\u00edfica para amiloidose, pode ser necess\u00e1rio tratamento de suporte para IC. A AC apresenta-se inicialmente como IC de fra\u00e7\u00e3o de eje\u00e7\u00e3o preservada (ICFEp) e padr\u00e3o restritivo do enchimento ventricular esquerdo, podendo ocorrer, tardiamente, a progress\u00e3o da doen\u00e7a e redu\u00e7\u00e3o da fra\u00e7\u00e3o de eje\u00e7\u00e3o. Esse mecanismo fisiopatol\u00f3gico pode explicar as dificuldades de manejo cl\u00ednico do paciente com amiloidose card\u00edaca ao se usar medica\u00e7\u00f5es consagradas para IC de fra\u00e7\u00e3o de eje\u00e7\u00e3o reduzida (ICFEr).172,173 Al\u00e9m disso, em pacientes com polineuropatia auton\u00f4mica, a presen\u00e7a de hipotens\u00e3o pode dificultar o uso de diur\u00e9ticos devido \u00e0 pr\u00e9-carga l\u00e1bil.A manuten\u00e7\u00e3o da euvolemia, com restri\u00e7\u00e3o h\u00eddrica e medicamentos, \u00e9 o foco. A medica\u00e7\u00e3o mais utilizada \u00e9 o diur\u00e9tico de al\u00e7a, indicado para reduzir a congest\u00e3o pulmonar e sist\u00eamica, podendo associar-se os antagonistas da aldosterona . O uso d,Em rela\u00e7\u00e3o aos f\u00e1rmacos modificadores de progn\u00f3stico usados para o tratamento da ICFER, os antagonistas neuro-hormonais como inibidores da enzima de convers\u00e3o da angiotensina II (iECA), bloqueadores dos receptores de angiotensina II (BRA) e betabloqueadores (BB), ou at\u00e9 mesmo subst\u00e2ncias recentemente descritas como os inibidores da neprilisina e antagonistas do receptor de angiotensina II (INRA) e inibidores de SGLT2, n\u00e3o h\u00e1 evid\u00eancias cient\u00edficas de benef\u00edcio na amiloidose card\u00edaca, al\u00e9m de haver risco de hipotens\u00e3o associada \u00e0 disfun\u00e7\u00e3o auton\u00f4mica. Aimo et al.As arritmias s\u00e3o muito comuns em pacientes com amiloidose card\u00edaca, e geralmente s\u00e3o sintom\u00e1ticas e pouco toleradas. A avalia\u00e7\u00e3o de arritmias nesta popula\u00e7\u00e3o deve envolver tr\u00eas diferentes situa\u00e7\u00f5es: arritmias atriais; arritmias ventriculares e doen\u00e7a do sistema de condu\u00e7\u00e3o.,O dep\u00f3sito amiloide leva a espessamento atrial com altera\u00e7\u00e3o de relaxamento atrial, aumento das press\u00f5es intracavit\u00e1rias, gerando dilata\u00e7\u00e3o atrial que, em associa\u00e7\u00e3o \u00e0 fibrose atrial, predisp\u00f5e \u00e0 fibrila\u00e7\u00e3o atrial (FA) ou a outras arritmias atriais. A preval\u00eancia de FA pode variar de 11% a 71% em pacientes com AC, sendo ainda mais elevada em pacientes com AC por ATTR, possivelmente por acometer pacientes mais idosos do sexo masculino.O manejo da FA nesses pacientes \u00e9, em geral, dif\u00edcil, uma vez que costumam n\u00e3o tolerar os principais f\u00e1rmacos utilizados para o controle de frequ\u00eancia, tais como BB, BCC ou digit\u00e1licos, por precipitarem hipotens\u00e3o postural e descompensa\u00e7\u00e3o da IC. Diante da necessidade de uso dessas medica\u00e7\u00f5es, aconselha-se uso de baixas doses com monitoramento hemodin\u00e2mico cauteloso, al\u00e9m de controle de n\u00edvel s\u00e9rico, se digoxina for utilizada. Para o controle de ritmo, uma an\u00e1lise retrospectiva n\u00e3o mostrou diferen\u00e7a em rela\u00e7\u00e3o \u00e0 sobrevida quando comparados pacientes que receberam f\u00e1rmacos antiarr\u00edtmicos com pacientes que foram tratados apenas com controle de frequ\u00eancia.,,,Por fim, a redu\u00e7\u00e3o da contratilidade atrial provocada pela infiltra\u00e7\u00e3o amiloide no tecido atrial pode ainda contribuir para a forma\u00e7\u00e3o de trombos, dado este demonstrado em estudos de aut\u00f3psia que revelam a presen\u00e7a de at\u00e9 33% de trombos intracavit\u00e1rios em pacientes com AC,Arritmias ventriculares s\u00e3o frequentes em pacientes com AC, especialmente amiloidose AL. Estudos pr\u00e9vios t\u00eam detectado a presen\u00e7a de arritmias ventriculares complexas em pelo menos 50% dos pacientes com AC forma AL, sendo taquicardia ventricular n\u00e3o sustentada (TVNS) a arritmia mais frequente e associada a menor sobrevida.\u2013Neste contexto, discute-se o papel do uso de cardiodesfibrilador implant\u00e1vel (CDI) na preven\u00e7\u00e3o de morte s\u00fabita de pacientes com AC. Na presen\u00e7a de taquicardia ventricular inst\u00e1vel ou sobreviventes de parada card\u00edaca sem causa revers\u00edvel com expectativa de vida maior que 1 ano com qualidade significativa, sugere-se potencial benef\u00edcio do uso de CDI.No entanto, como preven\u00e7\u00e3o prim\u00e1ria, a indica\u00e7\u00e3o de CDI \u00e9 mais dif\u00edcil por diferentes motivos. O primeiro deles \u00e9 que a maioria das causas de morte s\u00fabita descritas relaciona-se \u00e0 dissocia\u00e7\u00e3o eletromec\u00e2nica e n\u00e3o arritmias ventriculares;wild type) e 36% dos pacientes com ATTRv.Doen\u00e7a do sistema de condu\u00e7\u00e3o \u00e9 altamente prevalente entre pacientes com AC, sendo a condu\u00e7\u00e3o atrioventricular mais comumente afetada do que o n\u00f3 sinusal. A fisiopatologia envolvida n\u00e3o est\u00e1 totalmente definida, embora estudos pequenos sugiram envolvimento do sistema de condu\u00e7\u00e3o por prote\u00edna amiloide.,Na IC avan\u00e7ada associada \u00e0 AC, o uso de estrat\u00e9gias avan\u00e7adas de suporte incluindo dispositivos de assist\u00eancia circulat\u00f3ria mec\u00e2nica e transplante apresenta desafios e particularidades, especialmente por se tratar de doen\u00e7a multissist\u00eamica. Al\u00e9m disso, pelo tamanho reduzido da cavidade do ventr\u00edculo esquerdo e o frequente acometimento do ventr\u00edculo direito, pode haver limita\u00e7\u00e3o ao uso de dispositivos de assist\u00eancia circulat\u00f3ria mec\u00e2nica de longa dura\u00e7\u00e3o.A O acesso ao cuidado comprometido com melhoria cont\u00ednua da qualidade assistencial \u00e9 pilar fundamental para alcan\u00e7ar resultados de excel\u00eancia, sendo item cr\u00edtico nos casos das doen\u00e7as complexas como a AC. Os gestores p\u00fablicos e privados, pacientes, profissionais de sa\u00fade e todos envolvidos no cuidado ao paciente com amiloidose est\u00e3o hoje comprometidos na constru\u00e7\u00e3o de uma jornada que possibilite o cuidado certo e no lugar certo.\u2013A AC deve ser abordada tendo como ponto fundamental o r\u00e1pido diagn\u00f3stico, particularmente nas formas AL, e a disponibilidade de equipes multidisciplinares com comprovada experi\u00eancia e utilizando protocolos cl\u00ednicos embasados nas melhores evid\u00eancias cient\u00edficas. Ao lado disso, \u00e9 necess\u00e1rio promover cuidado integral e monitorar e publicar os resultados assistenciais. Nesses centros de refer\u00eancia, deve-se observar um modelo de pr\u00e1tica integrada e colaborativa, com estrutura de telemedicina para apoiar o diagn\u00f3stico de centros mais distantes, monitorar o paciente e promover pesquisa cl\u00ednica, base para incorpora\u00e7\u00e3o futura de novas tecnologias (terapias emergentes). Essa iniciativa est\u00e1 dispon\u00edvel em v\u00e1rios lugares do mundo e est\u00e1 sendo implementada com sucesso em algumas regi\u00f5es do nosso pa\u00eds.Adicionalmente, \u00e9 relevante considerar que os centros de refer\u00eancia devem ser os pilares para a constru\u00e7\u00e3o de um Registro Nacional de Amiloidose Card\u00edaca, permitindo melhor conhecimento da epidemiologia no nosso meio e da qualidade do cuidado, al\u00e9m de mensurar os desfechos cl\u00ednicos centrados no paciente, de forma a contribuir para elabora\u00e7\u00e3o de pol\u00edticas p\u00fablicas.,A constru\u00e7\u00e3o de novos modelos de financiamento dos medicamentos de alto custo para doen\u00e7as raras vem sendo discutida, testada em estudos de farmacoeconomia e implementada ao redor do mundo.De acordo com a avalia\u00e7\u00e3o da incorpora\u00e7\u00e3o de novos tratamentos no Sistema \u00danico de Sa\u00fade (SUS) para as doen\u00e7as raras com o apoio da CONITEC , foram incorporados cerca de 52% dos medicamentos avaliados. Nossa vis\u00e3o \u00e9 que estamos construindo um novo cen\u00e1rio de tal forma, que o embasamento t\u00e9cnico, \u00e0 luz de dados robustos e com o apoio de protocolos cl\u00ednicos e centros de refer\u00eancia, possa substituir o modelo da busca de acesso via judicializa\u00e7\u00e3o e seus impactos sobre os gastos da Uni\u00e3o.sitehttps://www.clinicaltrials.gov.Apesar dos amplos avan\u00e7os recentemente alcan\u00e7ados no entendimento da AC, ainda h\u00e1 in\u00fameras lacunas do conhecimento em torno desta doen\u00e7a complexa e plurifacetada que necessitam ser elucidadas. O diagn\u00f3stico da amiloidose, assim como seu tratamento, \u00e9 uma \u00e1rea em franca evolu\u00e7\u00e3o, o que pode ser ilustrado pelos 638 estudos recentemente completados ou em andamento registrados no 99mTc-pirofosfato, que permite a obten\u00e7\u00e3o de imagens moleculares do ac\u00famulo de fibras amiloides no mioc\u00e1rdio, temos como perspectivas futuras o uso de imagens de PET usando ,Na \u00e1rea da imagem cardiovascular para diagn\u00f3stico da AC, al\u00e9m da cintilografia card\u00edaca com radiotra\u00e7adores \u00f3sseos, como o ,Trabalhos recentes t\u00eam se concentrado na an\u00e1lise do transcriptoma, em busca de diferen\u00e7as de express\u00e3o entre indiv\u00edduos saud\u00e1veis e doentes, com an\u00e1lise molecular e gen\u00f4mica integrativa. O transcriptoma de pacientes com amiloidose AL \u00e9 mais semelhante ao dos pacientes com gamopatia monoclonal de significado indeterminado. Al\u00e9m disso, o n\u00edvel de microRNA circulante, que conhecido por se correlacionar com dano card\u00edaco, \u00e9 aumentado nos pacientes AL. Por meio da an\u00e1lise de componentes principais, t\u00eam sido demonstrados perfis fenot\u00edpicos altamente sobrepostos entre AL e gamopatia monoclonal de significado indeterminado e mieloma m\u00faltiplo.Adicionalmente, a intelig\u00eancia artificial (IA) aplicada na an\u00e1lise de dados dispon\u00edveis de banco de dados de prontu\u00e1rios m\u00e9dicos emerge como uma estrat\u00e9gia promissora para identificar indiv\u00edduos com sinais de alerta e permitir o diagn\u00f3stico mais precoce da AC, com potencial para reduzir o atraso para o in\u00edcio dos tratamentos modificadores da evolu\u00e7\u00e3o da doen\u00e7a.anti-sense no ensaio cl\u00ednico CARDIO-TTRansform, testando um novo f\u00e1rmaco de segunda gera\u00e7\u00e3o, AKCEA-TTR-LRx .O desenvolvimento de novas terapias espec\u00edficas para AC por ATTR \u00e9 um campo intenso de investiga\u00e7\u00e3o. Terapias de silenciamento g\u00eanico da TTR que se mostraram efetivas para o tratamento da polineuropatia amiloide heredit\u00e1ria est\u00e3o sendo atualmente testadas em grandes estudos multic\u00eantricos em pacientes com AC por ATTR, incluindo as plataformas de RNA de interfer\u00eancia, com o patisiran no estudo APOLLO-B ; e a tecnologia de oligonucleot\u00eddeos flutter atrial exibem alto risco de eventos cardioemb\u00f3licos e tem sido recomendado o tratamento com anticoagulantes. Essas interven\u00e7\u00f5es, apesar de terem um racional fisiopatol\u00f3gico para aplica\u00e7\u00e3o em pacientes com amiloidose, ainda precisam ser testadas em ensaios cl\u00ednicos adequados, sendo \u00e1reas importantes para investiga\u00e7\u00e3o cl\u00ednica futura. Al\u00e9m disso, o transplante card\u00edaco tem se mostrado como estrat\u00e9gia segura nesses pacientes, mas o uso de ventr\u00edculo artificial e a possibilidade de combina\u00e7\u00e3o das terapias devem ainda ser analisados em ensaios cl\u00ednicos adequados.Devido ao potencial arritmog\u00eanico e \u00e0 les\u00e3o do sistema de condu\u00e7\u00e3o associados ao dep\u00f3sito amiloide, \u00e9 frequente o uso de dispositivos implant\u00e1veis como marca-passo ou cardiodesfibriladores implant\u00e1veis como estrat\u00e9gia de redu\u00e7\u00e3o de mortalidade e aumento de sobrevida nessa popula\u00e7\u00e3o. Por outro lado, os indiv\u00edduos com AC e fibrila\u00e7\u00e3o ou \u00c9 fundamental que esperemos os resultados de estudos distintos testando diferentes interven\u00e7\u00f5es e terapias espec\u00edficas, para que possamos definir as melhores op\u00e7\u00f5es e combina\u00e7\u00f5es de tratamento para essa doen\u00e7a, tendo em mente que: 1) n\u00e3o conhecemos completamente todos os detalhes da sua fisiopatologia; 2) n\u00e3o temos compreens\u00e3o e experi\u00eancia ampla de como as medica\u00e7\u00f5es funcionam nessa doen\u00e7a; 3) ainda n\u00e3o temos uma avalia\u00e7\u00e3o acurada, detalhada e de longo prazo dos riscos e benef\u00edcios dos diferentes tratamentos; 4) e tamb\u00e9m n\u00e3o conhecemos a rela\u00e7\u00e3o entre dose e resposta das diferentes medica\u00e7\u00f5es para esse cen\u00e1rio cl\u00ednico.Dessa forma, julgamos que s\u00e3o a\u00e7\u00f5es relevantes e fundamentais a serem implantadas:Cria\u00e7\u00e3o de novos centros de refer\u00eancia/excel\u00eancia em AC.Capacita\u00e7\u00e3o de profissionais para o reconhecimento precoce e encaminhamento para centros especializados.Est\u00edmulo para elabora\u00e7\u00e3o de um Registro Nacional de Amiloidose Card\u00edaca.Organiza\u00e7\u00e3o e debate dos desafios para uma \u201cJornada do Paciente com Amiloidose\u201d com seguran\u00e7a e qualidade.Discuss\u00e3o de novos modelos de remunera\u00e7\u00e3o e de aten\u00e7\u00e3o na AC.Incentivo \u00e0 pesquisa cl\u00ednica da AC em nosso pa\u00eds. Significant advances in understanding cardiac amyloidosis (CA) have been made in recent years, leading to a thorough reformulation of its clinical significance. In addition to convincing evidence that CA is a relatively common cause of heart failure with preserved ejection fraction (HFpEF), we are witnessing the emergence of specific therapies that can change the course of the disease and prolong the survival of affected patients.In parallel, relevant advances in cardiovascular imaging techniques have greatly contributed to earlier and more accurate identification of the disease. Cardiac scintigraphy with bone-seeking radiotracers has allowed non-invasive diagnosis of transthyretin (TTR) cardiac amyloidosis (ATTR-CA), eliminating the need for endomyocardial biopsy, which has greatly simplified the diagnostic flow.Thus, we aim to present the most current recommendations for the diagnosis, prognostic staging, and treatment of CA based on a critical review of the current scientific evidence.In this position paper, the recommendations and levels of evidence are classified according to the following parameters:\u2013Systemic amyloidosis is caused by tissue deposition of fibrillar and insoluble protein aggregates in different organs, including the heart, which leads to organ dysfunction.TTR is a protein composed of four monomers, which circulate as a tetramer.,In ATTRwt, the amino acid sequence is normal and the process by which the wild-type protein becomes unstable and aggregates into amyloid fibrils is not completely clear. However, aging appears to be involved in its pathophysiology.In the AL form, amyloidogenic light chains originate from plasma cells or, less frequently, abnormal B lymphocytes. Thus, it is a clonal and neoplastic hematologic disease. In the heart, the deposition of amyloid fibrils causes structural damage by increasing cardiac and vascular rigidity, impairing cardiac contraction and relaxation and creating conduction disturbances. In parallel, circulating light chains are directly toxic to the myocardium through lysosomal dysfunction, defective autophagy, production of reactive oxygen species, cell and mitochondrial dysfunction, alterations in cardiomyocyte calcium homeostasis and, finally, cell death.\u2013Different subtypes of amyloidosis can lead to overlapping clinical manifestations and, once diagnosed, it is essential to correctly characterize the precursor protein to determine a specific treatment.Depending on the affected organs and degree of dysfunction, a wide spectrum of clinical manifestations can be observed, with a progressive and potentially fatal evolution. The main organs affected by systemic amyloidosis are the heart, kidneys, eyes, central and peripheral nervous system, and liver. Nonspecific clinical manifestations are frequently observed and include fatigue, weight loss, peripheral edema and orthostatic hypotension. For this reason, late diagnosis is common. Thus, knowledge of the disease and a high degree of clinical suspicion are necessary to complete the diagnosis.In ATTRv, depending on the mutation, the clinical picture is dominated by neuropathy or heart disease. In ATTRwt, heart disease is the main clinical manifestation, occurring mainly in elderly men who develop HFpEF without previously known risk factors.,Some extracardiac alterations may precede CA by several years, especially bilateral carpal tunnel syndrome and spontaneous rupture of the biceps tendon. It is essential to recognize such signs as part of the clinical picture of amyloidosis, which could lead to earlier diagnosis and specific treatments that could prevent the progression of heart disease.There is a higher incidence of ATTRwt in older patients, usually those over 70 years of age. However, since the clinical manifestations of ATTRv also usually occur in older adults, age should not be taken into account when differentiating between the two forms of ATTR. Regarding sex, there is a strong predominance (80% to 90%) of ATTRwt in men.,,,Regarding ATTRv, V30M is the most widespread mutation worldwide, being endemic in Portugal, Sweden, and Japan. It is probably the most common form in Brazil. Another common mutation is V122I, which is present in 3.4% of African-Americans and is related to the development of heart disease in patients over 60 years of age.,The incidence of AL-CA is 6-10/million people/year and it is considered the main cause of CA.Thus, CA could be considered an underdiagnosed condition, rather than a rare disease. Recent data from the USA indicate a progressive increase in CA prevalence ,,,Regarding prognosis, AL affects multiple organs and is more aggressive than other subtypes. Late diagnosis is associated with high early mortality in the first 6 to 12 months due to advanced heart disease complications.Mutations in the TTR gene are associated with a wide variety of clinical manifestations, which reflect the deposition of the variant protein in different types of tissues. Cardiac and peripheral nervous system tissue are the most frequent; the former is particularly associated with the V122I mutation, and the latter with the V30M mutation.In this chapter, we will describe the main neurological manifestations suggestive of ATTRv.Neurological manifestations in ATTRv can be divided into peripheral neuropathy, ie, late manifestations of central nervous system involvement linked to amyloid angiopathy, and central nervous system manifestations associated with oculoleptomeningeal infiltration.,The involvement of the peripheral nerves in ATTRv is typically mixed sensory and motor length-dependent axonal neuropathy, ie, it initially affects more distal segments of the limbs, especially the lower ones, progressing to the proximal segments and upper limbs.,In its early onset form (< 50 years of age), it is usually associated with the ATTRv V30M (V50M) mutation, and thin fibers with little or no myelination are initially affected. This is followed, in the degree that the disease progresses, by thick fibers, which are very myelinated and responsible for vibratory, postural-kinetic, and motor sensitivity. The initial symptoms are erectile dysfunction, early satiety, nausea, vomiting, diarrhea, constipation, alternating diarrhea with constipation, orthostatic hypotension, syncope, arrhythmias, altered atrioventricular conduction, dry eye, urinary retention or incontinence, neuropathic pain, lost sensitivity to heat and cold, and significant weight loss. The initial phase can include painless lesions, plantar perforating ulcers and their repercussions, such as localized infections, cellulitis, osteomyelitis, and even septicemia. After a few years, gait instability and muscle atrophy appear, always evolving from distal to proximal segments.In its late forms, neuropathy compromises all types of fibers, although dysautonomia is not as important, at least in the initial phase. These forms can be associated with V30M or a number of other mutations, and the evolution is usually more aggressive. In a Brazilian study, 26% of ATTRv patients with V30M had late onset.Bilateral carpal tunnel syndrome is a frequent manifestation in ATTRv and may be the initial manifestation. Although it can be associated with any mutation, it is particularly important in some of them, including TTR V122I. This mutation appears to be frequent in Brazil and is associated with heart disease, which it can precede by several years.Prolonging survival, which was initially associated with liver transplantation and is currently possible with new drugs, has enabled the appearance of previously uncommon manifestations. The long-term production of TTR by the choroid plexus is associated with both amyloid angiopathy and meningeal infiltration. Amyloid angiopathy manifests as focal stroke-like, transient ischemic attack-like, or aura-like episodes, as well as an irritative epileptic type. In more severe cases, ischemia or even intracranial hemorrhaging can occur. The following neurological manifestations stand out: hearing impairment, migraine, dementia, cerebellar syndrome, myelopathy, and radiculopathy.Some rare mutations have a predilection for oculocerebral involvement and lead to oculoleptomeningeal amyloidosis .Stages of neuropathy: The Coutinho stages for ATTRv are classified according to polyneuropathy. In stage 1 sensorimotor polyneuropathy affects gait, but walking support is not necessary. In stage 2, one or more support devices are needed for walking. In stage 3, the patient is confined to bed or a wheelchair.ATTRv is a disease of autosomal dominant inheritance but variable penetrance. It is mutation- and age-dependent, as well as regionally influenced.At least 140 different mutations have been described to date, but not all are pathogenic. Some polymorphisms are well-defined, while the significance of others is still indeterminate.A special topic among genetic aspects is pre-symptomatic testing, ie, testing the relatives of individuals known to be affected. Unlike diagnostic testing, it must be performed by specialized personnel and there must be a support team, including a psychologist. It must include a preparation phase, pre-diagnosis, genetic testing and a post-result support phase. Such testing should not be performed on children and should only be applied to individuals who expressly state that it is their wish and are considered psychologically prepared for the results.\u2013CA progresses as the cardiac extracellular matrix is infiltrated by amyloid fibrils, resulting in a progressive increase in the thickness of the ventricular wall and a marked increase in chamber stiffness, resulting in impaired diastolic function, which leads to heart failure with restrictive physiology.Thus, the most frequent clinical manifestation is heart failure syndrome, most commonly with preserved ejection fraction (HFpEF), although a drop in ejection fraction can occur in more advanced stages of the disease. The clinical syndrome may present predominant left HF symptoms with pulmonary congestion , right HF symptoms , or both sets of symptoms. Cardiac amyloidosis should be considered in the differential diagnosis of HFpEF etiology in older men,Syncope and orthostatic hypotension are common symptoms and indicate the presence of dysautonomia. One typical clinical aspect that may raise the suspicion of amyloidosis is the need for dose reduction or discontinuation of antihypertensive drugs in patients previously diagnosed with systemic arterial hypertension, especially beta-blockers and angiotensin-converting enzyme inhibitors/angiotensin receptor blockers.Amyloid infiltration can also occur, causing cardiac conduction system disease from the early stages, with variable degrees of atrioventricular block, which results in high-risk bradycardia in some cases, requiring pacemaker implantation. Another important change is the hardening of the atrial walls, which leads to high rates of atrial arrhythmias, including atrial fibrillation, as well as atrial thrombi, with cardioembolic stroke being a common clinical manifestation, even in individuals with sinus rhythm. Complex ventricular arrhythmias seem to be frequent in advanced stages of the disease, an aspect that is best documented in AL amyloidosis.CA, particularly the ATTR type, is often underdiagnosed due to factors associated with the medical evaluation, in addition to the characteristics of the disease itself, including: fragmented knowledge among different specialties and subspecialties, a scarcity of centers and specialists dedicated to the management of this disease, the mistaken belief that CA is a rare and incurable disease, and the phenotypic and genotypic heterogeneity of ATTR-CA.,Thus, recognizing \u201cred flags\u201d can help diagnose CA in HF patients,,Bilateral carpal tunnel syndrome, often one of the first indicators of ATTR-CA, is the most common non-cardiac manifestation and can precede symptoms of HF by several years. A recent study found that approximately 50% of individuals with ATTRwt had carpal tunnel syndrome 5 to 7 years prior to diagnosis.Other warning signs may emerge from typical changes in routine complementary cardiac examinations, which are covered as specific topics in this document.It should also be pointed out that CA can often simulate other heart diseases. Amyloidosis should be considered one possible etiology for patients with a hypertrophic cardiomyopathy phenotype, particularly if it developed after 60 years of age. The asymmetrical pattern of myocardial hypertrophy in ATTR-CA patients differs from that of AL-CA patients, which is usually symmetrical. In a study that compared 263 confirmed ATTR-CA patients with 50 AL-CA patients, among the ATTR cases, asymmetric hypertrophy was present in 79%, symmetric hypertrophy in 18%, and no myocardial hypertrophy in 3%.Older patients with severe low-flow/low-gradient aortic stenosis may have CA in 10% to 15% of cases, with an unfavorable prognosis.The electrocardiogram (ECG) is an essential test for diagnostic assessment and therapeutic planning, and its interpretation in conjunction with clinical and echocardiographic information is important. Although low-voltage ECG has great specificity in diagnosing myocardial infiltration secondary to CA, this is not the most prevalent finding. A lack of R-wave progression in precordial leads, which simulates an electrically inactive anteroseptal zone (pseudoinfarction pattern) is a much more frequent finding, with a prevalence of 60% to 70% in confirmed CA diagnoses, regardless of the type .In ATTR-CA, less than 40% of patients with a biopsy-confirmed diagnosis have low-voltage ECG.,Thus, without low-voltage ECG criteria or signs of left ventricle (LV) overload, diagnostic suspicion of CA should not be ruled out, especially ATTR-CA. Disproportionate voltage in relation to myocardial thickness is also an important warning sign, reaching a prevalence of 73% to 80% in CA patients, regardless of type.Among cardiac rhythm alterations, atrial fibrillation is more prevalent in ATTR patients, as well as atrioventricular blocks.,Echocardiography should be performed in all patients with clinical suspicion of the disease. Classic CA findings are usually present at an advanced stage of the disease and characterize a restrictive, infiltrative-type cardiomyopathy. The dimensions of the LV are not increased, volumes are normal or reduced, and there is a thickening of the ventricular walls. Increased atrial dimensions are common, reflecting early and progressive diastolic dysfunction, with increased filling pressures. Atrioventricular valves may be thickened and mitral and tricuspid regurgitations are functional. There may also be signs suggestive of infiltration of the interatrial septum, as well as an increase in the pulmonary artery systolic pressure. The right ventricle (RV) may also be affected. Pleural and pericardial effusions are very common and, in cases involving intense tissue infiltration, a granular sparkling appearance can be observed 35,54The LV ejection fraction is normally preserved until the more advanced stages of CA, although the longitudinal contractile function is reduced early.Assessing the ventricular diastolic filling pattern is essential and often demonstrates some degree of diastolic dysfunction. In the initial phases, alterations compatible with type I diastolic dysfunction can be observed may develop as a result of increased left atrial pressure.In more advanced phases of the disease, there is a restrictive pattern of ventricular filling .\u2013,Analyzing myocardial deformation allows the early identification of signs of myocardial dysfunction in relation to LV ejection fraction.Other analyses of myocardial deformation, such as RV systolic function,,Cardiovascular magnetic resonance (CMR) allows accurate assessment of myocardial tissue changes in CA.The global increase in myocardial tissue water content leads to an increase in mean hydrogen relaxation time, whether T1 or T2 (transverse).,\u2013,Thus, the combination of myofibril deposition and interstitial fibrosis can be detected easily and precisely, and can even be quantified by late enhancement (LE) techniquesThe CMR contrast medium is based on gadolinium, which is bound to a macromolecular chelator that does not allow it to pass through the entire cell membrane. Thus it is distributed exclusively in the myocardial extracellular volume. The distribution pattern in the LE image can raise suspicion of CA .,CA can modify the appearance of all cardiac chambers.,,,,CA is also commonly associated with increased myocardial thickness, which is, in most cases, more expressive than in cases secondary to hypertension. The thickness of the cardiac muscle is generally greater in ATTR-CA than AL-CA.As a result of diastolic dysfunction, it is not uncommon to observe pericardial or pleural effusions.,The LE technique after gadolinium contrast injection has been widely recognized as a pillar of CA imaging diagnosis.In a series of 250 patients with different forms of amyloidosis, LE with a transmural pattern was associated with a 5.4 times greater risk of death .,,,,,,,,,Different groups have investigated the utility of cardiovascular magnetic resonance-derived T1 maps to improve diagnostic and prognostic performance in CA.\u2013Technetium-99m-labeled bisphosphonate-derived radiotracers, originally developed for bone imaging, have found a new role as a non-invasive diagnostic tool for ATTR-CA.99mTc-labeled bone radiotracers used in ATTR-CA diagnosis are 99mTc-pyrophosphate, 99mTc-DPD , and 99mTc-HMDP (99mTc-labeled hydroxymethylene diphosphonate).\u201399mTc-pyrophosphate is the only one of these available in Brazil. It should be pointed out that although 99mTc-MDP (99mTc-labeled methylene diphosphonate) has proven efficient for bone scintigraphy, it has low sensitivity for diagnosing ATTR-CA and should not be used for this purpose.The main 99mTc-pyrophosphate binds in the heart is unknown, it is widely accepted that a calcium-dependent uptake mechanism is involved.99mTc-pyrophosphate uptake in the myocardium is probably related to the presence of microcalcifications.99mTc-pyrophosphate uptake, while AL-CA has little or no affinity for bone radiotracers. In addition, ATTR-CA has a more indolent evolution, providing more microcalcifications and, consequently, greater radiotracer accumulation.Although the structural component of the amyloid deposition to which 99mTc-pyrophosphate cardiac scintigraphy: it is a simple, easy, widely available method with low dosimetry that can differentiate ATTR-CA from AL-CA noninvasively and with high specificity, and thus guide treatment. This differentiation is useful, since AL-CA and ATTR-CA have completely different prognostic and therapeutic implications.Although the echocardiogram and cardiac magnetic resonance findings may be indicative of CA, they cannot differentiate ATTR-CA from AL-CA. This is the main advantage of 99mTc-pyrophosphate had an 88% sensitivity and 88% specificity for ATTR when only visual assessment was used (score \u2265 2).99mTc-pyrophosphate uptake was predictive of overall mortality and hospitalization for HF. In these studies, combined assessment of the intensity of radiotracer uptake in the myocardium with anatomical, functional and biomarker variables improved risk stratification.The role of bone radiotracers in ATTR diagnosis was recently re-evaluated by an international group of several centers with expertise in CA: in a cumulative analysis of 1,217 patients, 867 with biopsy-confirmed amyloidosis and 360 with non-amyloid cardiomyopathy, scintigraphy was highly sensitive (99%) and specific (86%) for ATTR-CA.99mTc-pyrophosphate (dosimetry: 3.2 mSv for the whole body for 15 mCi). Flat and single photon emission computed tomography (SPECT) images of the chest are taken 1 and 3 hours after administration of the radiopharmaceutical.No preparation required. Images are obtained after intravenous administration of 10 to 25 mCi (370 to 925 Mbq) of Planar chest images: can be obtained in the anterior, left anterior oblique, and left lateral projections using 99mTc photopic , low energy/high resolution collimator and a 256 \u00d7 256 matrix, 500,000-750,000 counts.SPECT chest images: obtained with a 128 \u00d7 128 matrix (64 \u00d7 64 is acceptable), 180-degree rotation from right anterior oblique to left posterior oblique (360-degree is acceptable), 1 image every 3 to 6 degrees. If available, SPECT/CT images (SPECT combined with computed tomography) provide greater confidence for interpreting the images.Minimum recommended images: 1 h SPECT and 1 h and 3 h flat images in the anterior projection. The use of SPECT images is recommended to differentiate diffuse uptake of the radiopharmaceutical in the myocardium from its persistence in the blood pool, focal uptake by myocardial infarction, and bone overlay. In addition, 1 h and 3 h flat images are useful for quantifying and monitoring the blood pool washout, which is variable, eg, much slower in patients with renal failure.Semiquantitative analysis (1 h planar image): For 99mTc-pyrophosphate, the semiquantitative analysis was defined as the ratio between uptake in the cardiac projection and uptake in the contralateral hemithorax, measured in a planar 1 h image in the anterior view. To do this, a circular area of interest is drawn over the cardiac projection without including the sternum, avoiding the inclusion of both the adjacent lung and areas of focal uptake in the costal arches. An identical (\u201cmirror\u201d) area of interest is placed in the contralateral hemithorax following the same procedure. The count within the cardiac area of interest is divided by the count in the contralateral area of interest, thus obtaining the heart/contralateral (H/CL) ratio. H/CL \u2265 1.5 at 1 h identifies ATTR-CA with high accuracy if systemic AL-CA has been excluded,Visual graduation (3 h image): visual grading is performed by comparing cardiac uptake with physiological uptake in adjacent costal arch, and can be performed with planar images in the anterior projection, in SPECT images, or even in whole-body images obtained 3 h after injecting the radiopharmaceutical. Uptake intensity is defined as grade 0 , grade 1 , grade 2 , and grade 3 . Grades 2 or 3 are strongly suggestive of ATTR if monoclonal gammopathy has been excluded entails a risk of inaccurate diagnosis. The most common cause of ATTR misdiagnosis is AL-CA. Recent studies indicate that up to 22% of patients with AL-CA may have grade 2 or 3 uptake in ,Finally, scintigraphy is not recommended for patient follow-up, since there is no current evidence of a correlation between a change in image pattern and disease progression or response to treatment.No specific laboratory marker can be used to diagnose CA.Troponin and natriuretic peptides have been found useful for assessing cardiac damage due to amyloidosis and are non-invasive, accessible, and relatively low-cost diagnostic aids.Analysis of data from the THAOS (Transthyretin Amyloidosis Outcomes Survey) registry showed that natriuretic peptide levels can be used as a diagnostic aid, with higher values observed in mutations associated with amyloid cardiomyopathy, such as V122I and late-onset V30M, than in those predominantly associated with neurologic manifestations, such as early-onset V30M. Higher levels of biomarkers were also observed in ATTRwt than ATTRv. However, it was observed that even in patients with a predominantly neurological phenotype, 45% to 90% had altered levels of these biomarkers, indicating some degree of subclinical myocardial involvement.Higher NT-proBNP values have been observed in AL-CA than ATTR-CA. This is because amyloidogenic light chains modulate p38 mitogen-activated protein kinase, which directly promotes NT-proBNP expression. Thus, despite the same degree of hemodynamic changes in both forms of amyloidosis, serum levels of NT-proBNP may be higher in AL-CA.Of note, patients with HFpEF due to ATTR have disproportionately high NT-ProBNP values with respect to HF severity compared to patients with non-amyloid HFpEF.Mild and persistent elevation of troponin levels is frequently observed and suggests subclinical myocardial damage in several non-ischemic cardiomyopathies.Several mechanisms have been postulated to explain the elevated troponin levels in these patients: myocardial ischemia, increased wall stress, direct myocyte damage by inflammatory cytokines and/or oxidative stress, neurohormonal activation, and microvascular dysfunction in heart failure. Microvascular dysfunction in amyloidosis is presumably caused by interstitial and perivascular deposition, increased ventricular filling pressure, and endothelial dysfunction due to immunoglobulin-induced toxicity in AL-CA. In addition, it has also been found that light chains have a direct cardiotoxic effect, regardless of extracellular deposition of fibrils, which might explain the higher troponin levels in AL-CA than in ATTR-CA, in which mild and persistent elevation is usually observed.It is important to point out that many ATTR patients have comorbidities, such as ischemic cardiomyopathy, which can cause altered troponin values. Therefore, troponin levels should not be used to rule out or confirm cardiac involvement. Rather, they should serve as a potential warning sign for the disease, which can be better evaluated through more specific tests.5.5.3A number of other biomarkers have been studied, some with high specificity for amyloidosis subtypes, such as retinol-binding protein 4 for hereditary amyloidosis through the Val142Ile mutation. However, further studies are necessary, as is the commercial availability of tests for routine analysis.Recent evidence indicates that CA, particularly the ATTRwt form, is more prevalent than previously estimated, and this is due in part to widespread underdiagnosis and the fact that this disease mimics other heart diseases, such as hypertrophic cardiomyopathy, non-amyloid HFpEF, and low-flow/low-gradient aortic stenosis.first and most important step is clinical suspicion, which is based on clinical history, a physical examination , and a tissue biopsy must be performed, which is fundamental for confirming amyloid protein deposition and definition of the therapeutic strategy.The patients with positive results for monoclonal light chains must be referred to a hematologist is available and monoclonal light chains are absent, grades 2 or 3 cardiac uptake and a \u2265 1.5 H/CL uptake ratio, with SPECT imaging showing that the increased uptake is in the ventricular walls, confirms ATTR-CA without the need for endomyocardial biopsy. TTR gene sequencing should then be performed to determine whether the ATTR is hereditary or wild type.When Differentiating between hereditary and wild-type ATTR has prognostic and therapeutic implications and is also important for family screening and genetic counseling.When cardiac scintigraphy with bone tracers is negative and is associated with an absence of monoclonal light chains, CA is unlikely. However, when clinical suspicion persists, based mainly on the results of other imaging methods highly suggestive of amyloidosis, endomyocardial biopsy can have a relevant diagnostic role and should be performed. Such cases could indicate ATTRv involving mutations and amyloid deposits, to which bone tracers do not adhere, such as early-onset V30M and P64I, in addition to other unusual types of amyloidosis.bone scintigraphy is unavailable (sub-route 2B), endomyocardial biopsy is recommended to clarify the diagnosis.When AL-CA patients have faster-forming and more toxic amyloid deposits than ATTR-CA patients. As a consequence, monthly increases of 1.45 to 2.16 mm can be observed in myocardial thickness, which are associated with higher elevations of biomarkers, such as troponin and BNP,Recent technological and laboratory advances indicate that the main prognostic determinant in AL-CA is the extent of amyloid deposition in the heart.Additionally, echocardiographic findings such as a marked increase in wall thickness, diastolic dysfunction, LV dysfunction, valve thickening, and reduction in LV global longitudinal strain, combined with BNP and troponin biomarkers and hematological status, are predictors of higher mortality.,\u2013\u2013,CMR also provides data that correlates with survival in CA patients.Biomarkers not only play a role in staging these patients, but also enable treatment response assessment, which is always a challenge in clinical practice, especially for AL-CA, since chemotherapy can be an additional factor in myocardial damage. What has been well established in the literature is the agreement between lower NT-proBNP levels and hematologic response to treatment, as well as improvement in New York Heart Association (NYHA) functional class. The reverse is also true, such as increased levels of NT-proBNP and troponin and reduced ejection fraction. , and CMR, provide complementary data that facilitate prognostic stratification.,,,\u2013Imaging findings, such as echocardiography, cardiac scintigraphy with technetium-pyrophosphate . Thus, specific treatment is based on eliminating this light chains production by eradicating plasma cell clones in bone marrow. The rapid reduction and, ideally, normalization of light chain levels (hematologic response) is a primary goal in AL-CA treatment. The reversal of damage to tissues and organs affected by amyloid deposits (organ response) is the second major objective of treatment. Further goals include improved quality of life and overall survival.AL-CA is caused by the aberrant production of kappa or lambda immunoglobulin light chains , as well as in those treated without this procedure. The median survival for stage I, II, III, and IV patients is 55, 19, 12 and 5 months, respectively, in patients who did not receive or failed HSCT, and 97, 58 and 22 months, respectively for stages II, III and IV, in patients who underwent HSCT.Low-risk patients are candidates for high-dose chemotherapy followed by autologous HSCT . This is,Three months after the end of treatment, the hematological and organ responses must be evaluated with specific tests for monoclonal gammopathy and each affected organ. The hematological and organ response criteria are summarized in Tables Even when a complete hematological response occurs, dysfunction improves more slowly, and may take months or even years to appear after the light chain ratio has been normalized.,,Only 20% of patients diagnosed with AL amyloidosis are eligible for HSCT, although this number could increase with organ response after induction regimens with effective anti-cancer therapies (bortezomib and daratumumab).HSCT eligibility assessment is central to therapeutic success in this strategy. A single prospective randomized study compared HSCT with conventional chemotherapy, finding no overall survival benefit compared to a less intensive strategy. However, in this study the high mortality rate associated with HSCT (24%) was related to the inclusion criteria, the inexperience of the transplant center and the use of a subtherapeutic dose of melphalan.\u2013Since the literature provides no precise well-established prospectively validated HSCT eligibility criteria, each center establishes its own. Although subjective assessment is relevant, HSCT can be guided by certain factors: \u201cphysiological age\u201d \u2264 70 years, creatinine clearance > 30 mL/min , troponin T < 0.06 ng/mL, Eastern Cooperative Oncology Group performance scale \u2264 2, NYHA functional class I or II, and systolic blood pressure > 90 mmHg. Adequate pre-treatment risk stratification, concomitant with the development of better supportive conditions, such as antibiotic therapy and intensive care, has led to a reduction in HSCT-associated mortality in recent decades, with rates of 2.4% to 3.4% in retrospective analyses.,,Therefore, HSCT should be recommended as the first-line therapy in eligible patients . This re,An upfront HSCT can be performed soon after diagnosis in patients with < 10% clonal plasma cells in the bone marrow or when preceded by induction therapy with regimens containing bortezomib and/or anti-CD38 monoclonal antibody. The latter should be considered when there is an association with multiple myeloma, a predictable delay in HSCT, poor performance that may improve with induction therapy, and > 10% infiltration of the bone marrow by clonal plasma cells, which is associated with worse prognosis.The main regimens used as pre-transplant therapy are cyclophosphamide, bortezomib and dexamethasone (CyBorD) and daratumumab, bortezomib, cyclophosphamide and dexamethasone (Dara-CyBorD).Given that new therapies for treating AL amyloidosis are emerging and the scarcity of randomized studies on transplantation, further studies will be needed to determine whether HSCT will remain the most effective therapy in coming decades.Most patients diagnosed with AL-CA are not eligible for higher intensity autologous HSCT therapy due to comorbidities, such as advanced heart disease, renal failure, the involvement of more than two organs, or advanced age. Thus, in this group of patients, chemotherapy with anti-cancer medications is the basis of treatment . The reg,\u2013The CyBorD regimen was found to be highly effective (81% to 94%), well tolerated, and capable of producing a rapid hematological response (within 3 months) in two retrospective studies with small samples (n = 17 and 43). Two-year overall survival reached 92% in these studies.Two recent randomized studies tested chemotherapy combinations as new treatments of choice in AL-CA patients who are ineligible for HSCT. The first compared a regimen of bortezomib, melphalan and dexamethasone to melphalan and dexamethasone, finding that adding bortezomib resulted in a higher overall hematologic response rate after 3 months (79% vs. 52%), a greater cardiac organ response (38% vs. 28%), and improved overall survival, with a 2-fold reduction in mortality.Several steps in the pathophysiological process of amyloid fibril formation and deposition in cardiac tissue are potential therapeutic targets in ATTR-CA, as illustrated in ,In the past, liver transplantation has been proposed as a treatment for patients with ATTRv-associated polyneuropathy.Tafamidis is a small molecule that selectively inhibits the dissociation of TTR tetramers by binding to thyroxine binding sites, thereby effectively inhibiting the cascade that results in amyloid fibril formation.2. The patients were randomized to receive tafamidis 80 mg/day, tafamidis 20 mg/day, or placebo in a 2:1:2 proportion. The study\u2019s primary endpoint was hierarchically assessed all-cause mortality, followed by the frequency of cardiovascular hospitalization over 30 months of follow-up. The main secondary outcomes were change in 6-minute walk test results and quality of life scores according to the Kansas City Cardiomyopathy Questionnaire. The tafamidis 80 and 20 mg groups were merged for the statistical analysis, and the results indicated that these patients (n = 264) had a 30% lower relative risk of all-cause mortality (RR = 0.70 [95% CI: 0 .51-0.96]), 32% fewer cardiovascular hospitalizations (RR = 0.68 [95% CI: 0.56-0.81]), a lower rate of decline in 6-minute walk test results (p < 0.001), and a lower rate of decline in Kansas City Cardiomyopathy Questionnaire scores (p < 0.001) than the placebo group (n = 177). The Kaplan-Meier survival curves showed that tafamidis resulted in lower all-cause mortality, with the curves diverging after approximately 18 months of treatment, a result that agrees with the concept that tafamidis modifies the natural history of the disease. Tafamidis was well tolerated, with a similar incidence of adverse effects in the treatment and placebo groups. In the patient subgroup analysis, tafamidis was associated with lower all-cause mortality than placebo independently of NYHA functional class or ATTR genotype (hereditary or wild-type). However, patients in NYHA functional class III at the time of inclusion who were allocated to the tafamidis group had higher rates of hospitalization than the placebo group, a result likely explained by longer survival during a more severe disease phase. This subgroup analysis demonstrates the need for a trial with a large enough sample size to specifically assess the effect of tafamidis in ATTR patients with more advanced symptoms of heart failure.Tafamidis is the only drug to have been specifically tested in CA patients in a prospective, randomized, placebo-controlled, multicenter clinical trial (ATTR-ACT).More recently, an open-label extension study of ATTR-ACT found that 80 mg/day of tafamidis resulted in significantly higher survival than 20 mg/day .Based on this evidence, tafamidis 80 mg/day is recommended for patients with ATTRv or ATTRwt, with NYHA I to III HF, without severe renal dysfunction, and who are beginning therapy at the earliest stages of the disease . In BrazAG10 is a selective TTR tetramer stabilizer designed to mimic the structural influence of a super-stabilizing mutation (T119M), which significantly reduces the dissociation rate of tetramers.,Therapies based on silencing the expression of genes that encode hepatic TTR production are very promising, including RNA interference (patisiran) and antisense oligonucleotide (inotersen) strategies. Both drugs have been tested in multicenter phase 3 trials in patients with ATTRv polyneuropathy and were shown to be effective in reducing the progression of neurological manifestations.In vitro and experimental studies have shown that certain compounds based on hydrophobic molecules promote the degradation of amyloid tissue deposits, allowing their resorption through the macrophage system.,In addition to specific therapy for amyloidosis, supportive treatment for HF may be necessary. CA initially presents as HFpEF and a restrictive pattern of LV filling, which could lead to disease progression and reduced EF. This pathophysiological mechanism could explain the difficulties in clinical management of CA patients when using HFrEF medications.,Maintaining euvolemia through water restriction and medications is the focus. Loop diuretics, the most frequently used medication, are indicated to reduce pulmonary and systemic congestion, and they may be associated with aldosterone antagonists . Using d,Regarding HFrEF drugs, there is no scientific evidence that neurohormonal antagonists such as angiotensin II-converting enzyme inhibitors, angiotensin II receptor blockers, and beta blockers, or even substances recently described as neprilysin inhibitors, angiotensin II receptor antagonists, and SGLT2 inhibitors, have any effect on CA, and they also involve the risk of hypotension and autonomic dysfunction. In a retrospective, single-center study of 99 CA patients (33% AL-CA and 67% ATTR-CA), Aimo et al.Arrhythmias are very common in CA patients and are usually symptomatic and poorly tolerated. Arrhythmia assessment in this population should involve three different situations: atrial arrhythmias, ventricular arrhythmias, and conduction system disease.,Amyloid deposition leads to atrial thickening, including changes in atrial relaxation and increased intracavitary pressure, which produces atrial dilatation that, in association with atrial fibrosis, predisposes patients to atrial fibrillation (AF) or other atrial arrhythmias. The prevalence of AF in CA patients can range from 11% to 71%, being even higher in ATTR-CA, possibly because it affects older men.Managing AF in these patients is generally difficult, since they usually cannot tolerate drugs such as beta-blockers, non-dihydropyridine calcium channel blockers or digitalis due to the fact that they cause postural hypotension and HF decompensation. Given the need for these medications, it is advisable to use low doses with careful hemodynamic monitoring, in addition to serum level control, if digoxin is used. Regarding rhythm control, a retrospective analysis found no difference in survival between patients who received antiarrhythmic drugs and those treated with frequency control alone.,,,Finally, the reduced contractility caused by amyloid infiltration in atrial tissue may also contribute to thrombus formation. Autopsy studies have found that up to 33% CA patients have intracavitary thrombi,,Ventricular arrhythmias are frequent in CA patients, especially AL-CA. Previous studies have detected complex ventricular arrhythmias in at least 50% of AL-CA patients, in whom non-sustained ventricular tachycardia was the most frequent arrhythmia and was associated with lower survival.\u2013In this context, implantable cardioverter-defibrillators (ICD) may have a role in preventing sudden death in CA patients. ICD may benefit patients with unstable ventricular tachycardia or who have survived cardiac arrest without a reversible cause and a life expectancy > 1 year with significant quality.However, recommending ICD as primary prevention is difficult for a number of reasons. The first is that most causes of sudden death are related to electromechanical dissociation and not ventricular arrhythmias.Conduction-system disease is highly prevalent among CA patients, with atrioventricular conduction being more commonly affected than the sinus node. The involved pathophysiology has not yet been fully defined, although small studies have suggested that the amyloid protein conduction system is involved.,In advanced HF associated with CA, advanced support strategies, such as mechanical circulatory assistance and transplantation, present challenges, especially since it is a multisystem disease. Moreover, due to the reduced size of the left ventricular cavity and the frequent involvement of the right ventricle, the use of long-term mechanical circulatory assistance devices may be limited.Access to care and a sustained commitment to improving care quality are fundamental for achieving excellent results, which are critical for complex diseases such as CA. Developing a course of care that provides the right care at the right time for CA requires the involvement of public and private health care managers, health professionals, and everyone involved in the care of these patients.\u2013CA, particularly AL-CA, must be approached with a view to rapid diagnosis, and multidisciplinary teams with proven experience and clinical protocols based on the best scientific evidence must be available. In addition, comprehensive care and the monitoring and publishing of care results should be promoted. An integrated and collaborative practice model should be observed at these reference centers, with a telemedicine structure to support diagnosis at more distant centers, patient monitoring, and clinical research, which is the basis for emerging therapies. Such initiatives have been undertaken at a number of centers worldwide, including some in Brazil.Additionally, such reference centers should contribute to a National Registry of Cardiac Amyloidosis, allowing better insight into regional epidemiology and care quality, in addition to measuring patient-centered clinical outcomes and contributing to new public policies.,New financing models for high-cost rare disease treatments have been discussed, tested in pharmacoeconomic studies, and implemented around the world.An assessment of new treatments for rare diseases by the National Commission for Technological Incorporation in the Unified Health System (CONITEC) indicated that approximately 52% of the evaluated medicines have already been incorporated. In our vision were are building a new, technologically-based scenario developed through robust data with the support of clinical protocols and reference centers that can replace the current model, where access is sought through judicialization, which has a heavy impact on the federal budget.Despite recent advances in CA research, there are still numerous knowledge gaps to be filled regarding this complex and multifaceted disease. Both the diagnosis and treatment of amyloidosis are evolving, which is exemplified by the 638 studies registered in the clinicaltrials.gov site that have recently been completed or are in progress.99mTc-pyrophosphate, which allows molecular images of amyloid fibril deposition in the myocardium. Although not yet available in Brazil, another promising technique is PET imaging with ,Current cardiovascular imaging for CA diagnosis involves cardiac scintigraphy with bone-seeking radiotracers, such as ,Recent studies have focused on transcriptome analysis, seeking differences in expression between healthy and sick individuals by means of molecular analysis and integrative genomics. The transcriptome of AL-CA patients is similar to that of patients with monoclonal gammopathy of undetermined significance. Furthermore, the level of circulating microRNA, which is known to correlate with cardiac damage, is increased in AL patients. Through principal component analysis, highly overlapping phenotypic profiles have been found between AL, monoclonal gammopathy of undetermined significance, and multiple myeloma.Additionally, using artificial intelligence to analyze data in medical databases is a promising strategy for identifying individuals with warning signs. This strategy could lead to earlier CA diagnosis and reduce treatment delay.New therapies specifically for ATTR-CA are under intense investigation. TTR gene silencing therapies that have been found effective for hereditary amyloid polyneuropathy are currently being tested in ATTR-CA patients in large multicenter studies. The APOLLO-B study is testing the RNA interference agent patisiran . The CARDIO-TTRansform clinical trial is testing antisense oligonucleotide technology in a new second-generation drug, AKCEA-TTR-LRx .Since amyloid deposition has arrhythmogenic potential and can damage the conduction system, implantable devices such as pacemakers or defibrillators are frequently used to reduce mortality and increase survival in this population. On the other hand, individuals with CA and atrial fibrillation or flutter are at high risk for cardioembolic events, and treatment with anticoagulants has been recommended. Although there is a pathophysiological rationale for using these interventions in CA patients, they must still be tested in appropriate clinical trials, and this is an important area for future clinical investigation. In addition, heart transplantation has proven to be a safe strategy in these patients, although artificial ventricles and combined therapies must still be assessed in clinical trials.To define the best treatment options and combinations for this disease, we must wait for the results of studies that are testing different interventions and specific therapies, bearing in mind that: 1) we are not fully aware of the details of the disease\u2019s pathophysiology; 2) we lack a broad understanding of how medications work in this disease; 3) we still do not have an accurate, detailed and long-term assessment of the risks and benefits of different treatments; 4) and we also do not know the dose-response relationship of different medications for this clinical scenario.Thus, we believe that the following basic steps are relevant and should be implemented:Creating new centers of reference/excellence in CA.Training professionals in early CA recognition and referral to specialized centers.Promoting the development of a National Registry of Cardiac Amyloidosis.Discussing the safety and care quality challenges involved in the journey of CA patients.Discussing new remuneration and care models for CA.Encouraging clinical research on CA in Brazil."} +{"text": "Realiza\u00e7\u00e3o:Departamento de Ergometria, Exerc\u00edcio, Cardiologia Nuclear e Reabilita\u00e7\u00e3o Cardiovascular da Sociedade Brasileira de CardiologiaConselho de Normatiza\u00e7\u00f5es e Diretrizes (2020-2021):Brivaldo Markman Filho, Antonio Carlos Sobral Sousa, Aurora Felice Castro Issa, Bruno Ramos Nascimento, Harry Correa Filho, Marcelo Luiz Campos VieiraCoordenador de Normatiza\u00e7\u00f5es e Diretrizes (2020-2021):Brivaldo Markman FilhoCoordenadores da Diretriz:Tales de Carvalho e Mauricio MilaniSum\u00e1rio1. Introdu\u00e7\u00e3o 9461.1. Classes (Graus) de Recomenda\u00e7\u00e3o 9471.2. N\u00edveis de Evid\u00eancia 9472. Estrutura de um Programa de Reabilita\u00e7\u00e3o Cardiovascular 9472.1. Equipe e Responsabilidades dos Profissionais 9472.1.1. M\u00e9dico Assistente 9472.1.2. M\u00e9dico-l\u00edder no Programa de Reabilita\u00e7\u00e3o Cardiovascular 9472.1.3. Outros Profissionais 9472.1.4. Fisioterapeutas e Profissionais de Educa\u00e7\u00e3o F\u00edsica 9472.1.5. Profissional de Enfermagem 9472.2. Estrutura F\u00edsica de um Servi\u00e7o de Reabilita\u00e7\u00e3o 9482.2.1. Aspectos Gerais 9482.2.2. Equipamentos para a Pr\u00e1tica de Exerc\u00edcios F\u00edsicos 9482.2.2.1. Exerc\u00edcios Aer\u00f3bicos 9482.2.2.2. Exerc\u00edcios de Fortalecimento Muscular 9482.2.2.3. Outros Exerc\u00edcios 9482.2.3. Monitoramento 9482.2.4. Seguran\u00e7a 9483. Fases da Reabilita\u00e7\u00e3o Cardiovascular e Estratifica\u00e7\u00e3o de Risco 9493.1. Risco Cl\u00ednico Alto 9493.2. Risco Cl\u00ednico Intermedi\u00e1rio 9503.3. Risco Cl\u00ednico Baixo 9514. Custo-efetividade da Reabilita\u00e7\u00e3o Cardiovascular 9525. Reabilita\u00e7\u00e3o Cardiovascular Domiciliar 9536. Reabilita\u00e7\u00e3o Cardiovascular Integrando o Tratamento Cl\u00ednico Pleno das Doen\u00e7as Cardiovasculares 9536.1. Recomenda\u00e7\u00f5es Gerais para Incremento da Atividade F\u00edsica e Pr\u00e1tica de Exerc\u00edcios F\u00edsicos 9546.2. Hipertens\u00e3o Arterial Sist\u00eamica 9556.2.1. Benef\u00edcios Terap\u00eauticos dos Exerc\u00edcios F\u00edsicos 9556.2.2. Indica\u00e7\u00f5es de Exerc\u00edcios F\u00edsicos na Hipertens\u00e3o Arterial Sist\u00eamica 9566.2.3. Avalia\u00e7\u00e3o Pr\u00e9-participa\u00e7\u00e3o 9566.2.4. Particularidades na Prescri\u00e7\u00e3o e no Acompanhamento dos Exerc\u00edcios F\u00edsicos 9576.3. Coronariopatia Est\u00e1vel ap\u00f3s Evento Agudo ou Revasculariza\u00e7\u00f5es 9576.3.1. Benef\u00edcios Terap\u00eauticos dos Exerc\u00edcios F\u00edsicos 9586.3.2. Quando Indicar Reabilita\u00e7\u00e3o 9586.3.3. Avalia\u00e7\u00e3o Pr\u00e9-participa\u00e7\u00e3o e Prescri\u00e7\u00e3o de Exerc\u00edcios 9596.3.4. Particularidades na Prescri\u00e7\u00e3o e Acompanhamento dos Exerc\u00edcios F\u00edsicos 9606.3.4.1. Angina Refrat\u00e1ria 9606.3.4.2. Treinamento com Indu\u00e7\u00e3o de Isquemia Mioc\u00e1rdica 9606.3.4.3. Ajustes de F\u00e1rmacos Diante da Assimila\u00e7\u00e3o do Treinamento F\u00edsico 9606.4. Insufici\u00eancia Card\u00edaca 9616.4.1. Prescri\u00e7\u00e3o dos Exerc\u00edcios F\u00edsicos e Avalia\u00e7\u00e3o Pr\u00e9-participa\u00e7\u00e3o 9616.4.2. Considera\u00e7\u00f5es Finais sobre a Insufici\u00eancia Card\u00edaca 9636.5. Transplante Card\u00edaco 9636.5.1. Benef\u00edcios dos Exerc\u00edcios F\u00edsicos 9636.5.2. Avalia\u00e7\u00e3o Pr\u00e9-participa\u00e7\u00e3o e Particularidades 9646.5.3. Prescri\u00e7\u00e3o do Treinamento F\u00edsico 9646.5.4. Reabilita\u00e7\u00e3o Cardiovascular Domiciliar 9656.5.5. Recomenda\u00e7\u00f5es 9656.6. Miocardiopatias 9666.6.1. Miocardiopatia Hipertr\u00f3fica 9666.6.1.1. Benef\u00edcios Terap\u00eauticos do Exerc\u00edcio F\u00edsico 9666.6.1.2. Quando Indicar Exerc\u00edcios F\u00edsicos 9676.6.1.3. Avalia\u00e7\u00e3o Pr\u00e9-participa\u00e7\u00e3o 9676.6.1.4. Particularidades na Prescri\u00e7\u00e3o e no Acompanhamento dos Exerc\u00edcios F\u00edsicos 9686.6.2. Miocardite 9686.6.3. Outras Miocardiopatias 9696.6.3.1. Cardiomiopatia Arritmog\u00eanica do Ventr\u00edculo Direito 9696.6.3.2. Miocardiopatia N\u00e3o Compactada 9696.7. Valvopatias 9706.7.1. Fase Pr\u00e9-interven\u00e7\u00e3o 9706.7.2. Fase P\u00f3s-interven\u00e7\u00e3o 9706.7.3. Avalia\u00e7\u00e3o Pr\u00e9-participa\u00e7\u00e3o 9706.7.4. Particularidades na Prescri\u00e7\u00e3o e no Acompanhamento dos Exerc\u00edcios F\u00edsicos 9716.8. Portadores de Marcapasso ou Cardioversor Desfibrilador Implant\u00e1vel 9716.8.1. Benef\u00edcios Terap\u00eauticos dos Exerc\u00edcios F\u00edsicos 9726.8.2. Quando Indicar Reabilita\u00e7\u00e3o Cardiovascular 9736.8.3. Avalia\u00e7\u00e3o Pr\u00e9-participa\u00e7\u00e3o 9736.8.4. Particularidades na Prescri\u00e7\u00e3o e no Acompanhamento dos Exerc\u00edcios F\u00edsicos 9736.8.5. Treinamento Resistido 9746.8.6. Estimula\u00e7\u00e3o El\u00e9trica Neuromuscular 9746.9. Doen\u00e7a Arterial Obstrutiva Perif\u00e9rica 975Refer\u00eancias 9771 bem como da taxa de hospitaliza\u00e7\u00e3o, 2 com expressivo ganho de qualidade de vida, 2 justificando a sua consensual e enf\u00e1tica recomenda\u00e7\u00e3o pelas principais sociedades m\u00e9dicas mundiais. 6Est\u00e1 cientificamente comprovado, sendo algo incorporado ao senso comum, que ser fisicamente ativo contribui para preservar e recuperar a boa sa\u00fade do corpo e da mente. Os efeitos favor\u00e1veis da reabilita\u00e7\u00e3o cardiovascular (RCV) com \u00eanfase nos exerc\u00edcios f\u00edsicos t\u00eam sido consistentemente documentados, inclusive em meta-an\u00e1lises de estudos cl\u00ednicos randomizados, que demonstram significativas redu\u00e7\u00f5es da morbimortalidade cardiovascular e global, 8 Em contrapartida, maiores volumes de atividade f\u00edsica s\u00e3o positivamente associados \u00e0 melhor qualidade e \u00e0 maior expectativa de vida, 13 existindo uma forte e inversa associa\u00e7\u00e3o dos diferentes componentes da aptid\u00e3o f\u00edsica com a mortalidade por todas as causas e com a ocorr\u00eancia de eventos cardiovasculares desfavor\u00e1veis. Ou seja, quanto menor o n\u00edvel de aptid\u00e3o f\u00edsica, maior tende ser a taxa de mortalidade. 21O sedentarismo, que apresenta elevada preval\u00eancia no Brasil e no mundo, est\u00e1 fortemente relacionado \u00e0s doen\u00e7as cardiovasculares (DCV) e \u00e0 mortalidade precoce. 21Portanto, o principal objetivo da RCV com \u00eanfase nos exerc\u00edcios f\u00edsicos \u00e9 propiciar uma melhora dos componentes da aptid\u00e3o f\u00edsica, tanto aer\u00f3bico quanto n\u00e3o aer\u00f3bicos , algo que exige a combina\u00e7\u00e3o de diferentes modalidades de treinamento. Assim, a RCV deve proporcionar os mais elevados n\u00edveis de aptid\u00e3o f\u00edsica pass\u00edveis de obten\u00e7\u00e3o, de modo a reduzir o risco de eventos cardiovasculares e promover todos os outros benef\u00edcios a serem auferidos pela pr\u00e1tica regular de exerc\u00edcios f\u00edsicos, culminando com a redu\u00e7\u00e3o da mortalidade geral. 23 a RCV \u00e9 mundialmente subutilizada. No Brasil, pa\u00eds de dimens\u00e3o continental e grande diversidade social e econ\u00f4mica, dentre as in\u00fameras barreiras ao acesso \u00e0 RCV, 25 vale destacar como algo presente em praticamente todas as regi\u00f5es: escassez de servi\u00e7os estruturados, dificuldade de deslocamento (mobilidade urbana ruim) e n\u00edveis altos de viol\u00eancia nas cidades. 27 Neste contexto, programas de reabilita\u00e7\u00e3o cardiovascular domiciliar (RCVD), em que a maioria das sess\u00f5es ocorre no ambiente domiciliar sob supervis\u00e3o indireta, surgem como complemento ou alternativa aos programas tradicionais, nos quais as sess\u00f5es s\u00e3o sempre realizadas sob supervis\u00e3o direta ou presencial.Entretanto, apesar dos benef\u00edcios documentados e do excelente significado em termos de custo-efetividade, 31 esta diretriz aborda exclusivamente a interven\u00e7\u00e3o com base na pr\u00e1tica de exerc\u00edcios f\u00edsicos direcionadas aos pacientes com DCV, sendo a classe (ou grau) de recomenda\u00e7\u00e3o sempre fundamentada no n\u00edvel de evid\u00eancia encontrado, conforme consta a seguir.A exemplo do que ocorreu nos documentos anteriormente publicados pela Sociedade Brasileira de Cardiologia sobre o tema, Classe I: condi\u00e7\u00f5es para as quais h\u00e1 evid\u00eancias conclusivas, ou, na sua falta, consenso de que o procedimento \u00e9 seguro e \u00fatil/eficaz;Classe II: condi\u00e7\u00f5es para as quais h\u00e1 evid\u00eancias conflitantes e/ou diverg\u00eancia de opini\u00e3o sobre seguran\u00e7a e utilidade/efic\u00e1cia do procedimento:Classe IIA: peso ou evid\u00eancia/opini\u00e3o a favor do procedimento. A maioria aprova;Classe IIB: seguran\u00e7a e utilidade/efic\u00e1cia menos bem estabelecida, n\u00e3o havendo predom\u00ednio de opini\u00f5es a favor.Classe III: condi\u00e7\u00f5es para as quais h\u00e1 evid\u00eancias e/ou consenso de que o procedimento n\u00e3o \u00e9 \u00fatil/eficaz e, em alguns casos, pode ser prejudicial.N\u00edvel A: dados obtidos a partir de m\u00faltiplos estudos randomizados de bom porte, concordantes e/ou de meta-an\u00e1lise robusta de estudos cl\u00ednicos randomizados;N\u00edvel B: dados obtidos a partir de meta-an\u00e1lise menos robusta, com base em um \u00fanico estudo randomizado ou em estudos n\u00e3o randomizados (observacionais);N\u00edvel C: dados obtidos de opini\u00f5es consensuais de especialistas.32A composi\u00e7\u00e3o das equipes profissionais de RCV deve ajustar-se aos objetivos, \u00e0 clientela e \u00e0s disponibilidades de recursos humanos e materiais, respeitadas as caracter\u00edsticas regionais, a modalidade (supervis\u00e3o direta ou indireta) e o local de realiza\u00e7\u00e3o . A equipe multiprofissional habitualmente \u00e9 composta por m\u00e9dicos, educadores f\u00edsicos, fisioterapeutas e profissionais de enfermagem, mas outros, como nutricionistas, psic\u00f3logos e assistentes sociais, podem compor a equipe. 31A RCV comp\u00f5e o tratamento cl\u00ednico pleno dos pacientes est\u00e1veis com DCV, o que exige a integra\u00e7\u00e3o do m\u00e9dico assistente, que, ao encaminhar o seu paciente, deve ter conhecimento das indica\u00e7\u00f5es e dos benef\u00edcios a serem obtidos, adotando as necess\u00e1rias provid\u00eancias cl\u00ednicas pr\u00e9-participa\u00e7\u00e3o. Tendo em vista o encaminhamento de relat\u00f3rios, eventuais necessidades de ajustes farmacol\u00f3gicos, intercorr\u00eancias m\u00e9dicas, entre outros, \u00e9 de grande relev\u00e2ncia que sejam criados mecanismos para uma f\u00e1cil comunica\u00e7\u00e3o entre o m\u00e9dico assistente e a equipe de RCV. 34Coordena as a\u00e7\u00f5es m\u00e9dicas, sendo no Brasil habitualmente o coordenador geral do programa de RCV. Ele deve conhecer em profundidade a tem\u00e1tica de RCV e ter conhecimento para atuar em emerg\u00eancias cardiovasculares. Algumas de suas principais atua\u00e7\u00f5es s\u00e3o:31Executar a avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o, com inclus\u00e3o de testes de exerc\u00edcio, de modo a subsidiar a programa\u00e7\u00e3o inicial das sess\u00f5es de treinamento da RCV; Treinar a equipe para identificar situa\u00e7\u00f5es de risco e realizar o atendimento apropriado em situa\u00e7\u00f5es emergenciais;Estabelecer restri\u00e7\u00f5es e limites para a prescri\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos;Liderar e interagir com os demais membros da equipe, com o objetivo de otimizar a qualidade e a seguran\u00e7a da prescri\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos;Programar reavalia\u00e7\u00f5es subsequentes, sempre interagindo com o m\u00e9dico assistente.31De modo semelhante aos m\u00e9dicos, os demais membros da equipe, ao executarem suas respectivas fun\u00e7\u00f5es, devem seguir as normas e regras que norteiam as atividades do programa, respeitando as recomenda\u00e7\u00f5es de seus respectivos conselhos profissionais. Atuam diretamente na prescri\u00e7\u00e3o e na supervis\u00e3o dos exerc\u00edcios f\u00edsicos, dentro das metas e dos limites definidos na orienta\u00e7\u00e3o m\u00e9dica, ap\u00f3s a avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o e subsequentes reavalia\u00e7\u00f5es. Devem ter conhecimentos espec\u00edficos sobre as DCV e fisiologia do exerc\u00edcio, al\u00e9m de receberem periodicamente treinamento de suporte b\u00e1sico de vida, incluindo o uso de desfibrilador autom\u00e1tico externo. Al\u00e9m da atua\u00e7\u00e3o nas sess\u00f5es de exerc\u00edcios f\u00edsicos, podem contribuir para as orienta\u00e7\u00f5es e demais medidas, visando a ado\u00e7\u00e3o de h\u00e1bitos saud\u00e1veis.Em um programa de RCV, o profissional de enfermagem pode auxiliar na avalia\u00e7\u00e3o cl\u00ednica, atuando na obten\u00e7\u00e3o e no fornecimento de informa\u00e7\u00f5es relacionadas \u00e0 situa\u00e7\u00e3o cl\u00ednica do paciente, inclusive em contato com os familiares. Pode ser respons\u00e1vel pelas dosagens de glicemia e verifica\u00e7\u00e3o de press\u00e3o arterial (PA), antes e durante as sess\u00f5es de exerc\u00edcios. Em caso de intercorr\u00eancias cl\u00ednicas, pode participar do atendimento e auxiliar o m\u00e9dico, com eventuais administra\u00e7\u00f5es de medicamentos. Deve tamb\u00e9m estar capacitado para atuar no suporte b\u00e1sico de vida, com uso de desfibrilador autom\u00e1tico externo.29Um programa de RCV pode funcionar em v\u00e1rios tipos de instala\u00e7\u00f5es, a depender dos objetivos e recursos dispon\u00edveis. Mais frequentemente, os programas de RCV s\u00e3o realizados em ambientes fechados e climatizados, sendo tamb\u00e9m poss\u00edvel realizar as sess\u00f5es de exerc\u00edcios f\u00edsicos em espa\u00e7os abertos, como pistas de atletismo, quadras, gin\u00e1sios poliesportivos, parques ou \u00e1reas p\u00fablicas de lazer. o C e 25 o C, al\u00e9m de umidade relativa do ar entre 40 e 65% durante as sess\u00f5es de exerc\u00edcio. A \u00e1rea dispon\u00edvel exclusivamente para a realiza\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos, desconsiderando vesti\u00e1rios, banheiros, recep\u00e7\u00e3o ou sala de espera, varia muito, podendo ir desde 20 m 2 a algumas centenas de metros quadrados. \u00c9 importante que existam locais pr\u00f3prios para a troca de roupas e instala\u00e7\u00f5es sanit\u00e1rias. Para minimizar o risco de acidentes e quedas, o piso deve ter propriedades antiderrapantes. 29Em ambientes fechados, o espa\u00e7o para a realiza\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos dever\u00e1 apresentar dimens\u00f5es e caracter\u00edsticas adequadas, vari\u00e1veis de acordo com os recursos locais e a capacidade de atendimento. O ambiente dever\u00e1 ser suficientemente amplo para a realiza\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos, com uma altura de p\u00e9 direito idealmente igual ou superior a 2,5 m. Tamb\u00e9m dever\u00e1 ser apropriadamente iluminado e bem ventilado, onde seja poss\u00edvel manter a temperatura entre 22 2.2.2.1. Exerc\u00edcios Aer\u00f3bicos29Os equipamentos mais usados s\u00e3o esteiras rolantes e cicloerg\u00f4metros de membros inferiores (MMII), mas tamb\u00e9m podem ser utilizados cicloerg\u00f4metros de membros superiores (MMSS), remoerg\u00f4metros, erg\u00f4metros de esqui, el\u00edpticos, entre outros. As esteiras rolantes devem ser el\u00e9tricas, com capacidade de suportar, pelo menos, 100 kg de peso corporal, com suportes frontal e lateral para as m\u00e3os e trava de seguran\u00e7a. Devem tamb\u00e9m permitir ajuste individualizado dentro de uma faixa ampla de velocidade e inclina\u00e7\u00e3o. Os cicloerg\u00f4metros podem ser de frenagem mec\u00e2nica ou eletromagn\u00e9tica. H\u00e1 modelos espec\u00edficos para MMSS ou ainda para que os quatro membros sejam exercitados simultaneamente. Para os modelos de MMII existem as op\u00e7\u00f5es vertical e horizontal. O ideal \u00e9 que o cicloerg\u00f4metro possibilite a leitura da cad\u00eancia ou velocidade e, principalmente, da pot\u00eancia em watts. H\u00e1 cicloerg\u00f4metros em que \u00e9 poss\u00edvel programar a intensidade diretamente em watts, de modo que a resist\u00eancia do pedal aumenta quando a cad\u00eancia diminui e vice-versa.Os remoerg\u00f4metros, erg\u00f4metros de esqui e el\u00edpticos podem ser particularmente \u00fateis para os pacientes com menor grau de limita\u00e7\u00e3o funcional ou que j\u00e1 tenham tido experi\u00eancias pr\u00e9vias com tais equipamentos. Eles apresentam como vantagem possibilitar o exerc\u00edcio simult\u00e2neo dos MMSS e MMII.2.2.2.2. Exerc\u00edcios de Fortalecimento MuscularH\u00e1 v\u00e1rios tipos de equipamentos que podem ser utilizados para o fortalecimento muscular. Por\u00e9m, \u00e9 poss\u00edvel realizar v\u00e1rios exerc\u00edcios utilizando somente o peso corporal, que representa um esfor\u00e7o, em geral, suficiente nos pacientes mais debilitados. Um exemplo pr\u00e1tico \u00e9 o exerc\u00edcio de sentar e levantar, cuja realiza\u00e7\u00e3o requer t\u00e3o somente uma cadeira ou um banco.medicine balls ), \u201cbolas su\u00ed\u00e7as\u201d e faixas ou bandas el\u00e1sticas com diferentes graus de resist\u00eancia. 29O uso de cordas ou faixas suspensas, bem fixadas ao teto ou alto da parede, podem permitir uma ampla variedade de exerc\u00edcios com a utiliza\u00e7\u00e3o do peso do pr\u00f3prio corpo. Pesos livres, halteres ou caneleiras com pesos variados s\u00e3o frequentemente adotados em programas de RCV e possibilitam uma ampla variedade de movimentos e est\u00edmulos adequados de diferentes grupos musculares. Podem ser tamb\u00e9m utilizados aparelhos espec\u00edficos, com pesos ligados a cabos e polias. Outros equipamentos que tamb\u00e9m podem ser usados: barras, bast\u00f5es, bolas com peso , glicos\u00edmetros e ox\u00edmetros digitais. Dependendo da complexidade cl\u00ednica e do risco de eventos cardiovasculares desfavor\u00e1veis, \u00e9 desej\u00e1vel o monitoramento eletrocardiogr\u00e1fico no repouso e durante o exerc\u00edcio, que pode ser obtido por equipamentos de conex\u00e3o direta ao paciente ou por sistemas de telemetria, sendo de fundamental import\u00e2ncia em caso de eventos cardiovasculares a possibilidade do r\u00e1pido acesso aos equipamentos, para identifica\u00e7\u00e3o do quadro cl\u00ednico e a subsequente conduta m\u00e9dica.Apesar de ser extremamente incomum, \u00e9 importante que o programa tenha um planejamento para o adequado atendimento de eventos cardiovasculares graves, como a parada cardiorrespirat\u00f3ria, que, na maioria dos casos em adultos, decorre de fibrila\u00e7\u00e3o ventricular ou taquicardia ventricular sem pulso. Portanto, o desfibrilador, manual ou autom\u00e1tico, \u00e9 um equipamento de seguran\u00e7a obrigat\u00f3rio. Ainda devem estar dispon\u00edveis outros materiais do suporte b\u00e1sico e avan\u00e7ado de vida, como laringosc\u00f3pio, tubos orotraqueais de tamanhos variados, m\u00e1scaras, ambu e oxigenioterapia suplementar.36Para orienta\u00e7\u00e3o mais detalhada de t\u00e9cnicas, equipamentos e medicamentos, orienta-se consultar diretrizes espec\u00edficas sobre os respectivos assuntos. 31Tradicionalmente, a RCV \u00e9 dividida em fases temporais, sendo a fase 1 intra-hospitalar e as fases 2 a 4 ambulatoriais. Nos prim\u00f3rdios, a fase 1 foi destinada \u00e0 recupera\u00e7\u00e3o ap\u00f3s infarto agudo do mioc\u00e1rdio (IAM) ou cirurgia de revasculariza\u00e7\u00e3o mioc\u00e1rdica (CRVM). Posteriormente, em contexto atualmente denominado reabilita\u00e7\u00e3o cardiopulmonar e metab\u00f3lica, foram inclu\u00eddos os pacientes internados submetidos a interven\u00e7\u00f5es coron\u00e1rias percut\u00e2neas (ICP), cirurgias valvares, cirurgias para cardiopatias cong\u00eanitas e transplante card\u00edaco (TxC), al\u00e9m dos portadores de insufici\u00eancia card\u00edaca (IC), doen\u00e7a arterial coronariana (DAC), diab\u00e9ticos, hipertensos, pneumopatas e nefropatas cr\u00f4nicos, assim que estabilizados clinicamente. Portanto, a RCV deve ser iniciada imediatamente ap\u00f3s o paciente ter sido considerado clinicamente compensado, como decorr\u00eancia do tratamento cl\u00ednico e/ou intervencionista. 31Na fase 1 da RCV objetiva-se que o paciente tenha alta hospitalar com as melhores condi\u00e7\u00f5es f\u00edsicas e psicol\u00f3gicas poss\u00edveis, municiado de informa\u00e7\u00f5es referentes ao estilo saud\u00e1vel de vida, em especial no que diz respeito ao exerc\u00edcio f\u00edsico. Prop\u00f5e-se a combina\u00e7\u00e3o de exerc\u00edcios f\u00edsicos de baixa intensidade, t\u00e9cnicas para o controle do estresse e programas de educa\u00e7\u00e3o em rela\u00e7\u00e3o aos fatores de risco e \u00e0 cardiopatia. A equipe de atendimento deve ser composta por, pelo menos, m\u00e9dico, fisioterapeuta e enfermeiro, capacitados para atuar em RCV, que n\u00e3o precisam dedicar tempo integral ao programa de reabilita\u00e7\u00e3o, podendo exercer outras atividades no hospital. O direcionamento \u00e0s fases ambulatoriais da RCV deve ser realizado na alta da interna\u00e7\u00e3o. A fase 2 come\u00e7a imediatamente ap\u00f3s a alta hospitalar e tem dura\u00e7\u00e3o m\u00e9dia de 3 meses. A fase 3 costuma ter dura\u00e7\u00e3o de 3 a 6 meses e a fase 4 tem dura\u00e7\u00e3o prolongada. Em todas as fases objetiva-se progress\u00e3o dos benef\u00edcios da RCV ou, pelo menos, a manuten\u00e7\u00e3o dos ganhos obtidos.31Em uma divis\u00e3o r\u00edgida da RCV em fases temporais, pode-se n\u00e3o levar em considera\u00e7\u00e3o que existem pacientes com cardiopatias graves, muito sintom\u00e1ticos e debilitados, que permanecem por longo prazo em uma reabilita\u00e7\u00e3o \u201cfase 2\u201d, pois continuam requerendo a supervis\u00e3o direta dos exerc\u00edcios f\u00edsicos, enquanto outros, de baixo risco, desde o in\u00edcio se enquadram em programas de fase 3 ou mesmo de fase 4, sendo potenciais candidatos a uma RCV domiciliar, em que a maioria das sess\u00f5es ocorrem sob supervis\u00e3o indireta, \u00e0 dist\u00e2ncia. Portanto, recomenda-se uma estratifica\u00e7\u00e3o do risco cl\u00ednico que possibilite o uso mais racional dos programas, com direcionamento individualizado \u00e0s modalidades de RCV. Nesse contexto, os pacientes de alto risco, com menor capacidade f\u00edsica e mais sintom\u00e1ticos, devem participar de sess\u00f5es supervisionadas por tempo indeterminado, enquanto os de menor risco, com maior capacidade f\u00edsica e menos sintom\u00e1ticos precocemente podem realizar, sem supervis\u00e3o direta, exerc\u00edcios mais intensos e diversificados .37 As notas de corte para o enquadramento s\u00e3o baseadas na opini\u00e3o de especialistas (evid\u00eancia n\u00edvel C), o que possibilita modifica\u00e7\u00f5es regionais de acordo com a experi\u00eancia da equipe da RCV e com o julgamento cl\u00ednico realizado na avalia\u00e7\u00e3o m\u00e9dica pr\u00e9-participa\u00e7\u00e3o e subsequentes reavalia\u00e7\u00f5es de repouso e teste cardiopulmonar de exerc\u00edcio (TCPE) ou teste ergom\u00e9trico (TE).Os pacientes de alto risco, com frequ\u00eancia, podem necessitar de atendimento m\u00e9dico imediato ou a curto prazo . Portanto, requerem maior monitoramento do treinamento pela equipe assistencial, a qual deve ser capaz de identificar sinais e sintomas de situa\u00e7\u00f5es de risco e atuar no atendimento de intercorr\u00eancias cl\u00ednicas, inclusive com material de suporte b\u00e1sico e avan\u00e7ado de vida, com cardiodesfibrilador manual ou autom\u00e1tico. \u00c9 preferencial, inclusive, que esse equipamento esteja dentro da sala de atendimento. A equipe m\u00e9dica deve estar prontamente dispon\u00edvel na localidade, com r\u00e1pido acesso ao paciente em caso de intercorr\u00eancias graves.Ressalte-se que a melhor maneira de prevenir intercorr\u00eancias durante um programa de reabilita\u00e7\u00e3o e, especialmente ap\u00f3s eventos e interven\u00e7\u00f5es, consiste na realiza\u00e7\u00e3o de qualificadas avalia\u00e7\u00f5es pr\u00e9-participa\u00e7\u00e3o e subsequentes, que devem ser sistem\u00e1ticas.O programa de exerc\u00edcios deve ser individualizado em termos de intensidade, dura\u00e7\u00e3o, frequ\u00eancia, modalidade de treinamento e progress\u00e3o, de acordo com os testes funcionais realizados inicialmente e no seguimento. Sempre devem ser adotados recursos para a correta determina\u00e7\u00e3o da FC e verifica\u00e7\u00e3o da PA, em repouso e em esfor\u00e7o, al\u00e9m da possibilidade de verifica\u00e7\u00e3o de satura\u00e7\u00e3o de oxig\u00eanio, determina\u00e7\u00e3o da glicemia capilar e monitoramento eletrocardiogr\u00e1fico.O atendimento tamb\u00e9m deve contemplar um programa educacional direcionado \u00e0 modifica\u00e7\u00e3o do estilo de vida, com \u00eanfase na reeduca\u00e7\u00e3o alimentar e em estrat\u00e9gias para cessa\u00e7\u00e3o do tabagismo, quando necess\u00e1rias. \u00c9 importante que o paciente obtenha conhecimentos sobre sua doen\u00e7a e aprendizado de automonitoramento, tanto na execu\u00e7\u00e3o dos exerc\u00edcios quanto na identifica\u00e7\u00e3o de sinais e sintomas de alerta para situa\u00e7\u00f5es cl\u00ednicas inst\u00e1veis ou de risco.As caracter\u00edsticas cl\u00ednicas dos pacientes que se enquadrariam inicialmente no risco cl\u00ednico alto s\u00e3o:Interna\u00e7\u00e3o por descompensa\u00e7\u00e3o cardiovascular recente (menos de 8 a 12 semanas) devido a quadros de: IAM ou angina inst\u00e1vel; revasculariza\u00e7\u00e3o cir\u00fargica ou percut\u00e2nea; arritmias complexas; morte s\u00fabita revertida; descompensa\u00e7\u00e3o de IC;Pacientes cardiopatas, com presen\u00e7a ou aus\u00eancia de evento cardiovascular e/ou interven\u00e7\u00f5es, mas com importantes altera\u00e7\u00f5es funcionais ao esfor\u00e7o f\u00edsico, ou seja:\u2013 Baixa capacidade funcional no TE ou no TCPE (classifica\u00e7\u00e3o de Weber C e D ou consumo de oxig\u00eanio [VO 2 ] abaixo de 60% do predito para idade e sexo);\u2013 Sinais e sintomas de isquemia mioc\u00e1rdica em baixa carga (abaixo de 6 MET ou de VO 2 de 15 ml.kg -1 .min -1 );\u2013 Sintomatologia exacerbada .Outras caracter\u00edsticas cl\u00ednicas de pacientes com risco aumentado aos exerc\u00edcios f\u00edsicos: doen\u00e7a renal cr\u00f4nica (DRC) dial\u00edtica, dessatura\u00e7\u00e3o de oxig\u00eanio em esfor\u00e7o e arritmia ventricular complexa em repouso ou esfor\u00e7o.Considerando que os pacientes de alto risco frequentemente necessitam de reajustes de f\u00e1rmacos e de reavalia\u00e7\u00f5es, com eventuais interven\u00e7\u00f5es (revasculariza\u00e7\u00f5es ou outros procedimentos), torna-se essencial comunica\u00e7\u00e3o constante da equipe assistencial da RCV com o(s) m\u00e9dico(s) assistente(s). \u00c9 importante destacar que alguns pacientes, devido a intercorr\u00eancias nas sess\u00f5es e/ou resultados nas avalia\u00e7\u00f5es subsequentes, podem permanecer classificados como de alto risco, mantendo a pr\u00e1tica de exerc\u00edcios f\u00edsicos sob supervis\u00e3o direta por tempo indeterminado.Os pacientes podem ter cumprido etapas anteriores da RCV, sendo reclassificados, ingressar diretamente nessa categoria sem participa\u00e7\u00f5es pr\u00e9vias ou ser oriundos de outros programas de exerc\u00edcios. A dura\u00e7\u00e3o da RCV sob essa classifica\u00e7\u00e3o tamb\u00e9m pode ser vari\u00e1vel, a depender do quadro cl\u00ednico e da evolu\u00e7\u00e3o do treinamento f\u00edsico, algo a ser definido nas reavalia\u00e7\u00f5es subsequentes.A supervis\u00e3o de exerc\u00edcios deve ser feita pelo fisioterapeuta ou professor de educa\u00e7\u00e3o f\u00edsica, e o servi\u00e7o deve, idealmente, contar com a coordena\u00e7\u00e3o geral de um m\u00e9dico com experi\u00eancia em RCV. \u00c9 recomendada a disponibilidade de recursos para a correta determina\u00e7\u00e3o da FC e verifica\u00e7\u00e3o de PA em repouso e esfor\u00e7o e, sempre que necess\u00e1rio, com possibilidade de verifica\u00e7\u00e3o da satura\u00e7\u00e3o de oxig\u00eanio, determina\u00e7\u00e3o da glicemia e monitoramento eletrocardiogr\u00e1fico.Para o atendimento de pacientes de risco intermedi\u00e1rio, caso n\u00e3o haja m\u00e9dico presente no local das atividades, deve haver possibilidade do seu r\u00e1pido acionamento remoto. A estrutura do servi\u00e7o deve apresentar material de suporte b\u00e1sico de vida e profissionais de sa\u00fade treinados em reanima\u00e7\u00e3o cardiopulmonar, com o uso de desfibrilador autom\u00e1tico externo, o qual deve estar presente no local de atendimento.\u00c9 fundamental que a equipe m\u00e9dica integrada ao servi\u00e7o de RCV realize a avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o, com adequada estratifica\u00e7\u00e3o do risco. O seguimento m\u00e9dico regular e as reavalia\u00e7\u00f5es sistem\u00e1ticas, al\u00e9m dos atendimentos eventuais quando necess\u00e1rios, s\u00e3o fundamentais para garantir a seguran\u00e7a dos exerc\u00edcios.As caracter\u00edsticas cl\u00ednicas dos pacientes com risco intermedi\u00e1rio s\u00e3o:Evento cardiovascular ou interven\u00e7\u00f5es com intervalo superior a 12 semanas, com estabilidade do quadro cl\u00ednico;Pacientes cardiopatas que ainda apresentam algumas altera\u00e7\u00f5es funcionais em esfor\u00e7o f\u00edsico:\u2013 Moderada capacidade funcional no TE (entre 5 e 7 MET) ou no TCPE (classifica\u00e7\u00e3o de Weber B ou VO 2 entre 60 e 85% do predito para idade e sexo);\u2013 Sinais e sintomas de isquemia em carga acima de 6 MET ou com VO 2 acima de 15 ml.kg -1 .min -1 ;\u2013 Sintomatologia de menor magnitude .Outras caracter\u00edsticas cl\u00ednicas que o m\u00e9dico respons\u00e1vel pela avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o julgue como de risco intermedi\u00e1rio aos exerc\u00edcios f\u00edsicos.O principal objetivo da RCV neste perfil de risco ainda \u00e9 o aprimoramento da aptid\u00e3o f\u00edsica, tanto aer\u00f3bica quanto n\u00e3o aer\u00f3bica , com melhor controle da(s) doen\u00e7a(s). Deve ser considerada a necessidade de promo\u00e7\u00e3o de bem-estar, com melhora da qualidade de vida, al\u00e9m de outros procedimentos que contribuam para a redu\u00e7\u00e3o do risco de complica\u00e7\u00f5es cl\u00ednicas, como \u00e9 o caso das estrat\u00e9gias para cessa\u00e7\u00e3o do tabagismo, reeduca\u00e7\u00e3o alimentar e controle de peso corporal. A \u00eanfase na manuten\u00e7\u00e3o e ades\u00e3o do tratamento farmacol\u00f3gico tamb\u00e9m \u00e9 fundamental para evitar a progress\u00e3o ou instabiliza\u00e7\u00e3o da DCV. A obten\u00e7\u00e3o de conhecimentos sobre a pr\u00f3pria doen\u00e7a, possibilitando melhor automonitoramento, aumentam a acur\u00e1cia na identifica\u00e7\u00e3o de sinais e sintomas relacionados \u00e0 progress\u00e3o da doen\u00e7a ou a situa\u00e7\u00f5es cl\u00ednicas inst\u00e1veis, que podem requerer interrup\u00e7\u00e3o do programa de exerc\u00edcios e reavalia\u00e7\u00f5es m\u00e9dicas.Os pacientes desta categoria, ap\u00f3s um per\u00edodo inicial de orienta\u00e7\u00f5es e de obten\u00e7\u00e3o de conhecimentos sobre os exerc\u00edcios e o automonitoramento, podem adequar-se a uma RCV domiciliar, na qual a pr\u00e1tica de exerc\u00edcio f\u00edsico \u00e9 realizada com supervis\u00e3o indireta, sob a responsabilidade de profissionais do servi\u00e7o. A avalia\u00e7\u00e3o das sess\u00f5es, com reajustes na prescri\u00e7\u00e3o e esclarecimentos de d\u00favidas, deve ser feita de maneira sistem\u00e1tica, presencial ou virtualmente, conforme o caso.Assim como os pacientes de risco intermedi\u00e1rio, os de baixo risco podem ter sido reclassificados ap\u00f3s cumprirem etapas anteriores da RCV, ingressarem diretamente nesta categoria sem participa\u00e7\u00f5es pr\u00e9vias na RCV, ou serem oriundos de outros programas de exerc\u00edcios f\u00edsicos. A dura\u00e7\u00e3o do treinamento destes pacientes \u00e9 de longo prazo, visando a manuten\u00e7\u00e3o da sa\u00fade geral e obten\u00e7\u00e3o dos maiores ganhos poss\u00edveis nos componentes da aptid\u00e3o f\u00edsica, com o objetivo de alcan\u00e7ar ao m\u00e1ximo o potencial de sa\u00fade do indiv\u00edduo.Dependendo da disponibilidade e das prefer\u00eancias individuais, os exerc\u00edcios podem ser realizados sob supervis\u00e3o presencial ou \u00e0 dist\u00e2ncia. Por\u00e9m, em virtude do menor risco cl\u00ednico e da menor necessidade de supervis\u00e3o, os pacientes deste est\u00e1gio se enquadram perfeitamente em um modelo domiciliar, de modo que a equipe da RCV possa dedicar aten\u00e7\u00e3o assistencial presencial principalmente aos pacientes de maior risco cl\u00ednico.Os pacientes devem ter reavalia\u00e7\u00f5es m\u00e9dicas peri\u00f3dicas, realizadas pelo seu m\u00e9dico assistente e pela equipe da RCV, com TCPE ou TE, cuja periodicidade, a princ\u00edpio, n\u00e3o deve exceder 12 meses. O objetivo das reavalia\u00e7\u00f5es m\u00e9dicas \u00e9 reajustar a prescri\u00e7\u00e3o do treinamento e identificar eventual piora da doen\u00e7a ou sinais de risco para situa\u00e7\u00f5es inst\u00e1veis ou eventos cardiovasculares, possibilitando eventuais reajustes do tratamento farmacol\u00f3gico e/ou interven\u00e7\u00f5es cir\u00fargicas ou percut\u00e2neas.Os pacientes em RCVD devem ser periodicamente avaliados e orientados para a pr\u00e1tica dos exerc\u00edcios, ocasi\u00f5es em que \u00e9 recomend\u00e1vel a participa\u00e7\u00e3o em algumas sess\u00f5es supervisionadas de exerc\u00edcios, especialmente para os menos experientes, possibilitando eventuais reajustes na prescri\u00e7\u00e3o e esclarecimento de d\u00favidas. \u00c9 tamb\u00e9m recomend\u00e1vel a realiza\u00e7\u00e3o de consultas peri\u00f3dicas com a equipe da RCV, por meio de contatos virtuais e/ou telef\u00f4nicos, pelo menos uma vez a cada seis meses, para estimular a ades\u00e3o ao programa de exerc\u00edcios f\u00edsicos.As caracter\u00edsticas cl\u00ednicas dos pacientes do est\u00e1gio 4 (presen\u00e7a de todas as caracter\u00edsticas a seguir) s\u00e3o:Evento cardiovascular ou interven\u00e7\u00f5es com intervalo superior a 6 meses e estabilidade cl\u00ednica;Pacientes cardiopatas que n\u00e3o apresentam altera\u00e7\u00f5es funcionais em esfor\u00e7o f\u00edsico ou que estas sejam muito discretas quando presentes;Os pacientes nessa classifica\u00e7\u00e3o costumam apresentar as seguintes caracter\u00edsticas:\u2013 Boa capacidade funcional no TE (superior a 7 MET) ou no TCPE (classifica\u00e7\u00e3o de Weber A ou VO 2 acima de 85% do predito para idade e sexo);\u2013 Aus\u00eancia de sinais e sintomas de isquemia mioc\u00e1rdica ou de outra sintomatologia anormal ao esfor\u00e7o f\u00edsico.38 Em rela\u00e7\u00e3o \u00e0s DCV, 863 bilh\u00f5es de d\u00f3lares foram gastos mundialmente em 2010, estimando-se que em 2030 chegue a 1,04 trilh\u00e3o de d\u00f3lares. 39Segundo a Organiza\u00e7\u00e3o Mundial da Sa\u00fade, entre 2000 e 2016 o aumento mundial dos gastos com sa\u00fade no mundo foi maior do que o crescimento da economia global, chegando a 7,5 trilh\u00f5es de d\u00f3lares em 2016. 40 observa-se situa\u00e7\u00e3o semelhante, pois as DCV constituem o grupo que ocasiona o maior gasto com interna\u00e7\u00f5es no Sistema \u00danico de Sa\u00fade, sendo a principal causa de aposentadorias por invalidez. 45 Em 2015, em rela\u00e7\u00e3o \u00e0s DCV, estima-se que o gasto p\u00fablico com interna\u00e7\u00f5es hospitalares e consultas tenha sido superior a 5 bilh\u00f5es de reais e o gasto por afastamentos tempor\u00e1rios ou permanentes superior a 380 milh\u00f5es de reais. 40No Brasil, onde quase 50% dos gastos com sa\u00fade s\u00e3o financiados pelo governo, 47 Al\u00e9m disso, sua utiliza\u00e7\u00e3o em maior escala proporcionaria redu\u00e7\u00e3o nos gastos com sa\u00fade, em decorr\u00eancia da diminui\u00e7\u00e3o de novos eventos cardiovasculares, reinterna\u00e7\u00f5es hospitalares e tratamentos intervencionistas. 49 Assim, sua dissemina\u00e7\u00e3o deveria ser considerada uma estrat\u00e9gia de sa\u00fade p\u00fablica priorit\u00e1ria.Portanto, o impacto econ\u00f4mico provocado pelas DCV, aliado \u00e0 obrigatoriedade do uso consequente e racional de recursos financeiros, exige a implementa\u00e7\u00e3o em larga escala de modelos de baixo custo, viabilizando o atendimento de maior n\u00famero de pacientes. Em coronariopatas est\u00e1veis, a RCV \u00e9 uma estrat\u00e9gia que, em termos de custo-efetividade, supera, com larga margem, procedimentos amplamente utilizados no pa\u00eds, tais como a interven\u00e7\u00e3o coronariana percut\u00e2nea (ICP). 52 De acordo com Georgiou et al., 53 s\u00e3o consideradas medidas de excelente custo-efetividade as que exigem investimentos inferiores a 20.000 d\u00f3lares para salvar uma vida por ano (VSA), sendo aceit\u00e1veis as que exigem investimentos entre 20.000 e 40.000 d\u00f3lares e inaceit\u00e1veis aquelas que exigem investimentos acima de 40.000 d\u00f3lares por VSA.A determina\u00e7\u00e3o da custo-efetividade, que se faz por an\u00e1lise combinada das consequ\u00eancias cl\u00ednicas (efetividade) e do gasto financeiro do sistema de sa\u00fade, \u00e9 fundamental para avaliar a pertin\u00eancia da implementa\u00e7\u00e3o em larga escala de determinado tratamento. 23 publicaram a primeira revis\u00e3o sistem\u00e1tica de estudos sobre custo-efetividade da RCV como preven\u00e7\u00e3o secund\u00e1ria em pacientes portadores de DAC e IC. 23 Em artigo de 2018, 54 a avalia\u00e7\u00e3o de estudos publicados ap\u00f3s 2001 mostrou uma rela\u00e7\u00e3o de custo-efetividade muito semelhante \u00e0 descrita anteriormente, sendo o aumento dos gastos com a adi\u00e7\u00e3o da RCV ao tratamento convencional situado entre 2.555 e 23.598 d\u00f3lares por VSA.De acordo com os dados dispon\u00edveis entre 1985 e 2004, a RCV foi considerada uma interven\u00e7\u00e3o com excelente rela\u00e7\u00e3o de custo-efetividade, na medida em que a sua adi\u00e7\u00e3o ao tratamento convencional resultou em um aumento de gastos de 2.193 a 28.193 d\u00f3lares por VSA. Em 2005, Papadakis et al. per capita , 55 h\u00e1 uma escassez de dados sobre custo-efetividade da RCV nesses pa\u00edses. 56 A maioria das informa\u00e7\u00f5es \u00e9 oriunda de na\u00e7\u00f5es de alta renda per capita , como Estados Unidos, Canad\u00e1 e pa\u00edses europeus, dificultando a extrapola\u00e7\u00e3o dos resultados para a realidade brasileira. Entretanto, vale ressaltar que os poucos estudos dispon\u00edveis nos pa\u00edses de m\u00e9dia e baixa renda mostram a mesma tend\u00eancia. No Brasil, a incorpora\u00e7\u00e3o da reabilita\u00e7\u00e3o ao tratamento convencional de pacientes com IC resultou em um aumento de gasto de 21.169 d\u00f3lares por VSA. 57Cabe ainda destacar que, apesar de mais de 75% das mortes por DCV ocorrerem em pa\u00edses de m\u00e9dia e baixa renda 59 pois na maioria dos estados, inclusive na maior parte das capitais e grandes cidades brasileiras, n\u00e3o existe sequer um \u00fanico servi\u00e7o de RCV.No entanto, apesar dos claros benef\u00edcios cl\u00ednicos e econ\u00f4micos da RCV, o percentual de pacientes eleg\u00edveis que efetivamente participam desse tipo de servi\u00e7o est\u00e1 muito aqu\u00e9m do desejado. Segundo dados internacionais, apenas em torno de 30% frequentam um programa de RCV e, no Brasil, estima-se que a situa\u00e7\u00e3o seja ainda pior, estando certamente muito abaixo de 15%, 62 al\u00e9m do avan\u00e7o tecnol\u00f3gico de dispositivos que permitem o monitoramento \u00e0 dist\u00e2ncia, tem sido ampliada a utiliza\u00e7\u00e3o deste tipo de servi\u00e7o para o atendimento de pacientes com perfil de risco mais elevado.Neste contexto, a utiliza\u00e7\u00e3o de modelos de RCVD tem crescido. Inicialmente, a preocupa\u00e7\u00e3o quanto \u00e0 seguran\u00e7a da pr\u00e1tica do exerc\u00edcio f\u00edsico fez com que a RCVD fosse destinada somente a pacientes de baixo risco. No entanto, com a demonstra\u00e7\u00e3o de que n\u00e3o h\u00e1 inferioridade quanto \u00e0 seguran\u00e7a e com benef\u00edcios cl\u00ednicos semelhantes em rela\u00e7\u00e3o \u00e0 estrat\u00e9gia convencional, 60 que compararam os efeitos dos dois modelos em pacientes com DAC de risco leve e moderado em interven\u00e7\u00f5es realizadas por 3 meses, ap\u00f3s evento coronariano agudo. Apesar de o grupo de pacientes que atendeu ao programa tradicional ter realizado um volume maior de exerc\u00edcios f\u00edsicos, n\u00e3o houve diferen\u00e7a quanto ao ganho em capacidade funcional ou em qualidade de vida entre os dois grupos. Jolly et al. 62 compararam os desfechos relacionados aos fatores de risco cardiovasculares entre os programas tradicional e domiciliar por um per\u00edodo mais longo, com seguimentos de 6, 12 e 24 meses, e n\u00e3o observaram diferen\u00e7as nos resultados.Estudos recentes mostram que a RCVD apresenta efetividade semelhante \u00e0 tradicional, conforme demonstraram Ades et al., 61 tamb\u00e9m n\u00e3o encontrou diferen\u00e7as significativas entre as duas propostas em rela\u00e7\u00e3o aos desfechos morte, eventos card\u00edacos, capacidade funcional, qualidade de vida e fatores de risco modific\u00e1veis, no curto prazo (3 a 12 meses) e no longo prazo (at\u00e9 24 meses).Recentemente, uma revis\u00e3o sistem\u00e1tica de estudos com pacientes ap\u00f3s IAM, CRVM ou IC, realizada por Anderson et al., 64 havendo uma grande lacuna de pesquisas que possibilitem a compara\u00e7\u00e3o das suas propostas em termos de custo-efetividade. 67Assim, programas de RCVD devem ser considerados como estrat\u00e9gia para facilitar o acesso, a ades\u00e3o e a consequente dissemina\u00e7\u00e3o da interven\u00e7\u00e3o. Entretanto, existem apenas poucos estudos demonstrando que a RCVD apresenta um custo semelhante ao dos programas tradicionais, Diante dos fatos, \u00e9 insustent\u00e1vel que pa\u00edses de todos os n\u00edveis de renda e, mais preocupantemente os de m\u00e9dia e baixa, continuem fornecendo massivamente e sem crit\u00e9rios de indica\u00e7\u00e3o mais rigorosos, interven\u00e7\u00f5es terap\u00eauticas de alto custo e persistam negligenciando em rela\u00e7\u00e3o a uma estrat\u00e9gia altamente efetiva, economicamente vi\u00e1vel e de grande aplicabilidade como a RCV. Portanto, h\u00e1 necessidade da implementa\u00e7\u00e3o de pol\u00edticas de sa\u00fade p\u00fablica, com o objetivo de aumentar a disponibilidade, a participa\u00e7\u00e3o e a ades\u00e3o dos pacientes eleg\u00edveis aos programas de RCV tradicionais e domiciliares.Por fim, considerando a relev\u00e2ncia da RCV, fundamentada em seu amplo benef\u00edcio cl\u00ednico e custo-efetividade, imp\u00f5e-se a ado\u00e7\u00e3o de estrat\u00e9gias que modifiquem a cultura m\u00e9dica e favore\u00e7am a dissemina\u00e7\u00e3o de programas estruturados. Nesse contexto, torna-se relevante que servi\u00e7os de refer\u00eancia em cardiologia ofere\u00e7am a RCV aos seus pacientes durante a interna\u00e7\u00e3o e ap\u00f3s a alta hospitalar. A disponibilidade de um servi\u00e7o de RCV deveria inclusive ser considerada como um pr\u00e9-requisito obrigat\u00f3rio para que uma institui\u00e7\u00e3o m\u00e9dica fosse reconhecida ou acreditada como de excel\u00eancia em cardiologia.68 que, aliadas a um reduzido encaminhamento m\u00e9dico e uma baixa disponibilidade de servi\u00e7os, conduzem a uma participa\u00e7\u00e3o efetiva muito reduzida dos pacientes em programas de exerc\u00edcios f\u00edsicos supervisionados. Nesse contexto, programas de supervis\u00e3o indireta, realizados no ambiente domiciliar (RCVD), surgem como alternativa ou complementa\u00e7\u00e3o aos programas tradicionais e presenciais de RCV. Em virtude da sua maior abrang\u00eancia, a RCVD pode ser considerada o principal modo de interven\u00e7\u00e3o quando se trata de estrat\u00e9gia de sa\u00fade p\u00fablica, visando \u00e0 massifica\u00e7\u00e3o da RCV na popula\u00e7\u00e3o.O acesso e a ades\u00e3o dos pacientes a um programa presencial de RCV apresenta diversas barreiras, 61 incluiu 23 estudos com 2.890 pacientes cardiopatas e foram comparados os efeitos das RCV convencional e domiciliar. N\u00e3o foram encontradas diferen\u00e7as em mortalidade, capacidade f\u00edsica e qualidade de vida. Portanto, a escolha da participa\u00e7\u00e3o em programas formais ou domiciliares depende da disponibilidade de servi\u00e7os e das prefer\u00eancias individuais dos pacientes.Uma revis\u00e3o sistem\u00e1tica da Cochrane Entende-se como RCVD a pr\u00e1tica de exerc\u00edcios f\u00edsicos sem supervis\u00e3o presencial, mas orientada e acompanhada pelos profissionais do servi\u00e7o de RCV. Por essa raz\u00e3o, tamb\u00e9m \u00e9 chamada de reabilita\u00e7\u00e3o semi-supervisionada, com supervis\u00e3o indireta ou \u00e0 dist\u00e2ncia. As indica\u00e7\u00f5es e os objetivos da RCVD s\u00e3o os mesmos do modelo convencional, exigindo os mesmos cuidados em rela\u00e7\u00e3o \u00e0 avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o e \u00e0 prescri\u00e7\u00e3o de exerc\u00edcios. A maioria das sess\u00f5es \u00e9 realizada sob supervis\u00e3o indireta, mas a participa\u00e7\u00e3o em algumas sess\u00f5es presenciais, especialmente no in\u00edcio do programa, \u00e9 de fundamental import\u00e2ncia para consolidar o aprendizado das orienta\u00e7\u00f5es sobre a prescri\u00e7\u00e3o e esclarecer d\u00favidas. Os exerc\u00edcios podem ser realizados no pr\u00f3prio domic\u00edlio ou em parques, vias p\u00fablicas, gin\u00e1sios, academias, entre outros, com automonitoramento pelos pacientes, seguindo as orienta\u00e7\u00f5es recebidas.Sendo assim, para se obter uma adequada RCVD como estrat\u00e9gia populacional, primeiramente \u00e9 necess\u00e1rio ampliar a disponibilidade e capacidade de atendimento de programas presenciais de RCV, a fim de possibilitar avalia\u00e7\u00e3o inicial, orienta\u00e7\u00f5es, prescri\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos e seguimento das sess\u00f5es domiciliares, com ajustes peri\u00f3dicos por reavalia\u00e7\u00f5es. A estrat\u00e9gia domiciliar deve estar alinhada com a da RCV convencional, pois as duas modalidades s\u00e3o paralelas, com pacientes de diferentes perfis de risco, ou sequenciais, com o mesmo paciente em dois momentos cl\u00ednicos diferentes.smartphones podem intermediar a troca de informa\u00e7\u00f5es entre os pacientes e a equipe assistencial.Portanto, assim como a RCV convencional, a primeira etapa da RCVD \u00e9 o encaminhamento pelo m\u00e9dico assistente, seguido da avalia\u00e7\u00e3o pelo m\u00e9dico da reabilita\u00e7\u00e3o e demais profissionais, idealmente com realiza\u00e7\u00e3o do teste de esfor\u00e7o (TCPE ou TE) e/ou outras avalia\u00e7\u00f5es de aptid\u00e3o f\u00edsica. Ap\u00f3s a avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o, os pacientes definidos como de alto risco podem ser priorizados para a RCV presencial. J\u00e1 aqueles de menor risco, capazes de automonitoramento e conforme prefer\u00eancias individuais, podem ser direcionados \u00e0 RCVD. Ap\u00f3s receberem as instru\u00e7\u00f5es sobre a prescri\u00e7\u00e3o dos exerc\u00edcios, os pacientes executam as sess\u00f5es por conta pr\u00f3pria, podendo haver documenta\u00e7\u00e3o dos exerc\u00edcios em planilhas impressas ou eletr\u00f4nicas, com utiliza\u00e7\u00e3o de recursos como cardiofrequenc\u00edmetros, ped\u00f4metros ou medidores de velocidade e dist\u00e2ncia percorrida por GPS. Aplicativos de Em alguns casos, um programa de RCV combinado, com sess\u00f5es presenciais e domiciliares, pode ser a op\u00e7\u00e3o para pacientes de risco moderado, em aprendizagem sobre o automonitoramento ou com dificuldade de comparecer \u00e0s sess\u00f5es presenciais por problemas sociais ou de deslocamento. A propor\u00e7\u00e3o dessa combina\u00e7\u00e3o pode variar de acordo com as caracter\u00edsticas cl\u00ednicas do paciente e a estrutura log\u00edstica do servi\u00e7o.Portanto, o foco \u00e9 tornar os pacientes fisicamente mais ativos, sendo imperativa a redu\u00e7\u00e3o do sedentarismo e suas nefastas consequ\u00eancias. Para tal, \u00e9 fundamental a utiliza\u00e7\u00e3o isolada ou combinada dos recursos dispon\u00edveis, seja a atividade f\u00edsica informal, a reabilita\u00e7\u00e3o domiciliar ou a convencional.70A RCV deve estar integrada ao tratamento cl\u00ednico pleno das DCV, que consiste na a\u00e7\u00e3o sin\u00e9rgica das mudan\u00e7as estruturadas de estilo de vida com o tratamento farmacol\u00f3gico otimizado, com interven\u00e7\u00f5es indicadas quando existe instabiliza\u00e7\u00e3o cl\u00ednica e/ou refratariedade ao tratamento cl\u00ednico inicial. Nos pacientes com DAC est\u00e1vel, at\u00e9 mesmo com isquemia moderada ou grave, a adi\u00e7\u00e3o de tratamentos intervencionistas n\u00e3o t\u00eam se mostrado superiores na redu\u00e7\u00e3o de desfechos maiores . 73 Neste contexto, podem ser necess\u00e1rios ajustes de doses e/ou adi\u00e7\u00e3o de f\u00e1rmacos previamente ao in\u00edcio do programa de exerc\u00edcios f\u00edsicos. Por outro lado, ap\u00f3s o in\u00edcio da RCV e adequada ades\u00e3o aos exerc\u00edcios, alguns pacientes podem requerer retirada ou redu\u00e7\u00f5es de doses de f\u00e1rmacos, em virtude da assimila\u00e7\u00e3o ao treinamento f\u00edsico, como, por exemplo, em casos de hipotens\u00e3o sintom\u00e1tica, bradicardia acentuada e hipoglicemia. 75Para aumentar a efic\u00e1cia e a seguran\u00e7a da RCV, \u00e9 importante que o tratamento farmacol\u00f3gico da DCV esteja adequadamente ajustado, visando aumentar a toler\u00e2ncia ao esfor\u00e7o, o que favorece a execu\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos, reduzindo o risco de eventos. 76 Por isso, para a promo\u00e7\u00e3o da sa\u00fade e preven\u00e7\u00e3o de DCV, as diretrizes m\u00e9dicas t\u00eam recomendado, no m\u00ednimo, a pr\u00e1tica de exerc\u00edcio f\u00edsico de intensidade moderada por, pelo menos, 150 minutos semanais ou de alta intensidade por 75 minutos semanais (recomenda\u00e7\u00e3o 1 B). 83 A pr\u00e1tica de mais de 300 minutos semanais de exerc\u00edcio de intensidade moderada a alta pode conferir benef\u00edcio adicional, conforme j\u00e1 foi evidenciado em pacientes com DAC. 84Existe associa\u00e7\u00e3o entre o tempo de sedentarismo, com atividades como assistir \u00e0 televis\u00e3o, e maior mortalidade por todas as causas, bem como mortalidade cardiovascular. De acordo com a avalia\u00e7\u00e3o individual, a prescri\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos pode variar em rela\u00e7\u00e3o \u00e0s suas diversas caracter\u00edsticas, como tipo , modalidade e dura\u00e7\u00e3o (tempo de execu\u00e7\u00e3o), devendo se considerar a frequ\u00eancia semanal e a intensidade e 3 .Pacientes sedent\u00e1rios devem iniciar os exerc\u00edcios no limite inferior da prescri\u00e7\u00e3o, progredindo gradativamente ao longo das semanas seguintes. A progress\u00e3o inicial pode ser feita na dura\u00e7\u00e3o da sess\u00e3o e, posteriormente, na intensidade dos exerc\u00edcios. Pacientes j\u00e1 fisicamente ativos podem realizar, desde o in\u00edcio, exerc\u00edcios em n\u00edveis mais intensos, objetivando um m\u00ednimo de 75 minutos, divididos em duas ou mais sess\u00f5es semanais.Os exerc\u00edcios de resist\u00eancia muscular localizada, seja de fortalecimento ou de pot\u00eancia, t\u00eam se mostrado bastante ben\u00e9ficos para a sa\u00fade geral e para os sistemas cardiovascular e osteomuscular, sendo de fundamental import\u00e2ncia nos pacientes com sarcopenia e/ou osteopenia. Devem ser realizados, pelo menos, duas vezes por semana, privilegiando grandes grupos musculares de MMSS, MMII e tronco. Podem ser feitos utilizando o pr\u00f3prio peso corporal do indiv\u00edduo ou usando implementos como pesos livres, caneleiras, faixas el\u00e1sticas, aparelhos de muscula\u00e7\u00e3o, entre outros recursos. A carga ou peso, para cada exerc\u00edcio ou movimento, deve ser individualmente ajustada, al\u00e9m de se ter a devida aten\u00e7\u00e3o \u00e0 execu\u00e7\u00e3o dos movimentos para que a t\u00e9cnica e a postura sejam corretas.Existem diferentes protocolos para exerc\u00edcios resistidos, com varia\u00e7\u00f5es no n\u00famero de exerc\u00edcios utilizados por sess\u00e3o, como, por exemplo, de 6 a 15; na quantidade de s\u00e9ries para cada exerc\u00edcio, em geral de 1 a 3; e no n\u00famero de repeti\u00e7\u00f5es, que pode oscilar entre 6 e 20. A intensidade dos exerc\u00edcios resistidos pode ser ajustada de acordo com a intensidade relativa da for\u00e7a m\u00e1xima e pode ser expressa em fun\u00e7\u00e3o da carga m\u00e1xima poss\u00edvel para realizar uma repeti\u00e7\u00e3o m\u00e1xima (Teste de 1 repeti\u00e7\u00e3o m\u00e1xima ou 1RM). Carga de intensidade leve seria at\u00e9 30% de 1RM; intensidade m\u00e9dia, entre 30 e 60 ou 70% de 1RM; e intensidade alta, acima de 60 ou 70% de 1RM. Outra possibilidade \u00e9 a prescri\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos resistidos de modo subjetivo, pela sensa\u00e7\u00e3o de esfor\u00e7o ver .Um modo pr\u00e1tico \u00e9 o m\u00e9todo de repeti\u00e7\u00e3o vari\u00e1vel, que tem como objetivo executar uma faixa de repeti\u00e7\u00f5es . Se o paciente n\u00e3o conseguir realizar corretamente o movimento na repeti\u00e7\u00e3o m\u00ednima prescrita, significa que a carga aplicada est\u00e1 elevada. Por outro lado, se conseguir a repeti\u00e7\u00e3o m\u00e1xima prescrita de modo f\u00e1cil, \u00e9 porque a carga est\u00e1 leve. Assim, a carga ser\u00e1 ajustada para que o treinamento ocorra dentro da faixa de repeti\u00e7\u00f5es proposta. Esse m\u00e9todo pode ser aplicado aos mais variados exerc\u00edcios localizados e pode ser utilizado na progress\u00e3o da prescri\u00e7\u00e3o, sendo que os limites da repeti\u00e7\u00e3o podem ser modificados, dependendo dos objetivos almejados .Os exerc\u00edcios de flexibilidade podem oferecer benef\u00edcios osteomioarticulares, na qualidade de vida relacionada \u00e0 sa\u00fade e na preven\u00e7\u00e3o de queda em idosos. Ao contribu\u00edrem para uma movimenta\u00e7\u00e3o articular mais f\u00e1cil e eficiente, reduzem a demanda por oxig\u00eanio em situa\u00e7\u00f5es de movimento, favorecendo o desempenho do sistema cardiovascular. Nesses exerc\u00edcios, procura-se alcan\u00e7ar a amplitude m\u00e1xima do movimento, chegando at\u00e9 o ponto de leve desconforto, devendo a posi\u00e7\u00e3o ser mantida estaticamente por 10 a 30 segundos.86Dependendo da faixa et\u00e1ria, das condi\u00e7\u00f5es cl\u00ednicas e dos objetivos do programa de exerc\u00edcio para um dado paciente, outros tipos de exerc\u00edcio podem ser inclu\u00eddos na prescri\u00e7\u00e3o, como os de coordena\u00e7\u00e3o motora e de equil\u00edbrio. Al\u00e9m disso, devem ser considerados os in\u00fameros benef\u00edcios decorrentes de formas mais l\u00fadicas e socializantes de exerc\u00edcios, como a dan\u00e7a e outras modalidades. 87 \u00c9 tamb\u00e9m poss\u00edvel obter subs\u00eddios cl\u00ednicos e funcionais que possibilitem um adequado aconselhamento da atividade sexual, com base no modelo do KiTOMI, que foi proposto por autores brasileiros em 2016. 88 Al\u00e9m disso, \u00e9 fundamental para o paciente a reavalia\u00e7\u00e3o, com o intuito de estimular o comprometimento e mensurar a evolu\u00e7\u00e3o e os benef\u00edcios obtidos.A avalia\u00e7\u00e3o da aptid\u00e3o f\u00edsica aer\u00f3bica e n\u00e3o aer\u00f3bica possibilita uma prescri\u00e7\u00e3o mais individualizada dos exerc\u00edcios f\u00edsicos, com o objetivo de se obterem os melhores resultados e, por meio da estratifica\u00e7\u00e3o de risco e da busca de eventuais anormalidades, minimizar os riscos da pr\u00e1tica. De modo geral, a avalia\u00e7\u00e3o inicial tem como base a anamnese, o exame f\u00edsico e o ECG. Avalia\u00e7\u00f5es mais detalhadas dever\u00e3o ser individualizadas, com realiza\u00e7\u00e3o de TCPE ou TE, avalia\u00e7\u00e3o antropom\u00e9trica, de for\u00e7a/pot\u00eancia muscular e de flexibilidade. Na avalia\u00e7\u00e3o inicial, pode-se quantificar o d\u00e9ficit funcional frente ao desej\u00e1vel, bem como estabelecer metas a serem alcan\u00e7adas. \u00c9 importante enfatizar que os pacientes com baixa aptid\u00e3o f\u00edsica inicial s\u00e3o os que mais se beneficiam da RCV, ap\u00f3s adequada ader\u00eancia ao programa de exerc\u00edcio supervisionado. Finalizando, vale ressaltar a fundamental import\u00e2ncia do estabelecimento de um sistem\u00e1tico esquema de reavalia\u00e7\u00f5es, que, al\u00e9m de estimular o comprometimento dos pacientes, torne poss\u00edvel mensurar a evolu\u00e7\u00e3o e os benef\u00edcios obtidos, produzindo relat\u00f3rios que estimulem os ajustes do tratamento e que, portanto, devem ser sempre encaminhados aos m\u00e9dicos assistentes, os quais obviamente devem integrar ativamente o tratamento cl\u00ednico pleno.89 Houve um aumento global do n\u00famero de hipertensos de 594 milh\u00f5es em 1975 para 1,13 bilh\u00e3o em 2015, em grande parte creditado aos pa\u00edses subdesenvolvidos e em desenvolvimento. 90 Considerando que a maioria dos casos est\u00e1 relacionada ao estilo de vida, com o sedentarismo ocupando lugar de destaque, fica clara a import\u00e2ncia dos exerc\u00edcios f\u00edsicos ao lado de outras medidas comportamentais, al\u00e9m do uso de medica\u00e7\u00f5es, sempre que indicadas. 72A hipertens\u00e3o arterial sist\u00eamica (HAS) permanece como um dos maiores fatores de risco para o desenvolvimento de DAC, IC, DRC e acidente vascular cerebral (AVC) isqu\u00eamico ou hemorr\u00e1gico, representando, social e economicamente, um enorme desafio \u00e0 sa\u00fade p\u00fablica mundial. 91 A pr\u00e1tica regular de exerc\u00edcios f\u00edsicos exerce efeito terap\u00eautico na reestrutura\u00e7\u00e3o fisiol\u00f3gica desses sistemas, com redu\u00e7\u00e3o do estresse oxidativo e da inflama\u00e7\u00e3o, corre\u00e7\u00e3o da disfun\u00e7\u00e3o barorreflexa, aumento do t\u00f4nus vagal, diminui\u00e7\u00e3o da atividade simp\u00e1tica, revers\u00e3o do remodelamento hipertr\u00f3fico arteriolar em tecidos exercitados e redu\u00e7\u00e3o da resist\u00eancia vascular perif\u00e9rica, com consequente diminui\u00e7\u00e3o da PA e controle dos n\u00edveis press\u00f3ricos semelhante, ou mesmo superior, ao proporcionado pela farmacoterapia. 93A HAS apresenta fisiopatologia complexa e multifatorial, com modifica\u00e7\u00f5es estruturais e fisiol\u00f3gicas, em particular, nos sistemas vascular , renal e neural . 94 Todas essas altera\u00e7\u00f5es estruturais da parede do vaso, que ocorrem tanto em territ\u00f3rio arterial como arteriolar, elevam a rigidez do sistema vascular, com consequente aumento da velocidade da onda de pulso do fluxo sangu\u00edneo, da press\u00e3o de pulso (diferen\u00e7a entre a PA sist\u00f3lica [PAS] e a diast\u00f3lica [PAD]) e da press\u00e3o hidrost\u00e1tica no capilar. Soma-se a todo esse desequil\u00edbrio estrutural a disfun\u00e7\u00e3o do endot\u00e9lio, com o aumento de subst\u00e2ncias vasoconstritoras, de mediadores inflamat\u00f3rios e de agentes oxidantes, em detrimento da produ\u00e7\u00e3o de agentes vasodilatadores e antioxidantes. 96No tecido vascular, a HAS caracteriza-se por desorganiza\u00e7\u00e3o das c\u00e9lulas musculares lisas, aumento dos dep\u00f3sitos de col\u00e1geno e diminui\u00e7\u00e3o da raz\u00e3o elastina/col\u00e1geno, al\u00e9m da forma\u00e7\u00e3o de fibra el\u00e1stica anormal e l\u00e2mina el\u00e1stica interna com menor \u00e1rea fenestrada. shear stress \u201d), estimula positivamente o tecido endotelial, com aumento da produ\u00e7\u00e3o de enzimas antioxidantes e agentes vasodilatadores, al\u00e9m de diminui\u00e7\u00e3o da a\u00e7\u00e3o dos radicais livres, das citocinas pr\u00f3-inflamat\u00f3rias, das mol\u00e9culas de ades\u00e3o e dos agentes vasoconstritores, restaurando, assim, o equil\u00edbrio do funcionamento endotelial. 98 Estudos experimentais 94 em ratos espontaneamente hipertensos demonstram a reorganiza\u00e7\u00e3o de todas as estruturas vasculares da art\u00e9ria aorta ap\u00f3s a implementa\u00e7\u00e3o de um per\u00edodo de exerc\u00edcio aer\u00f3bico. O treinamento aer\u00f3bico promove adapta\u00e7\u00f5es vasculares nas art\u00e9rias de condut\u00e2ncia , nas arter\u00edolas (pela diminui\u00e7\u00e3o da raz\u00e3o parede/luz do vaso) e nos capilares, estimulando a angiog\u00eanese. 100O exerc\u00edcio f\u00edsico, por meio do aumento do estresse tangencial derivado da fric\u00e7\u00e3o do fluxo sangu\u00edneo na superf\u00edcie endotelial da parede do vaso , que t\u00eam demonstrado efeitos expressivos na redu\u00e7\u00e3o dos n\u00edveis tensionais. 109 A aptid\u00e3o f\u00edsica, medida objetivamente por meio de testes de esfor\u00e7o graduados, atenua o aumento da press\u00e3o com a idade e impede o desenvolvimento de hipertens\u00e3o. Em uma coorte de homens de 20 a 90 anos de idade, que foram seguidos por 3 a 28 anos, uma maior aptid\u00e3o f\u00edsica diminuiu a taxa de aumento press\u00f3rico ao longo do tempo e atrasou o per\u00edodo at\u00e9 o in\u00edcio da HAS. 110 Estudos epidemiol\u00f3gicos t\u00eam revelado associa\u00e7\u00e3o inversa entre o n\u00edvel de atividade f\u00edsica e a aptid\u00e3o cardiorrespirat\u00f3ria, com a presen\u00e7a de hipertens\u00e3o arterial. 112Maiores n\u00edveis de atividade f\u00edsica t\u00eam sido associados a uma diminui\u00e7\u00e3o no risco de desenvolvimento de HAS. Com o advento dos rastreadores eletr\u00f4nicos de atividade e do monitoramento ambulatorial da PA, tornou-se cada vez mais vi\u00e1vel a realiza\u00e7\u00e3o de estudos que correlacionem a atividade f\u00edsica e a PA. 112 Al\u00e9m disso, a pr\u00e1tica constante de atividades f\u00edsicas pode ser ben\u00e9fica tanto na preven\u00e7\u00e3o quanto no tratamento da hipertens\u00e3o, reduzindo a morbimortalidade cardiovascular. Indiv\u00edduos ativos apresentam um risco at\u00e9 30% menor de desenvolver hipertens\u00e3o que os sedent\u00e1rios 111 e o aumento da atividade f\u00edsica di\u00e1ria reduz a press\u00e3o aterial de maneira significativa. 113Os grandes ensaios cl\u00ednicos randomizados e as meta-an\u00e1lises t\u00eam confirmado que o exerc\u00edcio f\u00edsico regular pode reduzir os n\u00edveis press\u00f3ricos. 114 por ser o mais prevalente dos fatores de risco cardiovasculares e um dos principais fatores contribuintes para mortalidade no mundo. 115 A sobrevida \u00e9 menor em pessoas que passam a maior parte do tempo sentadas do que naquelas que passam pouco tempo desse modo. 116 H\u00e1 rela\u00e7\u00e3o direta entre o per\u00edodo sentado ou o tempo de televis\u00e3o com n\u00edveis elevados de PA, morbidade e mortalidade cardiovascular. 117 Por esta raz\u00e3o, para a redu\u00e7\u00e3o do tempo sentado, recomenda-se levantar-se por, pelo menos, 5 minutos a cada 30 minutos sentado, como medida v\u00e1lida de preven\u00e7\u00e3o. A pr\u00e1tica de exerc\u00edcios f\u00edsicos est\u00e1 indicada para todos os pacientes com HAS de atividade aer\u00f3bia de moderada a alta intensidade, sendo aconselh\u00e1vel associar duas a tr\u00eas sess\u00f5es de exerc\u00edcios resistidos por semana. Na aus\u00eancia de contraindica\u00e7\u00f5es, pode ocorrer aumento gradativo, visando a meta de 300 min/semana de exerc\u00edcios aer\u00f3bios de intensidade moderada ou 150 min/semana de exerc\u00edcios aer\u00f3bios de alta intensidade, para obten\u00e7\u00e3o de maiores benef\u00edcios.37 ou a redu\u00e7\u00e3o da intensidade de treinamento at\u00e9 a obten\u00e7\u00e3o de melhor controle press\u00f3rico. Em programas de RCV supervisionados, flexibiliza\u00e7\u00f5es dessas recomenda\u00e7\u00f5es podem ser realizadas individualmente, de acordo com a avalia\u00e7\u00e3o do m\u00e9dico da reabilita\u00e7\u00e3o e as respostas observadas no teste de esfor\u00e7o e nas sess\u00f5es de exerc\u00edcios. Durante o exerc\u00edcio, \u00e9 recomendado que a PA se mantenha inferior a 220/105 mmHg. Se estiver superior a esse n\u00edvel, deve-se considerar a interrup\u00e7\u00e3o da sess\u00e3o ou a redu\u00e7\u00e3o da intensidade de cargas, considerando o ajuste das medica\u00e7\u00f5es. 37Durante o treinamento, \u00e9 importante que a PA seja avaliada em repouso e em esfor\u00e7o. Para pacientes com valores em repouso superiores a 160/100 mmHg ou com les\u00e3o de \u00f3rg\u00e3os-alvo , \u00e9 recomendado o ajuste dos f\u00e1rmacos anti-hipertensivos para melhor controle press\u00f3rico antes de iniciar ou retornar \u00e0s sess\u00f5es de exerc\u00edcio, 120 Esse efeito agudo do treinamento f\u00edsico pode causar hipotens\u00e3o sintom\u00e1tica ap\u00f3s o t\u00e9rmino, que geralmente melhora com repouso e hidrata\u00e7\u00e3o. Pacientes em uso de alfabloqueadores, betabloqueadores, bloqueadores de canais de c\u00e1lcio e vasodilatadores podem ter maior risco de hipotens\u00e3o p\u00f3s-exerc\u00edcio, necessitando de aten\u00e7\u00e3o especial no desaquecimento. A recorr\u00eancia dessa situa\u00e7\u00e3o, que costuma decorrer do resultado da assimila\u00e7\u00e3o do treinamento que se soma aos efeitos anti-hipertensivos dos f\u00e1rmacos, exige considerar a necessidade de ajustes das doses ou mesmo suspens\u00e3o de medicamentos.Ap\u00f3s a sess\u00e3o de exerc\u00edcio, a PA deve ser verificada e \u00e9 comum a identifica\u00e7\u00e3o de valor inferior ao observado antes do in\u00edcio das atividades. Em hipertensos, o efeito anti-hipertensivo agudo de uma sess\u00e3o tende a ser maior com n\u00edveis mais intensos de exerc\u00edcios aer\u00f3bios. 121 Embora tais efeitos precisem ser reproduzidos em mais estudos, exerc\u00edcio em \u00e1gua aquecida parece ser apropriado para pacientes com hipertens\u00e3o arterial resistente.H\u00e1 poucos dados quanto ao efeito do exerc\u00edcio em pacientes com hipertens\u00e3o resistente, que se caracteriza pela PA acima da meta apesar do uso de tr\u00eas ou mais medica\u00e7\u00f5es anti-hipertensivas. Em rela\u00e7\u00e3o a esses pacientes, que requerem maior monitoramento, um ensaio cl\u00ednico randomizado unic\u00eantrico mostrou que exerc\u00edcio em \u00e1gua aquecida (30 a 32\u00baC) resultou em pronunciada redu\u00e7\u00e3o da PA (36/12 mmHg) ap\u00f3s 3 meses. 124 Os mecanismos subjacentes da DAC est\u00e1vel incluem obstru\u00e7\u00e3o ateroscler\u00f3tica dos vasos epic\u00e1rdicos, doen\u00e7a microvascular e espasmo coron\u00e1rio, isolados ou em associa\u00e7\u00e3o. 5 Clinicamente a manifesta\u00e7\u00e3o mais comum da DAC est\u00e1vel \u00e9 a angina do peito, que se caracteriza por epis\u00f3dios revers\u00edveis de dor tor\u00e1cica por isquemia mioc\u00e1rdica, decorrentes do desequil\u00edbrio entre oferta e consumo de oxig\u00eanio pelo mioc\u00e1rdio, em geral desencadeados pelo esfor\u00e7o f\u00edsico ou estresse emocional, que cessam com o repouso ou uso de nitrato de a\u00e7\u00e3o r\u00e1pida. 5As DCV, lideradas pela DAC, s\u00e3o respons\u00e1veis pela maior parte das mortes da popula\u00e7\u00e3o adulta. 125 sendo fundamental o tratamento cl\u00ednico pleno, com otimiza\u00e7\u00e3o do tratamento farmacol\u00f3gico e pr\u00e1tica de exerc\u00edcios f\u00edsicos regulares, al\u00e9m de outras modifica\u00e7\u00f5es comportamentais relacionadas a tabagismo, dieta e composi\u00e7\u00e3o corporal. Revasculariza\u00e7\u00f5es eletivas tamb\u00e9m podem ser indicadas nos pacientes com DAC est\u00e1vel, a depender da sintomatologia e do risco cardiovascular. 5 Por\u00e9m, vale ressaltar que, quando o quadro \u00e9 est\u00e1vel, mesmo nos pacientes com angina, o tratamento exclusivamente cl\u00ednico n\u00e3o tem se mostrado inferior ao tratamento com adi\u00e7\u00e3o de abordagem intervencionista. 127 A ocorr\u00eancia de eventos agudos de instabiliza\u00e7\u00e3o da doen\u00e7a, com quadros de IAM ou angina inst\u00e1vel, est\u00e1 relacionada com elevado aumento do risco cardiovascular, frequentemente exigindo ajustes da terapia farmacol\u00f3gica e revasculariza\u00e7\u00e3o cir\u00fargica ou percut\u00e2nea de urg\u00eancia. 131A DAC est\u00e1vel tem bom progn\u00f3stico, com mortalidade anual estimada em 1,5% e incid\u00eancia de infarto n\u00e3o fatal de 1,4%, 136 a melhora da capacidade funcional cardiorrespirat\u00f3ria 136 e a melhora perfusional cintilogr\u00e1fica. 140 Os benef\u00edcios adquiridos persistem com a manuten\u00e7\u00e3o da pr\u00e1tica regular de exerc\u00edcios f\u00edsicos, 144 que contribui para a melhora da qualidade de vida 146 e redu\u00e7\u00e3o das taxas de mortalidade cardiovascular e hospitaliza\u00e7\u00e3o. 148Est\u00e3o cientificamente demonstrados os efeitos ben\u00e9ficos do exerc\u00edcio f\u00edsico regular realizado em curto e longo prazos nos portadores de DAC est\u00e1vel. Em um per\u00edodo inicial de reabilita\u00e7\u00e3o cardiovascular, de 8 a 12 semanas, destacam-se o aumento do limiar isqu\u00eamico, 144 Destaca-se ainda um aumento da perfus\u00e3o mioc\u00e1rdica decorrente da melhora da resposta vasodilatadora dependente do endot\u00e9lio 151 e do aumento do recrutamento de vasos colaterais durante o exerc\u00edcio, 152 algo que reflete na atenua\u00e7\u00e3o da depress\u00e3o do segmento ST durante o exerc\u00edcio. 137 Ressalta-se, ainda, que o treinamento f\u00edsico associado \u00e0 dieta pobre em gorduras pode influenciar na progress\u00e3o da placa ateroscler\u00f3tica. 153Em pacientes com DAC est\u00e1vel, diferentes mecanismos explicam o aumento do limiar isqu\u00eamico, permitindo gradativamente cargas superiores de atividade f\u00edsica. A redu\u00e7\u00e3o do duplo produto para cargas subm\u00e1ximas de trabalho est\u00e1 associada, dentre outros mecanismos, \u00e0 melhora da modula\u00e7\u00e3o auton\u00f4mica card\u00edaca. 1 de 63 estudos envolvendo 14.486 pacientes com idade entre 47 e 71 anos revelaram que a RCV reduziu a mortalidade cardiovascular em 26% e a hospitaliza\u00e7\u00e3o global em 18%, com melhora adicional na qualidade de vida nessa popula\u00e7\u00e3o, devendo ser encorajada sempre que poss\u00edvel.A RCV \u00e9 uma terapia adjuvante eficaz no tratamento de pacientes ap\u00f3s evento coronariano agudo, CRVM e ICP. Revis\u00e3o sistem\u00e1tica e meta-an\u00e1lise 154 Outros estudos similares reportaram redu\u00e7\u00e3o da mortalidade card\u00edaca ou total entre 8 e 34% para cada MET de melhora na capacidade cardiorrespirat\u00f3ria. 156A melhora da capacidade cardiorrespirat\u00f3ria \u00e9 um dos fatores respons\u00e1veis pelos achados na redu\u00e7\u00e3o da mortalidade total. Em coorte realizada com 5.641 pacientes participantes de RCV no Canad\u00e1, verificou-se que cada 1 MET de aumento na capacidade cardiorrespirat\u00f3ria durante a RCV reduziu a mortalidade total em 25%. Exercise Training Intervention After Coronary Angioplasty ), em que houve aumento de 26% no VO 2 pico, melhora de 27% na qualidade de vida e redu\u00e7\u00e3o de 20% nos eventos card\u00edacos, incluindo diminui\u00e7\u00e3o de IAM e menor n\u00famero de hospitaliza\u00e7\u00f5es em pacientes que realizaram RCV ap\u00f3s angioplastia, quando comparados aos que permaneceram sedent\u00e1rios. 157Al\u00e9m disso, a RCV oferece efeito adicional na redu\u00e7\u00e3o de eventos cardiovasculares ap\u00f3s ICP, conforme evidenciado pelo estudo ETICA , com diminui\u00e7\u00e3o desses efeitos entre 7 e 28 dias e superior a 29 dias, quando progressivamente se perdia o efeito positivo sobre o remodelamento ventricular. \u00c9 importante ressaltar que n\u00e3o houve diferen\u00e7a em rela\u00e7\u00e3o a eventos entre as fases de in\u00edcio do treinamento e que a amostra estudada foi prioritariamente de homens jovens, o que refor\u00e7a a necessidade de mais estudos, principalmente em outras popula\u00e7\u00f5es, como a de idosos e mulheres. Para cada 1 semana de atraso no in\u00edcio dos exerc\u00edcios ap\u00f3s o infarto, poder\u00e1 ser necess\u00e1rio 1 m\u00eas adicional de treinamento para obten\u00e7\u00e3o de benef\u00edcios similares no volume sist\u00f3lico final e na fra\u00e7\u00e3o de eje\u00e7\u00e3o do ventr\u00edculo esquerdo (FEVE). 160Uma das maiores preocupa\u00e7\u00f5es do treinamento f\u00edsico precoce refere-se ao seu efeito no processo de remodelamento ventricular. Enquanto alguns autores reportam efeitos negativos, 168 Sendo assim, mudan\u00e7as pol\u00edticas, sociais, estruturais e na cultura m\u00e9dica s\u00e3o necess\u00e1rias para modificar esse cen\u00e1rio.Embora referendada amplamente pela literatura m\u00e9dica por seus efeitos ben\u00e9ficos e custo-efetividade, somente uma minoria dos pacientes eleg\u00edveis participa de programas de RCV, algo explic\u00e1vel por m\u00faltiplas barreiras, como inexist\u00eancia de programas, dificuldade de acesso aos servi\u00e7os, reduzido n\u00famero de encaminhamentos, mobilidade urbana de m\u00e1 qualidade, entre outros, afetando principalmente mulheres, idosos e minorias \u00e9tnicas. Tanto nos pacientes com DAC est\u00e1vel como naqueles ap\u00f3s evento coron\u00e1rio e/ou revasculariza\u00e7\u00f5es, \u00e9 fundamental a estratifica\u00e7\u00e3o de risco para a RCV, por meio de avalia\u00e7\u00e3o cl\u00ednica focada no conhecimento detalhado da DCV e nos tratamentos realizados, sejam medicamentosos ou intervencionistas. Quest\u00f5es relacionadas com exist\u00eancia de sintomas, fun\u00e7\u00e3o ventricular, capacidade funcional, presen\u00e7a de arritmias e possibilidade de isquemia residual auxiliam na estratifica\u00e7\u00e3o e devem fazer parte da avalia\u00e7\u00e3o inicial. O ideal \u00e9 que essa avalia\u00e7\u00e3o m\u00e9dica seja realizada por profissional integrado \u00e0 equipe da RCV (m\u00e9dico da reabilita\u00e7\u00e3o).O perfil de um paciente encaminhado \u00e0 RCV pode ser bastante variado, desde o que \u00e9 submetido a tratamento de maneira eletiva at\u00e9 aquele com s\u00edndrome coronariana aguda complicada e interna\u00e7\u00e3o prolongada. Uma avalia\u00e7\u00e3o mais ampla, incluindo quest\u00f5es nutricionais, psicol\u00f3gicas e osteomusculares, deve fazer parte da anamnese, pois esses fatores podem impactar diretamente no processo da RCV. Sempre devem ser realizadas cuidadosas avalia\u00e7\u00f5es, seja do local de pun\u00e7\u00e3o arterial, principalmente no acesso femoral, nos pacientes submetidos \u00e0 ICP, assim como das feridas cir\u00fargicas, em especial quanto \u00e0 estabilidade esternal e eventuais infec\u00e7\u00f5es, nos pacientes submetidos \u00e0 CRVM. A presen\u00e7a de situa\u00e7\u00e3o que implique necessidade de cuidados especiais exige as pertinentes orienta\u00e7\u00f5es \u00e0 equipe respons\u00e1vel pelo treinamento f\u00edsico dos pacientes.A avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o para a RCV, por meio de provas funcionais, objetiva o melhor conhecimento da capacidade funcional, a avalia\u00e7\u00e3o de isquemia residual e a pesquisa de arritmias induzidas pelo esfor\u00e7o. A identifica\u00e7\u00e3o de isquemia mioc\u00e1rdica ao esfor\u00e7o \u00e9 realizada por meio da ocorr\u00eancia de sintomas como angina de peito e/ou por altera\u00e7\u00f5es eletrocardiogr\u00e1ficas. O limiar isqu\u00eamico identificado no TE pelo in\u00edcio dessas altera\u00e7\u00f5es cl\u00ednicas e/ou eletrocardiogr\u00e1ficas, pode eventualmente ser caracterizado segundo a carga de trabalho e FC, a partir dos quais a isquemia se manifesta, algo que poderia ser utilizado na prescri\u00e7\u00e3o de exerc\u00edcio.O TE, para fins de prescri\u00e7\u00e3o, dever\u00e1 ser\u00e1 realizado sob o uso das medica\u00e7\u00f5es habituais, principalmente as que causam interfer\u00eancia na FC, para que haja reprodu\u00e7\u00e3o na avalia\u00e7\u00e3o da condi\u00e7\u00e3o que estar\u00e1 presente durante as sess\u00f5es de treinamento. Por exemplo, em situa\u00e7\u00f5es de pacientes que alteram a dose de betabloqueador durante a reabilita\u00e7\u00e3o, o ideal seria realizar um novo teste para ajuste da prescri\u00e7\u00e3o. Em caso de impossibilidade dessa conduta, o uso da percep\u00e7\u00e3o subjetiva de esfor\u00e7o poder\u00e1 auxiliar na prescri\u00e7\u00e3o at\u00e9 a realiza\u00e7\u00e3o de novo exame.Em alguns casos, os pacientes ingressantes na RCV podem estar com alguma limita\u00e7\u00e3o cl\u00ednica para realiza\u00e7\u00e3o de um teste funcional m\u00e1ximo. Nestes, pode-se realizar um teste inicial subm\u00e1ximo para guiar a prescri\u00e7\u00e3o, com posterior teste m\u00e1ximo ap\u00f3s a melhora cl\u00ednica e/ou otimiza\u00e7\u00e3o do tratamento farmacol\u00f3gico. Considerando a possibilidade de grandes erros, devido \u00e0 intensa varia\u00e7\u00e3o individual da resposta cronotr\u00f3pica, n\u00e3o devem ser usadas f\u00f3rmulas que consideram a idade para defini\u00e7\u00e3o da FC pico, sendo este erro ainda maior nos pacientes em uso de betabloqueadores.131 A intensidade tamb\u00e9m pode ser determinada pela ventila\u00e7\u00e3o pulmonar, sendo a atividade considerada de moderada intensidade quando o indiv\u00edduo permanece apenas discretamente ofegante, conseguindo falar frases completas sem interrup\u00e7\u00f5es e com limita\u00e7\u00e3o da FC de treinamento de modo arbitr\u00e1rio, ou seja, a utiliza\u00e7\u00e3o da FC de repouso + 20 bpm para pacientes que tiveram s\u00edndrome coronariana aguda, ou FC de repouso + 30 bpm para aqueles ap\u00f3s cirurgia ou tratamento intervencionista eletivo. \u00e7\u00f5es ver .Quando o TE for realizado, a intensidade dos exerc\u00edcios prescritos poder\u00e1 situar-se entre 40 e 80% da FC de reserva [m\u00e9todo de Karvonen: (FC pico \u2013 FC de repouso) x percentual de intensidade + FC de repouso]. Nesses casos, habitualmente se inicia com a FC no limite inferior da prescri\u00e7\u00e3o, sendo realizadas progress\u00f5es, conforme a evolu\u00e7\u00e3o cl\u00ednica e melhora da capacidade funcional. A maioria dos pacientes ter\u00e1 intensidade prescrita entre 50 e 70% da FC de reserva. Os mais limitados ou com disfun\u00e7\u00e3o ventricular importante poder\u00e3o trabalhar em intensidades menores, entre 40 e 60%, e aqueles previamente ativos e com melhor capacidade funcional, entre 50 e 80%. Os percentuais da FC pico tamb\u00e9m podem ser utilizados, sendo que intensidades moderadas correspondem de 70 a 85% da FC pico ver .169 Na presen\u00e7a de plat\u00f4 precoce do pulso de oxig\u00eanio ou, principalmente, queda dessa vari\u00e1vel durante o esfor\u00e7o, a prescri\u00e7\u00e3o da intensidade do exerc\u00edcio pode ser limitada \u00e0s cargas abaixo dessa ocorr\u00eancia. Desse modo, o TCPE \u00e9 considerado o padr\u00e3o-ouro na avalia\u00e7\u00e3o para a prescri\u00e7\u00e3o dos exerc\u00edcios e deve ser utilizado sempre que estiver dispon\u00edvel. 171 Nesses casos, a recomenda\u00e7\u00e3o de intensidades moderadas poder\u00e1 ser realizada entre os limiares ventilat\u00f3rios (limiar anaer\u00f3bico e ponto de compensa\u00e7\u00e3o respirat\u00f3ria), com evolu\u00e7\u00e3o da intensidade de maneira progressiva.O TCPE, por meio da an\u00e1lise da resposta do pulso de oxig\u00eanio, contribui para o aumento da sensibilidade e especificidade para o diagn\u00f3stico da isquemia mioc\u00e1rdica. Em rela\u00e7\u00e3o ao volume dos exerc\u00edcios, recomenda-se, no m\u00ednimo, 150 minutos por semana, que poder\u00e1 ser distribu\u00eddo em 3 a 5 sess\u00f5es. De acordo com a toler\u00e2ncia, adapta\u00e7\u00f5es ao treinamento e prefer\u00eancias individuais, al\u00e9m das considera\u00e7\u00f5es sobre o quadro cl\u00ednico, este volume poder\u00e1 aumentar para 300 minutos ou mais por semana.Em rela\u00e7\u00e3o ao treinamento resistido, o m\u00e9todo considerado padr\u00e3o-ouro para a prescri\u00e7\u00e3o da intensidade seria o teste de uma repeti\u00e7\u00e3o m\u00e1xima. Entretanto, na pr\u00e1tica, muitos programas de reabilita\u00e7\u00e3o n\u00e3o o utilizam pela limita\u00e7\u00e3o de tempo para aplic\u00e1-lo, ou mesmo por raz\u00f5es cl\u00ednicas, como em pacientes submetidos a CRVM, que podem ter limita\u00e7\u00f5es n\u00e3o apenas pela esternotomia, mas tamb\u00e9m por les\u00f5es da safenectomia. Nesses casos, a percep\u00e7\u00e3o subjetiva de esfor\u00e7o \u00e9 um m\u00e9todo pr\u00e1tico e \u00fatil.173Em pacientes com esternotomia, trabalhos com MMSS devem ter cargas restritas durante 5 a 8 semanas e limitadas a baixas intensidades. Exerc\u00edcios com amplitude de movimentos com os bra\u00e7os poder\u00e3o ser permitidos ap\u00f3s essa fase, se n\u00e3o existir instabilidade de esterno, embora novos estudos estejam avaliando a seguran\u00e7a de prescri\u00e7\u00e3o mais precoce do exerc\u00edcio ap\u00f3s a CRVM. Os pacientes devem sempre ser orientados quanto \u00e0 maneira correta de execu\u00e7\u00e3o do movimento e \u00e0 respira\u00e7\u00e3o, evitando a manobra de Valsalva. O intervalo entre as s\u00e9ries dos exerc\u00edcios resistidos pode ser entre 45 segundos e 1 minuto, a depender das cargas aplicadas e da toler\u00e2ncia do paciente.6.3.4.1. Angina Refrat\u00e1ria175 Tais pacientes geralmente n\u00e3o s\u00e3o referenciados aos programas de RCV, devido ao temor de eventos adversos durante o treinamento f\u00edsico, embora a reabilita\u00e7\u00e3o j\u00e1 tenha sido considerada como uma possibilidade terap\u00eautica exequ\u00edvel e segura para esses pacientes. 175Pacientes com angina refrat\u00e1ria s\u00e3o caracterizados por angina limitante com evolu\u00e7\u00e3o superior a 3 meses, em tratamento cl\u00ednico otimizado, com documenta\u00e7\u00e3o de isquemia mioc\u00e1rdica e n\u00e3o considerados eleg\u00edveis para interven\u00e7\u00e3o coronariana percut\u00e2nea e/ou cir\u00fargica. 178 H\u00e1 um \u00fanico estudo controlado envolvendo RCV em pacientes com angina refrat\u00e1ria. Ele avaliou 42 indiv\u00edduos, randomizados para o programa de exerc\u00edcios de RCV ou manuten\u00e7\u00e3o do tratamento cl\u00ednico habitual, durante 8 semanas. Os pacientes do grupo de exerc\u00edcio receberam a prescri\u00e7\u00e3o de FC de treino entre 60 e 75% da FC de reserva (para aqueles com fun\u00e7\u00e3o ventricular preservada) e entre 40 e 60% da FC de reserva quando a FEVE era inferior a 40%. Os pacientes do grupo de reabilita\u00e7\u00e3o aumentaram em 50 m a dist\u00e2ncia total no teste de caminhada , sem mudan\u00e7a na intensidade ou frequ\u00eancia da angina e sem eventos adversos em ambos os grupos. 161O objetivo das interven\u00e7\u00f5es terap\u00eauticas nesse cen\u00e1rio contribui para a melhora da qualidade de vida, facilitando a realiza\u00e7\u00e3o de atividades da vida di\u00e1ria. 162 est\u00e1 avaliando a seguran\u00e7a e efic\u00e1cia de um programa de exerc\u00edcios realizado durante um per\u00edodo de 12 semanas, supervisionado em ambiente hospitalar e com monitoramento eletrocardiogr\u00e1fico cont\u00ednuo. A prescri\u00e7\u00e3o est\u00e1 sendo individualizada e pautada nos par\u00e2metros do TCPE e no limiar de isquemia e/ou angina. At\u00e9 o momento, 42 pacientes foram inclu\u00eddos, e n\u00e3o houve documenta\u00e7\u00e3o de eventos cardiovasculares e hospitaliza\u00e7\u00f5es relacionadas aos exerc\u00edcios. A troponina T ultrassens\u00edvel, preditor conhecido de pior progn\u00f3stico, 179 n\u00e3o apresentou oscila\u00e7\u00e3o no seu n\u00edvel s\u00e9rico em 32 pacientes submetidos a uma sess\u00e3o aguda de exerc\u00edcio aer\u00f3bico (no limiar de isquemia) de 40 min de dura\u00e7\u00e3o, no momento da inclus\u00e3o no estudo ( dados n\u00e3o publicados ).Um estudo brasileiro randomizado ainda em andamento 180Nos pacientes com angina refrat\u00e1ria e baixo limiar isqu\u00eamico, a utiliza\u00e7\u00e3o de nitratos de a\u00e7\u00e3o r\u00e1pida antes do in\u00edcio da sess\u00e3o de treinamento f\u00edsico pode contribuir para um treino mais duradouro e at\u00e9 maiores intensidades de exerc\u00edcios. 6.3.4.2. Treinamento com Indu\u00e7\u00e3o de Isquemia Mioc\u00e1rdica182 pois as altera\u00e7\u00f5es contr\u00e1teis e os defeitos perfusionais precedem as altera\u00e7\u00f5es cl\u00ednicas e eletrocardiogr\u00e1ficas isqu\u00eamicas. 184Tradicionalmente, existe a recomenda\u00e7\u00e3o de que os exerc\u00edcios f\u00edsicos nos pacientes com DAC sejam realizados abaixo do limiar isqu\u00eamico cl\u00ednico e eletrocardiogr\u00e1fico. Por\u00e9m, isso pode ser dif\u00edcil de controlar. Estudos pr\u00e9vios j\u00e1 demonstraram que os exerc\u00edcios f\u00edsicos, prescritos conforme recomenda\u00e7\u00f5es da literatura, podem desencadear defeitos de perfus\u00e3o cintilogr\u00e1ficos, que n\u00e3o s\u00e3o evidenciados por meio de anormalidades no ECG e angina, 185 Outros autores demonstraram em pequena s\u00e9rie de pacientes que, ap\u00f3s seis semanas de treinamento em pacientes com DAC, est\u00edmulos isqu\u00eamicos repetitivos tamb\u00e9m n\u00e3o resultaram em danos, disfun\u00e7\u00f5es mioc\u00e1rdicas e arritmias significativas. 187O significado funcional da indu\u00e7\u00e3o de isquemia cintilogr\u00e1fica ainda \u00e9 incerto, mas estudos com realiza\u00e7\u00e3o de treinamentos acima do limiar isqu\u00eamico j\u00e1 foram realizados. Em um estudo com realiza\u00e7\u00e3o de uma \u00fanica sess\u00e3o de treinamento com dura\u00e7\u00e3o de 20 min acima do limiar isqu\u00eamico, n\u00e3o foram identificados ind\u00edcios de dano mioc\u00e1rdico agudo. 188 Adicionalmente, cabe enfatizar as evid\u00eancias da superioridade da combina\u00e7\u00e3o de treinos aer\u00f3bicos e resistidos em rela\u00e7\u00e3o a treinos aer\u00f3bicos isolados nos pacientes com DAC. 189Portanto, existem evid\u00eancias que sugerem a possibilidade da aplica\u00e7\u00e3o de treinos intervalados em pacientes com DAC est\u00e1vel, modalidade que tem se revelado segura e efetiva em melhorar o condicionamento f\u00edsico, a fun\u00e7\u00e3o endotelial e a fun\u00e7\u00e3o ventricular esquerda, acima dos resultados obtidos no treinamento moderado cont\u00ednuo. 6.3.4.3. Ajustes de F\u00e1rmacos Diante da Assimila\u00e7\u00e3o do Treinamento F\u00edsicoOs pacientes portadores de DAC est\u00e1vel geralmente utilizam medicamentos para al\u00edvio da sintomatologia, redu\u00e7\u00e3o de isquemia, melhora da fun\u00e7\u00e3o endotelial, estabiliza\u00e7\u00e3o da placa ateroscler\u00f3tica, controle dos fatores de risco e adequa\u00e7\u00e3o do padr\u00e3o hemodin\u00e2mico. Por exemplo, n\u00edveis elevados de PAS e/ou FC (aumento do duplo produto), aumentando o consumo de oxig\u00eanio mioc\u00e1rdico, obviamente contribuem para piorar a toler\u00e2ncia ao esfor\u00e7o e situa\u00e7\u00e3o cl\u00ednica.Nos programas de RCV, particular aten\u00e7\u00e3o deve ser dada \u00e0 melhora do limiar anginoso antes do in\u00edcio do treinamento, j\u00e1 que possibilita maior toler\u00e2ncia \u00e0 progress\u00e3o da intensidade de exerc\u00edcios e, com isso, a obten\u00e7\u00e3o dos efeitos ben\u00e9ficos almejados. Sendo assim, a otimiza\u00e7\u00e3o do tratamento farmacol\u00f3gico \u00e9 fundamental para uma RCV segura e eficaz.190 o que pode significar menor necessidade de f\u00e1rmacos usados no tratamento das DCV, sendo papel do m\u00e9dico de reabilita\u00e7\u00e3o discutir com o m\u00e9dico assistente sobre a eventual necessidade de ajustes farmacol\u00f3gicos.O paciente submetido \u00e0 RCV pode apresentar uma s\u00e9rie de adapta\u00e7\u00f5es fisiol\u00f3gicas relacionadas ao exerc\u00edcio, como, por exemplo, modula\u00e7\u00e3o favor\u00e1vel do sistema nervoso aut\u00f4nomo , com maior variabilidade de FC, redu\u00e7\u00e3o da FC basal, redu\u00e7\u00e3o do duplo produto de repouso e melhora da fun\u00e7\u00e3o endotelial, 191 deteriora\u00e7\u00e3o da vasorreatividade perif\u00e9rica com disfun\u00e7\u00e3o endotelial 192 e inflama\u00e7\u00e3o cr\u00f4nica. 193 Nesse contexto, o exerc\u00edcio f\u00edsico se estabeleceu como estrat\u00e9gia terap\u00eautica segura, que atenua os efeitos do descondicionamento f\u00edsico progressivo decorrente da evolu\u00e7\u00e3o natural da doen\u00e7a. 194A IC cr\u00f4nica \u00e9 uma s\u00edndrome complexa que compromete m\u00faltiplos sistemas, ocasionando como principais sintomas a dispneia e intoler\u00e2ncia progressiva ao esfor\u00e7o f\u00edsico. Apesar dos recentes avan\u00e7os na terap\u00eautica farmacol\u00f3gica, com redu\u00e7\u00e3o da elevada morbimortalidade, os sintomas tendem a persistir, comprometendo a qualidade de vida dos pacientes. Existem evid\u00eancias consistentes de que a redu\u00e7\u00e3o do n\u00edvel de atividade f\u00edsica na IC desencadeia um c\u00edrculo vicioso, que contribui para aumentar os sintomas e a intoler\u00e2ncia ao exerc\u00edcio, secund\u00e1rios \u00e0 redu\u00e7\u00e3o da capacidade funcional, produzindo efeitos psicol\u00f3gicos negativos, 197 No entanto, um \u00fanico e grande estudo randomizado multic\u00eantrico, o HF-ACTION, 198 revelou apenas uma modesta, mas n\u00e3o significativa, redu\u00e7\u00e3o nos desfechos prim\u00e1rios de morte e hospitaliza\u00e7\u00f5es por todas as causas, embora tenha demonstrado benef\u00edcios importantes na qualidade de vida e redu\u00e7\u00e3o da taxa de hospitaliza\u00e7\u00f5es por IC. Como cr\u00edtica \u00e0 pesquisa, h\u00e1 que se considerar que a baixa ades\u00e3o aos exerc\u00edcios provavelmente prejudicou a efic\u00e1cia da interven\u00e7\u00e3o, hip\u00f3tese que foi confirmada posteriormente em outro estudo, que demonstrou ser a ades\u00e3o aos exerc\u00edcios fator determinante para a obten\u00e7\u00e3o de benef\u00edcios a m\u00e9dio prazo. 199Estudos randomizados pequenos, revis\u00f5es sistem\u00e1ticas e meta-an\u00e1lises t\u00eam consistentemente demonstrado que o treinamento f\u00edsico regular \u00e9 seguro, aumenta a toler\u00e2ncia aos exerc\u00edcios, melhora a qualidade de vida e reduz hospitaliza\u00e7\u00f5es por IC. 2 sobre treinamento f\u00edsico em pacientes com IC, que analisou 33 estudos randomizados com inclus\u00e3o de 4.740 pacientes com predom\u00ednio FEVE reduzida, houve tend\u00eancia \u00e0 redu\u00e7\u00e3o da mortalidade total com os exerc\u00edcios f\u00edsicos ap\u00f3s um ano de seguimento. Comparado ao controle, o grupo de treinamento f\u00edsico teve menor taxa de hospitaliza\u00e7\u00e3o por IC e melhora da qualidade de vida. Quanto aos benef\u00edcios nas mulheres com IC, os estudos dispon\u00edveis sugerem que s\u00e3o positivos e equivalentes aos observados nos homens. 200Em uma revis\u00e3o sistem\u00e1tica New York Heart Association ), ainda n\u00e3o h\u00e1 dados suficientes para indicar programas de exerc\u00edcio, pois apenas um estudo randomizado brasileiro testou um programa de exerc\u00edcios di\u00e1rios em cicloerg\u00f4metro com ventila\u00e7\u00e3o n\u00e3o invasiva. Foram avaliados pacientes internados com IC descompensada, sendo observados benef\u00edcios funcionais e redu\u00e7\u00e3o do tempo de interna\u00e7\u00e3o. 201 Portanto, para um grau de recomenda\u00e7\u00e3o mais forte, h\u00e1 necessidade de mais estudos que confirmem os resultados iniciais.Para pacientes com sintomas avan\u00e7ados .Internacionalmente, os programas de RCV s\u00e3o implementados com v\u00e1rios formatos, utilizando-se modalidades isoladas ou associadas. Os exerc\u00edcios aplicados podem ser aer\u00f3bicos , de resist\u00eancia muscular localizada e treinamento de musculatura respirat\u00f3ria .208 Os testes funcionais devem ser realizados em uso das medica\u00e7\u00f5es prescritas para mimetizar a condi\u00e7\u00e3o que estar\u00e1 presente durante o treinamento.Antes de iniciar o programa de treinamento, \u00e9 fundamental que o paciente esteja com o tratamento farmacol\u00f3gico otimizado e clinicamente est\u00e1vel; o ideal \u00e9 que realize uma avalia\u00e7\u00e3o funcional, preferencialmente com TCPE ou TE. Na indisponibilidade das provas funcionais citadas, o teste de caminhada de 6 minutos pode servir de par\u00e2metro de acompanhamento dos ganhos funcionais. Os treinamentos aer\u00f3bicos recomendados podem ser cont\u00ednuos de moderada intensidade (TCMI), que correspondem \u00e0 zona de FC delimitada pelos limiares ventilat\u00f3rios do TCPE, ou, no caso do TE, \u00e0 zona situada entre 60 e 80% da FC pico ou 50 e 70% da FC de reserva. Pacientes mais graves e com maior limita\u00e7\u00e3o funcional podem iniciar no limite inferior da prescri\u00e7\u00e3o. Progress\u00f5es de intensidade at\u00e9 o limite superior podem ser realizadas com a evolu\u00e7\u00e3o do treinamento.Recentemente, tem aumentado a utiliza\u00e7\u00e3o de exerc\u00edcios aer\u00f3bicos de alta intensidade realizados de modo intervalado, denominado de treinamento intervalado de alta intensidade (TIAI). Ele alterna per\u00edodos mais intensos com momentos de recupera\u00e7\u00e3o passiva ou ativa, o que possibilita maior dura\u00e7\u00e3o total de exerc\u00edcios na alta intensidade e, consequentemente, pode produzir maior est\u00edmulo para adapta\u00e7\u00f5es fisiol\u00f3gicas centrais e perif\u00e9ricas.209 demonstraram que o TIAI foi superior ao TCMI em promover a melhora na capacidade funcional e em diferentes par\u00e2metros cardiovasculares. Posteriormente, outros ensaios cl\u00ednicos foram realizados e meta-analisados. No que tange ao efeito do TIAI sobre a capacidade funcional, a superioridade do m\u00e9todo em rela\u00e7\u00e3o ao TCMI foi confirmada em uma meta-an\u00e1lise. 210 Entretanto, o maior estudo multic\u00eantrico publicado, o Smartex-HF, 211 comparou as modalidades de exerc\u00edcios cont\u00ednuos de moderada intensidade com os intervalados de alta intensidade. A conclus\u00e3o foi que os benef\u00edcios s\u00e3o semelhantes, n\u00e3o havendo superioridade de modalidade em nenhum aspecto. Portanto, a escolha do protocolo vai depender de experi\u00eancia da equipe, condi\u00e7\u00f5es cl\u00ednicas, capacidade f\u00edsica e prefer\u00eancias do paciente.Em pacientes com IC e FEVE reduzida, Wisl\u00f8ff et al. 212 Um deles \u00e9 composto por 4 minutos de exerc\u00edcios de alta intensidade (90 a 95% da FC m\u00e1xima), alternados com 3 minutos de leve intensidade (70% da FC m\u00e1xima). 209 Protocolos com dura\u00e7\u00e3o da carga intensa bem menores, com 30 ou 90 s, j\u00e1 foram descritos, e a toler\u00e2ncia a diferentes modelos de TIAI pode variar de acordo com a escolha e a capacidade f\u00edsica do paciente. 213 Portanto, a utiliza\u00e7\u00e3o e o modo de execu\u00e7\u00e3o vai depender das caracter\u00edsticas cl\u00ednicas e escolhas do paciente, bem como da experi\u00eancia e das prefer\u00eancias da equipe de RCV.Al\u00e9m disso, o modo de utilizar o TIAI pode variar bastante, com v\u00e1rios protocolos descritos. 214 Eles podem ser prescritos como percentuais da contra\u00e7\u00e3o volunt\u00e1ria m\u00e1xima ou de acordo com a percep\u00e7\u00e3o subjetiva ao esfor\u00e7o. As cargas e repeti\u00e7\u00f5es recomendadas podem variar de acordo com as limita\u00e7\u00f5es funcionais do paciente e devem ser individualizadas, com progress\u00e3o de acordo com evolu\u00e7\u00e3o na RCV.Al\u00e9m do treinamento aer\u00f3bico, a adi\u00e7\u00e3o de exerc\u00edcios de resist\u00eancia muscular localizada tem sido sugerida para obten\u00e7\u00e3o de benef\u00edcios adicionais. 215 A meta-an\u00e1lise de Smart et al., 216 que avaliou 11 estudos com 287 participantes com IC, sendo 148 submetidos ao treinamento da musculatura inspirat\u00f3ria (TMI) comparados com 139 controles sedent\u00e1rios, mostrou significativos ganhos devidamente documentados: a) no TCPE, pelo aumento de consumo de oxig\u00eanio mioc\u00e1rdico no pico do esfor\u00e7o (VO 2 pico) e melhora da efici\u00eancia ventilat\u00f3ria observada na rela\u00e7\u00e3o da ventila\u00e7\u00e3o pulmonar com a produ\u00e7\u00e3o de di\u00f3xido de carbono (VE/VCO 2 slope); b) na espirometria, pelo aumento da press\u00e3o inspirat\u00f3ria m\u00e1xima; c) no teste de caminhada de 6-minutos, pela maior dist\u00e2ncia percorrida; d) melhora da qualidade de vida. Portanto, o TMI proporcionou ganhos da aptid\u00e3o cardiorrespirat\u00f3ria e na qualidade de vida de similar magnitude aos obtidos com o treinamento convencional, devendo ser considerado alternativa v\u00e1lida para os pacientes com IC gravemente descondicionados fisicamente e muito debilitados, em uma transi\u00e7\u00e3o para os exerc\u00edcios f\u00edsicos convencionais.Os exerc\u00edcios respirat\u00f3rios t\u00eam sido indicados para programas de treinamento de pacientes com fraqueza da musculatura respirat\u00f3ria. 217\u00c9 fundamental que pacientes com IC realizem exerc\u00edcios f\u00edsicos, idealmente com prescri\u00e7\u00e3o individualizada, no contexto de um programa de RCV, levando-se em considera\u00e7\u00e3o a combina\u00e7\u00e3o de treinamentos aer\u00f3bicos de moderada e/ou alta intensidade, exerc\u00edcios de resist\u00eancia muscular localizada e treinamento da musculatura respirat\u00f3ria (treinamento ventilat\u00f3rio). Para isso, devem ser levadas em considera\u00e7\u00e3o, al\u00e9m do quadro cl\u00ednico e limita\u00e7\u00f5es funcionais a ele relacionadas, prefer\u00eancias do paciente e a experi\u00eancia da equipe. Por fim, \u00e9 relevante ressaltar a exist\u00eancia de alternativas v\u00e1lidas mesmo para os pacientes muito debilitados e gravemente descondicionados. O transplante card\u00edaco (TxC) \u00e9 o tratamento de escolha para pacientes com IC refrat\u00e1ria, que permanecem com sintomas graves mesmo em uso de todo o arsenal farmacol\u00f3gico dispon\u00edvel e na realiza\u00e7\u00e3o de procedimentos cir\u00fargicos indicados.218Nos \u00faltimos anos ocorreram avan\u00e7os significativos no TxC, com surgimento de novas t\u00e9cnicas cir\u00fargicas e desenvolvimento de subst\u00e2ncias imunossupressoras mais eficientes. No Brasil, houve um crescimento substancial na quantidade de procedimentos, o que estava estagnado desde 2015, com taxa de 1,7 TxC por milh\u00e3o da popula\u00e7\u00e3o (pmp). Em 2019 houve um crescimento de 17,6%, chegando a 2 TxC pmp, muito pr\u00f3ximo da meta estabelecida para o ano . Em 2018 foram realizados 357 procedimentos, e at\u00e9 mar\u00e7o de 2019, 104 cora\u00e7\u00f5es j\u00e1 foram transplantados no Brasil. 220 Os receptores s\u00e3o capazes de retornar ao trabalho e ter uma vida normal, com m\u00ednimos sintomas ou mesmo assintom\u00e1ticos. 221 A taxa de sobrevida no 1\u00ba ano \u00e9 estimada em 90% e em 5 anos em cerca de 70%. 222O TxC tem como objetivo promover a melhora na qualidade de vida, assim como aumento da sobrevida. 2 pico ainda se encontra reduzido quando comparado ao de indiv\u00edduos saud\u00e1veis, pareados por idade. 224 Dentre outros fatores, isso pode ser explicado por: 1) imediatamente no per\u00edodo p\u00f3s-transplante, o aloenxerto apresenta aus\u00eancia de inerva\u00e7\u00e3o simp\u00e1tica e parassimp\u00e1tica (denerva\u00e7\u00e3o auton\u00f4mica), provocando aumento da FC de repouso, o que atenua a sua eleva\u00e7\u00e3o natural como resposta ao exerc\u00edcio e prejudica a recupera\u00e7\u00e3o ap\u00f3s o esfor\u00e7o 225 ; 2) ocorr\u00eancia de disfun\u00e7\u00e3o muscular esquel\u00e9tica (\u00e0s vezes chegando \u00e0 caquexia), na qual a terapia imunossupressora, associada \u00e0 IC pr\u00e9via, exerce papel de destaque 226 ; 3) comprometimento da fun\u00e7\u00e3o vascular e diast\u00f3lica. 227 Em pacientes com TxC, na fase aguda do exerc\u00edcio, o aumento do d\u00e9bito card\u00edaco depende fundamentalmente do mecanismo de Frank-Starling, do aumento do retorno venoso, do inotropismo, do cronotropismo e da redu\u00e7\u00e3o da p\u00f3s-carga. 229 Al\u00e9m disso, ocorre aumento das concentra\u00e7\u00f5es de catecolaminas circulantes, 227 que reduzem lentamente ap\u00f3s o t\u00e9rmino do exerc\u00edcio, justificando uma lenta recupera\u00e7\u00e3o da FC. 230Embora o TxC melhore significativamente a capacidade funcional dos pacientes, o VO 231 e os pacientes transplantados podem evoluir com desenvolvimento de HAS, diabetes melito e coronariopatia. 232 Por sua vez, o treinamento f\u00edsico \u00e9 conhecido como terap\u00eautica de excel\u00eancia para o manejo dessas doen\u00e7as cr\u00f4nicas, 233 sendo eficaz na otimiza\u00e7\u00e3o do controle auton\u00f4mico. 234A imunossupress\u00e3o pode predispor a maior risco de outras complica\u00e7\u00f5es, 2 pico e a melhora do controle hemodin\u00e2mico, da for\u00e7a muscular e da densidade mineral \u00f3ssea, 236 contribuindo para melhorar o progn\u00f3stico. 19 Embora existam in\u00fameras possibilidades de prescri\u00e7\u00e3o de treinamento, o principal m\u00e9todo preconizado permanece sendo o exerc\u00edcio aer\u00f3bico, que pode ser realizado de maneira cont\u00ednua ou intervalada, 170 sempre que poss\u00edvel devem ser realizados tamb\u00e9m exerc\u00edcios resistidos. 6O treinamento f\u00edsico ap\u00f3s o TxC contribui para o aumento do VO 237 os pesquisadores observaram que, ap\u00f3s um per\u00edodo de 46 meses p\u00f3s TxC, pacientes que realizaram treinamento aer\u00f3bico, apresentaram capacidade funcional e fun\u00e7\u00e3o cronotr\u00f3pica semelhantes \u00e0s verificadas em indiv\u00edduos saud\u00e1veis. 240No estudo pioneiro de Richard et al., 234 evidenciando aumento m\u00e9dio do VO 2 pico de 2,5 ml.kg - 1 .min - 1 nos que realizaram treinamento, em rela\u00e7\u00e3o aos alocados para cuidados usuais. Rosembaun et al. 241 estudaram a rela\u00e7\u00e3o entre a participa\u00e7\u00e3o precoce em um programa de RCV de pacientes ap\u00f3s TxC e verificaram que o n\u00famero de sess\u00f5es realizadas nos primeiros 90 dias esteve relacionado diretamente com melhor sobrevida em 10 anos.Uma meta-an\u00e1lise da Cochrane, que reuniu nove ensaios cl\u00ednicos randomizados, totalizando 284 pacientes, comparou o efeito do treinamento f\u00edsico aos cuidados usuais em pacientes p\u00f3s TxC, 242 descreveram melhoras significativas no VO 2 pico de pacientes ap\u00f3s TxC, com aumento m\u00e9dio de 3,1 ml.kg - 1 .min - 1 ap\u00f3s 12 semanas de treinamento combinado (resistido e aer\u00f3bico). Kobashigawa et al. 243 estudaram 27 pacientes ap\u00f3s TxC, os quais foram submetidos a uma combina\u00e7\u00e3o de treinamento aer\u00f3bico, resistido e de flexibilidade durante 6 meses versus grupo-controle. A dura\u00e7\u00e3o e a intensidade das sess\u00f5es de exerc\u00edcios aer\u00f3bicos tiveram como meta, no m\u00ednimo, 30 min de exerc\u00edcio cont\u00ednuo de intensidade moderada em bicicleta estacion\u00e1ria. O grupo interven\u00e7\u00e3o apresentou aumento m\u00e9dio de 4,4 ml - 1 .kg - 1 .min no VO 2 pico, versus 1,9 ml.kg - 1 .min - 1 no grupo-controle. Esses dados fornecem informa\u00e7\u00f5es valiosas da import\u00e2ncia de ambos os tipos de treinamento ap\u00f3s TxC.Haykowsky et al. crossover , Dall et al. 244 verificaram efeito superior do TIAI em rela\u00e7\u00e3o ao TCMI no VO 2 pico, com ganho adicional de 2,3 ml.kg - 1 .min - 1 e melhora superior na qualidade de vida. Em meta-an\u00e1lise que reuniu tr\u00eas ensaios cl\u00ednicos randomizados que compararam o TIAI (blocos intensos: 80 a 100% do VO 2 pico ou 85 a 95% da FC pico) aos cuidados usuais, os pacientes que realizaram TIAI apresentaram aumento adicional no VO 2 pico de 4,45 ml.kg - 1 .min - 1 ap\u00f3s o per\u00edodo de interven\u00e7\u00e3o, que variou de 8 a 12 semanas, com tr\u00eas a cinco sess\u00f5es semanais. 233Em rela\u00e7\u00e3o ao treinamento de alta intensidade em pacientes ap\u00f3s TxC, os resultados s\u00e3o motivadores, mas ainda h\u00e1 um pequeno n\u00famero de estudos. Em um estudo 224 avaliaram os efeitos de um programa de TIAI ap\u00f3s TxC em 43 pacientes. Foram observados os efeitos na progress\u00e3o da vasculopatia do enxerto, avaliada por ultrassom intracoron\u00e1rio, constatando menor progress\u00e3o de placas de ateroma no grupo TIAI. Por\u00e9m, mais estudos ainda s\u00e3o necess\u00e1rios para esclarecer melhor os benef\u00edcios dessa modalidade de treinamento. 245Nytr\u00f8en et al. 235 estudaram, pela primeira vez, o efeito do treinamento resistido na miopatia induzida por glicocorticoide em receptores de TxC. Um grupo realizou treinamento e foi comparado com um grupo-controle. Ap\u00f3s 6 meses, apesar de ambos terem apresentado aumento na for\u00e7a muscular do quadr\u00edceps e dos extensores lombares, houve um aumento at\u00e9 6 vezes maior no grupo treinado.Sabe-se que alguns dos efeitos adversos comuns ao uso de glicocorticoides ap\u00f3s o TxC s\u00e3o atrofia e fraqueza musculares. Em 1998, Braith et al. 236O treinamento resistido tamb\u00e9m parece ter importante efeito terap\u00eautico para melhoria do metabolismo \u00f3sseo. Ap\u00f3s o TxC, os pacientes n\u00e3o raramente apresentam perda \u00f3ssea significativa na cabe\u00e7a do f\u00eamur e perda \u00f3ssea mineral total. Em pacientes arrolados para treinamento resistido, ap\u00f3s 2 meses da realiza\u00e7\u00e3o do TxC, o treinamento de for\u00e7a se mostrou capaz de restaurar a densidade mineral \u00f3ssea a n\u00edveis pr\u00e9-transplante. 246 Os fisioterapeutas e profissionais de educa\u00e7\u00e3o f\u00edsica devem atuar na prescri\u00e7\u00e3o, aplica\u00e7\u00e3o, supervis\u00e3o e orienta\u00e7\u00e3o dos exerc\u00edcios, mas seguindo os limites de seguran\u00e7a recomendados pelos m\u00e9dicos respons\u00e1veis pela avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o. 247Os pacientes ap\u00f3s TxC devem realizar anamnese, exame f\u00edsico, ECG de repouso de 12 deriva\u00e7\u00f5es, ecocardiograma com Doppler colorido ou outros exames a crit\u00e9rio dos profissionais envolvidos. O ideal \u00e9 a realiza\u00e7\u00e3o de um teste funcional em exerc\u00edcio, preferencialmente o TCPE realizado por m\u00e9dico experiente com o m\u00e9todo. O TCPE considerado o padr\u00e3o-ouro para avalia\u00e7\u00e3o da capacidade funcional, permite a determina\u00e7\u00e3o das respostas cardiopulmonar e metab\u00f3licas ao esfor\u00e7o f\u00edsico, por meio de diversas vari\u00e1veis que s\u00e3o de grande utilidade para a avalia\u00e7\u00e3o cl\u00ednica e prescri\u00e7\u00e3o otimizada dos exerc\u00edcios f\u00edsicos. 170 Quando nem mesmo este estiver dispon\u00edvel, o teste de caminhada de 6 minutos poder\u00e1 auxiliar na avalia\u00e7\u00e3o cl\u00ednica e prescri\u00e7\u00e3o de exerc\u00edcio, sendo par\u00e2metro v\u00e1lido de compara\u00e7\u00e3o da capacidade funcional no decorrer do treinamento. 249A impossibilidade da realiza\u00e7\u00e3o do TCPE n\u00e3o deve ser um impedimento para pr\u00e1tica dos exerc\u00edcios; na aus\u00eancia dele, sugere-se um TE convencional. 170O treinamento aer\u00f3bico, ressaltando que o TCMI tem sido o m\u00e9todo usado na maioria dos estudos, tem sido o mais preconizado, devendo ser complementado pelo treinamento resistido a partir da 6\u00aa semana ap\u00f3s TxC. Mas, diferentes metodologias de treinamento v\u00eam sendo estudadas de maneira isolada ou combinada e t\u00eam se mostrado eficazes na promo\u00e7\u00e3o da sa\u00fade cardiovascular nos indiv\u00edduos em RCV, abrindo um leque de possibilidades a serem consideradas. 250 Nesse sentido, programas que inclu\u00edram o treinamento intervalado, at\u00e9 mesmo de alta intensidade, mostraram bons resultados. 233 Por\u00e9m, para uma otimizada e segura prescri\u00e7\u00e3o, deve haver adequada individualiza\u00e7\u00e3o de cada componente da sess\u00e3o de treinamento. 170De acordo com a condi\u00e7\u00e3o cl\u00ednica do paciente, a intensidade do exerc\u00edcio aer\u00f3bico pode aumentar gradualmente de moderada a alta ao longo do treinamento, pois a intensidade est\u00e1 diretamente associada \u00e0 magnitude das desej\u00e1veis adapta\u00e7\u00f5es cardiovasculares. 170 Entretanto, tendo em vista a resposta cronotr\u00f3pica ainda comprometida, 251 a prescri\u00e7\u00e3o com base nos percentuais da FC m\u00e1xima ou nas FC dos limiares n\u00e3o \u00e9 poss\u00edvel nas primeiras sess\u00f5es de treinamento, podendo ser \u00fateis quando ocorre melhora na resposta auton\u00f4mica. 224 Por esse motivo, a cont\u00ednua avalia\u00e7\u00e3o do comportamento da FC durante o exerc\u00edcio e na recupera\u00e7\u00e3o se torna de suma import\u00e2ncia. Quando um TCPE for dispon\u00edvel, a prescri\u00e7\u00e3o do exerc\u00edcio aer\u00f3bico poder\u00e1 ser pautada nas cargas atingidas nos limiares ventilat\u00f3rios ou nos percentuais estabelecidos do VO 2 pico. Outra estrat\u00e9gia simples e vi\u00e1vel \u00e9 a avalia\u00e7\u00e3o da percep\u00e7\u00e3o subjetiva do esfor\u00e7o, por meio da escala BORG, 252 devendo haver empenho da equipe multiprofissional em educar o paciente em rela\u00e7\u00e3o \u00e0 percep\u00e7\u00e3o de esfor\u00e7o e \u00e0s manifesta\u00e7\u00f5es de sintomas. 6A determina\u00e7\u00e3o de zonas-alvo de treinamento \u00e9 recomend\u00e1vel, visando otimiza\u00e7\u00e3o da prescri\u00e7\u00e3o do exerc\u00edcio. 253 que foi validado em idosos ativos e se mostrou razoavelmente confi\u00e1vel em fornecer informa\u00e7\u00f5es sobre a for\u00e7a de MMII, sendo bastante utilizado em centros de reabilita\u00e7\u00e3o e em estudos cient\u00edficos de diferentes condi\u00e7\u00f5es cl\u00ednicas. 256Al\u00e9m da avalia\u00e7\u00e3o e prescri\u00e7\u00e3o dos exerc\u00edcios aer\u00f3bicos, \u00e9 fundamental a realiza\u00e7\u00e3o de exerc\u00edcios resistidos. Tradicionalmente t\u00eam sido utilizados, para a avalia\u00e7\u00e3o e prescri\u00e7\u00e3o destes exerc\u00edcios, os testes de carga de uma repeti\u00e7\u00e3o m\u00e1xima, cuja aplica\u00e7\u00e3o, entretanto, \u00e9 question\u00e1vel, principalmente ap\u00f3s o TxC recente, pois sua seguran\u00e7a carece de investiga\u00e7\u00f5es cl\u00ednicas nos transplantados. Uma alternativa seria o teste de sentar e levantar da cadeira em 30 segundos, Uma possibilidade de grande aplicabilidade \u00e9 a prescri\u00e7\u00e3o dos exerc\u00edcios f\u00edsicos resistidos de modo subjetivo, segundo a percep\u00e7\u00e3o de esfor\u00e7o considerado moderado, associado ao m\u00e9todo de repeti\u00e7\u00e3o vari\u00e1vel, com o objetivo de executar uma faixa de repeti\u00e7\u00f5es . Se o paciente n\u00e3o conseguir executar o m\u00ednimo, a carga aplicada est\u00e1 elevada; se executar o m\u00e1ximo de modo f\u00e1cil, a carga est\u00e1 leve. Desse modo, a carga pode ser ajustada para que o treinamento seja realizado dentro da faixa de repeti\u00e7\u00f5es proposta.258 o que deixa evidente a import\u00e2ncia da supervis\u00e3o dos pacientes ao longo do treinamento, com eventual necessidade de que as sess\u00f5es sejam temporariamente interrompidas. Tendo em vista o exposto, alguns autores sugerem que os pacientes n\u00e3o devam realizar exerc\u00edcio f\u00edsico durante o per\u00edodo de administra\u00e7\u00e3o de terapia com pulsos de esteroides e nos dias de bi\u00f3psia mioc\u00e1rdica. 170Durante o treinamento, especial aten\u00e7\u00e3o deve ser dada \u00e0s complica\u00e7\u00f5es como as infec\u00e7\u00f5es relacionadas ao procedimento do transplante. Em levantamento realizado nos Estados Unidos, foi evidenciado que 36% dos receptores s\u00e3o hospitalizados ao longo do primeiro ano, e 61%, dentro de um per\u00edodo de 4 anos ap\u00f3s TxC, 1 sendo recomend\u00e1veis como uma alternativa para a RCV tradicional em pacientes de menor risco. 71Estudos t\u00eam demonstrado que os programas de RCVD s\u00e3o seguros e efetivos, 259 conduziram um estudo prospectivo e randomizado para avaliar o efeito de um programa de exerc\u00edcios domiciliares durante 2 meses em 37 pacientes ap\u00f3s TxC. O grupo controle manteve o estilo de vida habitual durante o per\u00edodo de estudo. Os indiv\u00edduos do grupo interven\u00e7\u00e3o realizaram um programa de exerc\u00edcios no m\u00ednimo tr\u00eas vezes na semana, que englobou 5 min de aquecimento, atividades de fortalecimento de MMSS e MMII, 15 a 20 min de exerc\u00edcio aer\u00f3bico em intensidade de 60 a 70% do VO 2 pico, al\u00e9m de 5 min de desaquecimento. Para garantir a execu\u00e7\u00e3o domiciliar correta, inicialmente foi realizado um per\u00edodo supervisionado para orienta\u00e7\u00e3o e prescri\u00e7\u00e3o. Ao final de 2 meses, os pacientes melhoraram for\u00e7a e resist\u00eancia muscular, \u00edndice de fadiga e qualidade de vida no dom\u00ednio f\u00edsico. Por meio do TCPE, foi observado aumento da carga de trabalho, mas sem modifica\u00e7\u00e3o do VO 2 pico, provavelmente pelo curto per\u00edodo de seguimento ou pela metodologia da prescri\u00e7\u00e3o do treinamento, que foi de menor intensidade. Outro estudo, 260 com protocolo de treinamento aer\u00f3bico equivalente, por\u00e9m maior dura\u00e7\u00e3o, cinco vezes por semana durante 6 meses, documentou melhora no VO 2 pico, na carga de trabalho e na PA de indiv\u00edduos ap\u00f3s TxC. Al\u00e9m disso, ocorreram sinais de reinerva\u00e7\u00e3o simp\u00e1tica card\u00edaca e restaura\u00e7\u00e3o da sensibilidade \u00e0 modula\u00e7\u00e3o auton\u00f4mica nas art\u00e9rias, sendo que nenhuma altera\u00e7\u00e3o foi observada no grupo controle.Wu et al. 2 pico. 261Mesmo com per\u00edodo superior a 5 anos ap\u00f3s o TxC, a RCVD melhora a capacidade funcional, conforme demonstra um estudo em que 21 pacientes foram instru\u00eddos a realizar um programa de treinamento f\u00edsico domiciliar por 1 ano em bicicleta ergom\u00e9trica, enquanto nove pacientes serviram como controle. Para garantir o adequado controle, os pacientes receberam um cart\u00e3o inteligente, programado para um aquecimento de 6 min e uma carga de trabalho constante durante 20 min, com ajuste de carga de acordo com a prescri\u00e7\u00e3o e o monitoramento da FC. Ao final de 12 meses, houve modesta melhora no VO 262 em estudo publicado em 2008, compararam os efeitos de programa domiciliar e presencial sobre a capacidade de exerc\u00edcio e vari\u00e1veis cronotr\u00f3picas em 28 pacientes ap\u00f3s TxC. Foram observadas melhoras significativas no VO 2 pico e na FC de reserva apenas no grupo da RCV tradicional. Por\u00e9m, novos estudos, com inclus\u00e3o de um maior n\u00famero de pacientes, s\u00e3o necess\u00e1rios para melhor elucida\u00e7\u00e3o desta superioridade da RCV presencial observada neste estudo.Karapolat et al. Com base nas diversas evid\u00eancias expostas, o efeito ben\u00e9fico do treinamento f\u00edsico em indiv\u00edduos ap\u00f3s TxC \u00e9 inequ\u00edvoco e essa terapia se mostra segura e exequ\u00edvel, podendo ser realizada no ambiente hospitalar ou domiciliar . No entA RCV deve ser iniciada entre 6 e 8 semanas ap\u00f3s o TxC, sendo o direcionamento realizado na alta hospitalar. Em casos selecionados e ap\u00f3s criteriosa avalia\u00e7\u00e3o da equipe, o in\u00edcio pode ser mais precoce. Assim como em qualquer situa\u00e7\u00e3o na qual o paciente seja submetido a esternotomia, um cuidado especial em rela\u00e7\u00e3o a n\u00e3o realizar exerc\u00edcios que sobrecarreguem a musculatura tor\u00e1cica e levem \u00e0 tra\u00e7\u00e3o do esterno deve ser salientado, principalmente nos primeiros 90 dias ap\u00f3s o procedimento cir\u00fargico.A prescri\u00e7\u00e3o ideal inclui exerc\u00edcios para promo\u00e7\u00e3o das diferentes val\u00eancias f\u00edsicas, sempre enfatizando o que \u00e9 preconizado para cada condi\u00e7\u00e3o. No cen\u00e1rio ap\u00f3s Txc, assim como em outras indica\u00e7\u00f5es de RCV, o exerc\u00edcio aer\u00f3bico \u00e9 a parte principal das sess\u00f5es de treinamento, devendo ser complementado pelos resistidos e de flexibilidade, dentro de um programa individualizado e periodizado. As sess\u00f5es devem sempre iniciar com um per\u00edodo de aquecimento, assim como encerrar com um desaquecimento controlado. Tal estrat\u00e9gia visa, al\u00e9m do aquecimento muscular, um per\u00edodo para ajuste da FC e da PA, que est\u00e3o alteradas nesses pacientes pela denerva\u00e7\u00e3o do cora\u00e7\u00e3o, especialmente no in\u00edcio do programa de treinamento ap\u00f3s o procedimento.indoor , utilizando recursos como esteiras e/ou bicicletas ergom\u00e9tricas, ou quanto outdoor . Recomenda-se frequ\u00eancia semanal de tr\u00eas a cinco sess\u00f5es, com dura\u00e7\u00e3o de 20 a 40 minutos. A frequ\u00eancia e dura\u00e7\u00e3o das sess\u00f5es ser\u00e3o ajustadas conforme condi\u00e7\u00f5es pr\u00e9vias do paciente e devem progredir ao longo do treinamento. O controle da intensidade \u00e9 fundamental e, devido ao maior n\u00famero de evid\u00eancias, preconiza-se o TCMI (Entre o 1\u00ba e o 2\u00ba limiar ventilat\u00f3rio), com uma percep\u00e7\u00e3o de esfor\u00e7o referida entre 11 e 13 na Escala Borg modificada. O treinamento intervalado pode ser adotado, em casos selecionados, com objetivo de varia\u00e7\u00e3o na forma do treino e busca de um ganho funcional potencialmente maior.O exerc\u00edcio aer\u00f3bico pode ser realizado em forma de caminhada ou ciclismo, tanto Os exerc\u00edcios resistidos t\u00eam papel fundamental, principalmente na fase inicial ap\u00f3s TxC. Muitos apresentaram IC de longa dura\u00e7\u00e3o, estiveram internados por longos per\u00edodos e passaram pelo estresse cir\u00fargico. No in\u00edcio do treinamento, atividades sem carga externa, ou seja, apenas com peso corporal, podem ser consideradas como est\u00edmulo suficiente para esses pacientes. Em seguida, bandas el\u00e1sticas, halteres, caneleiras e aparelhos de muscula\u00e7\u00e3o podem ser inclu\u00eddos no programa de treinamento. Maior cuidado deve ser dado aos exerc\u00edcios de MMSS, devido \u00e0 toracotomia, levando em considera\u00e7\u00e3o que, com o uso de corticosteroides, o per\u00edodo de cicatriza\u00e7\u00e3o pode ser maior.265Outras informa\u00e7\u00f5es e exemplos de protocolos de treinamento nesses pacientes podem ser obtidos em outras publica\u00e7\u00f5es. Nesta se\u00e7\u00e3o, ser\u00e3o abordadas a miocardiopatia hipertr\u00f3fica (MCH), a miocardite e outras miocardiopatias, cujas indica\u00e7\u00f5es de RCV est\u00e3o listadas na 271 sendo a doen\u00e7a card\u00edaca herdada mais comum na popula\u00e7\u00e3o e causada por uma gama de muta\u00e7\u00f5es de genes respons\u00e1veis pelas prote\u00ednas do sarc\u00f4mero card\u00edaco. 268 A principal caracter\u00edstica \u00e9 uma express\u00e3o cl\u00ednica heterog\u00eanea, com altera\u00e7\u00f5es fisiopatol\u00f3gicas peculiares e uma hist\u00f3ria natural vari\u00e1vel. At\u00e9 10% dos casos s\u00e3o causados por outras doen\u00e7as gen\u00e9ticas, incluindo metab\u00f3licas e neuromusculares heredit\u00e1rias, anormalidades cromoss\u00f4micas e s\u00edndromes gen\u00e9ticas. 272 Alguns pacientes apresentam outros dist\u00farbios que podem mimetizar formas da doen\u00e7a, como, por exemplo, amiloidose. 273A MCH \u00e9 uma doen\u00e7a caracterizada por hipertrofia do ventr\u00edculo esquerdo, geralmente com c\u00e2maras ventriculares n\u00e3o dilatadas, na aus\u00eancia de outra doen\u00e7a card\u00edaca ou sist\u00eamica capaz de produzir a magnitude da hipertrofia evidenciada, 268 No entanto, essa estimativa parece ser distinta na pr\u00e1tica cl\u00ednica, o que permite inferir que uma parcela dos indiv\u00edduos afetados s\u00e3o assintom\u00e1ticos. Diversos padr\u00f5es de hipertrofia assim\u00e9trica do ventr\u00edculo esquerdo s\u00e3o comuns \u00e0 MCH, e pode haver fen\u00f3tipos diversos em familiares de primeiro grau. Tipicamente, uma ou mais regi\u00f5es do ventr\u00edculo esquerdo t\u00eam espessura parietal aumentada quando comparadas com outras, e podem ocorrer transi\u00e7\u00f5es e varia\u00e7\u00f5es de espessura em \u00e1reas adjacentes ou \u00e1reas n\u00e3o cont\u00edguas. Contudo, apesar da hipertrofia septal assim\u00e9trica ser a mais comumente debatida, n\u00e3o existe um padr\u00e3o cl\u00e1ssico de MCH e, virtualmente, todos os padr\u00f5es poss\u00edveis de hipertrofia ventricular esquerda podem ocorrer. Mesmo a aus\u00eancia de hipertrofia pode ser encontrada em indiv\u00edduos geneticamente acometidos (fen\u00f3tipo negativo).A preval\u00eancia populacional \u00e9 estimada em torno de 0,2% ou 1:500. 274 Notavelmente, apenas em um pequeno subgrupo de pacientes com MCH ocorre morte prematura e complica\u00e7\u00f5es significativas relacionadas \u00e0 doen\u00e7a, as quais podem ocorrer por obstru\u00e7\u00e3o da via de sa\u00edda do ventr\u00edculo esquerdo, IC com disfun\u00e7\u00e3o diast\u00f3lica e/ou sist\u00f3lica e morte s\u00fabita (MS) ou arritmias card\u00edacas . 275 A MS na MCH pode acontecer em qualquer faixa et\u00e1ria, embora seja mais comum em adolescentes e adultos jovens; por isso, a identifica\u00e7\u00e3o de indiv\u00edduos sob maior risco \u00e9 muito importante na avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o esportiva. 276Diferentes estudos de coorte retrospectivos e observacionais, de popula\u00e7\u00f5es multic\u00eantricas, esclareceram a hist\u00f3ria natural e o curso cl\u00ednico dessa cardiopatia. Alguns mais recentes t\u00eam mostrado mortalidade anual em torno de 1%, valor muito menor do que em pesquisas mais antigas. 279 Portanto, incentivar um estilo de vida saud\u00e1vel para pacientes com MCH \u00e9 essencial para reduzir o risco global de doen\u00e7a.Em muitos casos, a MS pode ser a primeira manifesta\u00e7\u00e3o da doen\u00e7a nesses indiv\u00edduos, ocorrendo mais comumente naqueles sem sintomas de alerta, os quais n\u00e3o haviam sido diagnosticados previamente ao evento. Entretanto, a maioria dos pacientes com MCH apresenta uma expectativa de vida normal ou quase normal, com mortalidade relacionada a outras doen\u00e7as, algumas, inclusive, de etiologia n\u00e3o cardiovascular. 6.6.1.1. Benef\u00edcios Terap\u00eauticos do Exerc\u00edcio F\u00edsico19 Em paciente com MCH, forma obstrutiva e minimamente sintom\u00e1tica, tamb\u00e9m j\u00e1 foi observada associa\u00e7\u00e3o de mortalidade com a aptid\u00e3o aer\u00f3bica. 281 Pacientes com VO 2 pico abaixo de 18 ml.kg - 1 .min - 1 no TCPE apresentaram maior mortalidade e sintomatologia mais exuberante em compara\u00e7\u00e3o aos que obtiveram valores iguais ou superiores. A presen\u00e7a de VO 2 pico inferior a 60% do previsto significou pior sobrevida em 4 anos, em torno de 60%. 280O n\u00edvel de aptid\u00e3o cardiorrespirat\u00f3ria est\u00e1 associado ao risco de mortalidade cardiovascular e por todas as causas na popula\u00e7\u00e3o em geral. 271 Evid\u00eancias sugerem que o treinamento f\u00edsico intenso poderia acelerar tais altera\u00e7\u00f5es, mas esse ainda \u00e9 um tema controverso. Entretanto, sabe-se que o aumento da fibrose mioc\u00e1rdica est\u00e1 associado a menor VO 2 pico nessa popula\u00e7\u00e3o. 282O aumento da fibrose mioc\u00e1rdica e o desarranjo miofibrilar podem estar por tr\u00e1s do risco aumentado de MS na MCH, pois essa altera\u00e7\u00e3o atua como substrato para arritmias fatais. 281 Quando existe redu\u00e7\u00e3o do VO 2 pico, a pr\u00e1tica de exerc\u00edcios f\u00edsicos pode contribuir para aumentar a capacidade funcional.Sendo assim, avaliar a aptid\u00e3o aer\u00f3bica, preferencialmente pelo TCPE, \u00e9 importante nos pacientes com MCH. 2 pico ap\u00f3s treinamento de moderada intensidade depois de 16 semanas de interven\u00e7\u00e3o . 266 Outro estudo prospectivo n\u00e3o randomizado incluiu 20 pacientes com MCH e mostrou aumento significativo na dura\u00e7\u00e3o do TE, assim como na capacidade funcional estimada . 267 Nesse estudo, os pacientes completaram um programa de RCV com sess\u00f5es de 60 min de exerc\u00edcio moderado a vigoroso, realizados em esteira ou cicloerg\u00f4metro, 2 vezes por semana. A intensidade do exerc\u00edcio progrediu de 50 para 85% da FC de reserva, o que resultou em aumento gradual do condicionamento e pode ter minimizado o risco de eventos adversos, como arritmias induzidas pelo exerc\u00edcio. Em nenhum desses estudos com treinamento houve ocorr\u00eancia de eventos adversos s\u00e9rios, como morte, MS card\u00edaca abortada, terapia do cardioversor desfibrilador implant\u00e1vel (CDI) ou taquicardia ventricular sustentada. 267At\u00e9 o momento, apenas um ensaio cl\u00ednico randomizado controlado examinou o efeito do treinamento f\u00edsico em pacientes com MCH (RESET-MCH). Esse estudo, que contemplou 136 pacientes, demonstrou aumento no VO 6.6.1.2. Quando indicar exerc\u00edcios f\u00edsicos281A intensidade de exerc\u00edcio liberado para pacientes com MCH representa um grande desafio. Se por um lado, o exerc\u00edcio f\u00edsico intenso pode ser delet\u00e9rio, com aumento do risco de arritmias potencialmente fatais; por outro, a restri\u00e7\u00e3o excessiva de atividade f\u00edsica conduz ao descondicionamento e pode ter efeitos negativos na sa\u00fade e na qualidade de vida do paciente, podendo at\u00e9 aumentar o risco cardiovascular, visto que existe associa\u00e7\u00e3o entre aptid\u00e3o f\u00edsica e mortalidade. American Heart Association , em seu posicionamento oficial para o tratamento da MCH, desencoraja os pacientes com a doen\u00e7a a se envolverem em esportes competitivos de intensidade moderada a vigorosa , pois ambos poderiam aumentar o risco cardiovascular.Em virtude das novas evid\u00eancias, pode-se considerar haver efeito positivo dos exerc\u00edcios f\u00edsicos moderados em pacientes selecionados com MCH, com a avalia\u00e7\u00e3o de risco e a prescri\u00e7\u00e3o realizadas individualmente. Ressalta-se que as evid\u00eancias sugerem benef\u00edcios para o treinamento moderado cont\u00ednuo, sendo que outras modalidades ainda devem ser mais estudadas.Entretanto, a presen\u00e7a das seguintes caracter\u00edsticas poderiam ser consideradas contraindica\u00e7\u00f5es maiores \u00e0 pr\u00e1tica de exerc\u00edcio: hist\u00f3ria de MS abortada e aus\u00eancia de CDI; hist\u00f3ria de s\u00edncope aos esfor\u00e7os; ocorr\u00eancia de taquicardia ventricular induzida pelo exerc\u00edcio; aumento do gradiente com o exerc\u00edcio (superior a 50 mmHg) e resposta press\u00f3rica anormal ao esfor\u00e7o.6.6.1.3. Avalia\u00e7\u00e3o Pr\u00e9-participa\u00e7\u00e3oA defini\u00e7\u00e3o da libera\u00e7\u00e3o para iniciar os exerc\u00edcios deve ser realizada pela avalia\u00e7\u00e3o m\u00e9dica pr\u00e9-participa\u00e7\u00e3o, realizando-se anamnese, exame f\u00edsico e ECG de 12 deriva\u00e7\u00f5es.285 As altera\u00e7\u00f5es eletrocardiogr\u00e1ficas podem preceder doen\u00e7a estruturalmente detectada por alguns anos, o que torna a realiza\u00e7\u00e3o do ECG de suma import\u00e2ncia nesse cen\u00e1rio. 269 Somente uma minoria dos pacientes com MCH apresentam ECG normal, em geral aqueles sem outra manifesta\u00e7\u00e3o fenot\u00edpica da doen\u00e7a (gen\u00f3tipo positivo/fen\u00f3tipo negativo).Uma grande parcela dos indiv\u00edduos com MCH \u00e9 assintom\u00e1tica ou oligossintom\u00e1tica e a suspeita cl\u00ednica \u00e9 dada por altera\u00e7\u00f5es no ECG de repouso, o qual \u00e9 anormal em at\u00e9 95% dos pacientes com a doen\u00e7a. 286O ecocardiograma segue como o exame mais empregado para o diagn\u00f3stico de MCH, restando a resson\u00e2ncia magn\u00e9tica nuclear (RMN) como alternativa para os casos em que o primeiro n\u00e3o for conclusivo, ou para avaliar situa\u00e7\u00f5es de hipertrofia mais localizada . Em atletas jovens, a diferencia\u00e7\u00e3o entre a hipertrofia fisiol\u00f3gica do cora\u00e7\u00e3o de atleta e a hipertrofia patol\u00f3gica da MCH \u00e9 um desafio. Isso porque, tal como acontece com indiv\u00edduos sedent\u00e1rios com MCH, a maioria dos atletas com a doen\u00e7a mostra um padr\u00e3o assim\u00e9trico de hipertrofia do ventr\u00edculo esquerdo. Em contraste, aqueles com hipertrofia fisiol\u00f3gica do ventr\u00edculo esquerdo mostram distribui\u00e7\u00e3o mais homog\u00eanea e sim\u00e9trica da espessura da parede, com apenas pequenas diferen\u00e7as entre os segmentos cont\u00edguos e um padr\u00e3o sim\u00e9trico de hipertrofia do ventr\u00edculo esquerdo. 287O teste de exerc\u00edcio, previamente ao in\u00edcio da RCV, est\u00e1 sempre recomendado nesses pacientes, seja para avalia\u00e7\u00e3o da capacidade funcional ou para detec\u00e7\u00e3o de respostas anormais da PA e sinais de aumento da obstru\u00e7\u00e3o din\u00e2mica da via de sa\u00edda com o esfor\u00e7o. Para melhor detec\u00e7\u00e3o de obstru\u00e7\u00e3o da via de sa\u00edda durante o exerc\u00edcio, a associa\u00e7\u00e3o de exame de imagem (ecocardiograma) com teste de esfor\u00e7o \u00e9 a melhor modalidade dispon\u00edvel e deve ser encorajada sempre que poss\u00edvel. Pacientes com aus\u00eancia de obstru\u00e7\u00e3o no repouso podem apresentar gradientes significativos no esfor\u00e7o, sendo reclassificados em rela\u00e7\u00e3o ao progn\u00f3stico. 2 pico, em virtude do seu documentado valor progn\u00f3stico. 281 Al\u00e9m disso, a obten\u00e7\u00e3o dos limiares ventilat\u00f3rios contribui para uma prescri\u00e7\u00e3o mais individualizada.Quando estiver dispon\u00edvel, sugere-se a realiza\u00e7\u00e3o do TCPE para uma melhor avalia\u00e7\u00e3o em esfor\u00e7o, com medida direta do VO 6.6.1.4. Particularidades na Prescri\u00e7\u00e3o e no Acompanhamento dos Exerc\u00edcios F\u00edsicosAlgumas particularidades dos exerc\u00edcios em pacientes com MCH devem ser destacadas:Atividades do tipo \u201cexplos\u00e3o\u201d , em que h\u00e1 potencial para r\u00e1pida acelera\u00e7\u00e3o e desacelera\u00e7\u00e3o, devem ser evitadas;Atividades com consumo de energia est\u00e1vel e constante s\u00e3o preferidas;Exerc\u00edcio em condi\u00e7\u00f5es ambientais adversas, incluindo calor ou frio extremos, deve ser evitado, pois h\u00e1 um aumento do risco de exacerba\u00e7\u00e3o das altera\u00e7\u00f5es fisiol\u00f3gicas induzidas pelo exerc\u00edcio;Programas de treinamento que visem competitividade, ou obten\u00e7\u00e3o de n\u00edveis mais altos de condicionamento f\u00edsico e excel\u00eancia, devem ser evitados, pois normalmente motivam os pacientes a se esfor\u00e7arem al\u00e9m dos limites.Exerc\u00edcios est\u00e1ticos (isom\u00e9tricos) intensos, como levantamento de peso, devem ser evitados, pois h\u00e1 risco aumentado de provocar obstru\u00e7\u00e3o da via de sa\u00edda do ventr\u00edculo esquerdo, devido \u00e0 intensa manobra de Valsalva;Treinamento resistido com baixa carga e maior n\u00famero de repeti\u00e7\u00f5es foi considerado seguro para pacientes com DCV, embora n\u00e3o haja evid\u00eancia s\u00f3lida do seu uso na MCH.Em rela\u00e7\u00e3o ao uso de medica\u00e7\u00f5es, algumas observa\u00e7\u00f5es s\u00e3o importantes. O uso de betabloqueadores e antagonistas do c\u00e1lcio pode estar indicado no tratamento da MCH. Como essas medica\u00e7\u00f5es atenuam a resposta da FC, \u00e9 poss\u00edvel ocorrer uma resposta cronotr\u00f3pica muito reduzida ao esfor\u00e7o, o que pode ocasionar aumento da intoler\u00e2ncia ao exerc\u00edcio e indicar necessidade de ajuste da medica\u00e7\u00e3o. O uso de diur\u00e9ticos em excesso pode ser delet\u00e9rio por aumentar o gradiente da via de sa\u00edda. Sendo assim, devem ser utilizados com cautela. Do mesmo modo que os diur\u00e9ticos, a desidrata\u00e7\u00e3o pelo exerc\u00edcio pode elevar o gradiente. Assim, \u00e9 importante aten\u00e7\u00e3o a uma adequada hidrata\u00e7\u00e3o durante o treinamento.A patog\u00eanese da miocardite consiste em tr\u00eas fases: les\u00e3o aguda, geralmente de etiologia viral; resposta imune do hospedeiro; e recupera\u00e7\u00e3o, ou transi\u00e7\u00e3o para fibrose e miocardiopatia dilatada, sendo que, clinicamente, n\u00e3o existe uma distin\u00e7\u00e3o clara entre essas fases. A les\u00e3o inicial pode causar dano agudo ao mioc\u00e1rdio, com comprometimento contr\u00e1til mediado por citocinas produzidas pelo processo inflamat\u00f3rio local. Esse quadro inflamat\u00f3rio agudo pode evoluir, na fase tardia, para fibrose extensa, o que pode causar dilata\u00e7\u00e3o e disfun\u00e7\u00e3o ventricular.283A miocardite aguda \u00e9 suspeitada quando existe a presen\u00e7a dos seguintes crit\u00e9rios: S\u00edndrome cl\u00ednica com IC aguda, dor tor\u00e1cica do tipo angina ou miopericardite com menos de 3 meses de dura\u00e7\u00e3o;Eleva\u00e7\u00e3o inexplicada na dosagem de troponina s\u00e9rica;Altera\u00e7\u00f5es eletrocardiogr\u00e1ficas sugestivas de isquemia mioc\u00e1rdica;Anormalidades contr\u00e1teis globais ou segmentares e/ou derrame peric\u00e1rdico na ecocardiografia.RMN com altera\u00e7\u00f5es caracter\u00edsticas no sinal tecidual em T2, ou imagens ponderadas em T1, e presen\u00e7a de realce tardio com gadol\u00ednio.290Em rela\u00e7\u00e3o \u00e0 participa\u00e7\u00e3o em programas de RCV de pacientes com miocardites ap\u00f3s a resolu\u00e7\u00e3o da fase aguda, o assunto \u00e9 muito pouco estudado e n\u00e3o existem estudos cient\u00edficos sobre a seguran\u00e7a e efic\u00e1cia da interven\u00e7\u00e3o. No entanto, relatos de casos de RCV nesse perfil de pacientes t\u00eam demonstrado benef\u00edcios na qualidade de vida e na aptid\u00e3o f\u00edsica, especialmente quando h\u00e1 comprometimento funcional, mesmo ap\u00f3s a melhora do quadro agudo e otimiza\u00e7\u00e3o do tratamento medicamentoso. 283 Depois dessa avalia\u00e7\u00e3o, casos selecionados podem iniciar exerc\u00edcios moderados na RCV, visando os benef\u00edcios gerais obtidos com os pacientes com IC.Antes de iniciar a pr\u00e1tica de exerc\u00edcios, os pacientes com quadro pr\u00e9vio de miocardite devem ser submetidos a ecocardiograma, Holter de 24 horas e teste de exerc\u00edcio em um per\u00edodo n\u00e3o inferior a 3 a 6 meses ap\u00f3s a doen\u00e7a aguda. 269No \u00e2mbito esportivo, \u00e9 razo\u00e1vel que atletas retornem a sua rotina normal de treinamento apenas se houver: retorno da fun\u00e7\u00e3o sist\u00f3lica a valores normais; marcadores de necrose mioc\u00e1rdica e inflama\u00e7\u00e3o dentro dos valores normais e aus\u00eancia de arritmias clinicamente significativas no Holter e no teste de exerc\u00edcio. Ressalta-se que o significado cl\u00ednico da perman\u00eancia de realce tardio na resson\u00e2ncia de pacientes p\u00f3s-miocardite, com resolu\u00e7\u00e3o do quadro, ainda \u00e9 desconhecido. Sendo assim, parece razo\u00e1vel que aqueles com pequenas \u00e1reas de realce e sem arritmias significativas no Holter e no exerc\u00edcio possam retornar \u00e0 atividade esportiva, mantendo acompanhamento cl\u00ednico. Em casos cr\u00f4nicos, em que a disfun\u00e7\u00e3o ventricular persiste ao longo do seguimento, o paciente deve seguir as recomenda\u00e7\u00f5es gerais para a RCV descritas para a IC cr\u00f4nica ver .6.6.3.1. Cardiomiopatia Arritmog\u00eanica do Ventr\u00edculo Direito291A cardiomiopatia arritmog\u00eanica do ventr\u00edculo direito (CAVD) \u00e9 uma doen\u00e7a heredit\u00e1ria que est\u00e1 associada \u00e0 MS em jovens e atletas. Patologicamente, ocorre perda de mi\u00f3citos, com substitui\u00e7\u00e3o fibroadiposa, principalmente no mioc\u00e1rdio do ventr\u00edculo direito, embora o acometimento isolado do ventr\u00edculo esquerdo ou o biventricular tamb\u00e9m possam ocorrer. 292 Em indiv\u00edduos com gen\u00f3tipos positivos, o aumento do risco de arritmias com o exerc\u00edcio tamb\u00e9m j\u00e1 foi confirmado. Os eventos de taquiarritmias ventriculares e MS geralmente ocorrem durante o esfor\u00e7o, incluindo esportes e exerc\u00edcios de endurance , com um aumento no risco de taquicardia, fibrila\u00e7\u00e3o ventricular e IC. 293H\u00e1 evid\u00eancias, em modelo experimental animal, de que o exerc\u00edcio aumenta a penetr\u00e2ncia e o risco de arritmias em portadores de muta\u00e7\u00f5es tradicionais da CAVD. 270 A redu\u00e7\u00e3o da intensidade do exerc\u00edcio foi associada \u00e0 diminui\u00e7\u00e3o substancial do risco de taquiarritmias ventriculares ou morte, principalmente nos pacientes sem muta\u00e7\u00e3o desmossomal detectada e com CDI para preven\u00e7\u00e3o prim\u00e1ria. 294 Portanto, a evid\u00eancia cient\u00edfica sugere que a participa\u00e7\u00e3o em esportes e exerc\u00edcio intenso est\u00e1 associada ao in\u00edcio precoce dos sintomas e maior risco de arritmias ventriculares e eventos maiores em pacientes com CAVD. Sendo assim, devem ser desqualificados para a pr\u00e1tica esportiva. 276J\u00e1 foi demonstrado que indiv\u00edduos com CAVD envolvidos em esportes competitivos apresentaram maior ocorr\u00eancia de taquiarritmias ventriculares e MS, al\u00e9m de in\u00edcio mais precoce dos sintomas, comparados com aqueles que participaram apenas de atividade f\u00edsica leve e sedent\u00e1rios. 21Em rela\u00e7\u00e3o \u00e0 participa\u00e7\u00e3o em programas de RCV, n\u00e3o h\u00e1 dados cient\u00edficos que indiquem ou que sugiram benef\u00edcios dos exerc\u00edcios f\u00edsicos para os pacientes com CAVD. Por outro lado, mant\u00ea-los sedent\u00e1rios, contribuindo para a baixa aptid\u00e3o f\u00edsica, tamb\u00e9m pode n\u00e3o ser apropriado, visto que existe associa\u00e7\u00e3o geral de baixa aptid\u00e3o f\u00edsica com mortalidade. 270 Desse modo, pode-se supor que a participa\u00e7\u00e3o dos pacientes em um programa de RCV supervisionado, com exerc\u00edcios de leve a moderada intensidade, pode n\u00e3o ser delet\u00e9ria. Dependendo de outras caracter\u00edsticas cl\u00ednicas dos indiv\u00edduos, como presen\u00e7a de fatores de risco cardiovasculares, os exerc\u00edcios f\u00edsicos poderiam ser prescritos para o controle dessas condi\u00e7\u00f5es.Em um pequeno estudo observacional com pacientes com CAVD, n\u00e3o foi observada diferen\u00e7a na taxa de mortalidade entre os indiv\u00edduos inativos e os que realizaram apenas atividades f\u00edsicas recreacionais. versus riscos potenciais do sedentarismo e baixa aptid\u00e3o f\u00edsica. Cabe ao paciente escolher a op\u00e7\u00e3o, de acordo com suas prefer\u00eancias pessoais.Portanto, a inclus\u00e3o de um paciente com CAVD em programas de RCV somente deve ser realizada ap\u00f3s a avalia\u00e7\u00e3o m\u00e9dica pr\u00e9-participa\u00e7\u00e3o e o rigoroso ponderamento entre os riscos e benef\u00edcios dos exerc\u00edcios f\u00edsicos. Devem ser discutidas as op\u00e7\u00f5es com o paciente, expondo a aus\u00eancia de benef\u00edcios comprovados No contexto da RCV, extrapolando os achados em atletas, sugere-se tamb\u00e9m restri\u00e7\u00e3o a maiores intensidades de treinamento. Os pacientes com CAVD poderiam realizar exerc\u00edcios f\u00edsicos supervisionados de leve a moderada intensidade.6.6.3.2. Miocardiopatia N\u00e3o Compactada296A miocardiopatia n\u00e3o compactada (MNC) \u00e9 uma doen\u00e7a card\u00edaca que ocorre devido \u00e0 interrup\u00e7\u00e3o embrion\u00e1ria da compacta\u00e7\u00e3o mioc\u00e1rdica. Caracteriza-se por espessamento segmentar das paredes do ventr\u00edculo esquerdo, consistindo em duas camadas: uma epic\u00e1rdica compactada e uma endoc\u00e1rdica com marcadas trabecula\u00e7\u00f5es e recessos intratrabeculares profundos, onde os espa\u00e7os s\u00e3o preenchidos pelo fluxo sangu\u00edneo. 297 Clinicamente, pode ser assintom\u00e1tica ou cursar com sintomas de IC, arritmias ventriculares e/ou atriais, pr\u00e9-excita\u00e7\u00e3o, eventos tromboemb\u00f3licos ou MS. N\u00e3o existem crit\u00e9rios universalmente aceitos para o diagn\u00f3stico morfol\u00f3gico. Contudo, a rela\u00e7\u00e3o entre mioc\u00e1rdio n\u00e3o compactado/compactado superior a 2,1:1 no final da s\u00edstole ao ecocardiograma ou 2,3:1 no final da s\u00edstole na RMN tem sido o crit\u00e9rio proposto mais aceito atualmente. 298Sua incid\u00eancia e preval\u00eancia s\u00e3o incertas, segundo alguns registros ecocardiogr\u00e1ficos, em torno de 0,02 a 0,05%. 300 Em estudos recentes, atletas revelaram alta preval\u00eancia de aumento da trabecula\u00e7\u00e3o ventricular, quando comparados a um grupo controle . Acredita-se que o aumento da trabecula\u00e7\u00e3o ventricular ou a exist\u00eancia de crit\u00e9rios ecocardiogr\u00e1ficos isolados para miocardiopatias tenham, provavelmente, pequena signific\u00e2ncia e possam ser parte do espectro do cora\u00e7\u00e3o de atleta. 301 Portanto, nem todos os atletas com n\u00e3o compacta\u00e7\u00e3o ventricular isolada t\u00eam o diagn\u00f3stico de MNC. Diante disso, existe a necessidade de se considerarem par\u00e2metros funcionais, como a FEVE, para decis\u00e3o de conduta. 301Ainda n\u00e3o est\u00e1 estabelecido como o treinamento f\u00edsico pode influenciar a MNC ou a frequ\u00eancia de aparecimento da morfologia de n\u00e3o compacta\u00e7\u00e3o na popula\u00e7\u00e3o. N\u00e3o existem, at\u00e9 o momento, evid\u00eancias de estudos com RCV ou treinamento na MNC. Sendo assim, pacientes que apresentem disfun\u00e7\u00e3o ventricular esquerda devem seguir as mesmas recomenda\u00e7\u00f5es de exerc\u00edcio para aqueles com IC cr\u00f4nica ver .304Pacientes com valvopatias representam um grupo bastante heterog\u00eaneo e podem ter grande variabilidade quanto a faixa et\u00e1ria, etiologia, valvas acometidas e gravidade das les\u00f5es, seja por estenose, insufici\u00eancia ou les\u00f5es mistas. Entretanto, a maioria das valvopatias tem caracter\u00edsticas em comum, que s\u00e3o as manifesta\u00e7\u00f5es cl\u00ednicas induzidas pelo esfor\u00e7o f\u00edsico, como dor tor\u00e1cica, dispneia e/ou limita\u00e7\u00f5es funcionais. A gravidade desses sintomas em pacientes com valvopatias graves pode ser utilizada como um dos crit\u00e9rios para a indica\u00e7\u00e3o de interven\u00e7\u00e3o cir\u00fargica ou percut\u00e2nea. Al\u00e9m disso, a identifica\u00e7\u00e3o de redu\u00e7\u00e3o da aptid\u00e3o aer\u00f3bica, documentada pelo TCPE ou pelo TE, tamb\u00e9m se constitui em crit\u00e9rio utilizado para defini\u00e7\u00e3o de indica\u00e7\u00e3o de interven\u00e7\u00f5es. Um dos problemas no seguimento cl\u00ednico dos pacientes com valvopatias \u00e9 que a doen\u00e7a tem longa evolu\u00e7\u00e3o. Os sintomas e limita\u00e7\u00f5es funcionais podem ter lenta instala\u00e7\u00e3o e progress\u00e3o, o que pode levar o paciente a, espontaneamente, reduzir a sua pr\u00e1tica de atividade f\u00edsica, em virtude de sintomatologia aos esfor\u00e7os. Com isso, o sedentarismo pode contribuir para a redu\u00e7\u00e3o da aptid\u00e3o f\u00edsica aer\u00f3bica e amplificar sintomas.Desse modo, d\u00favidas podem surgir na condu\u00e7\u00e3o cl\u00ednica sobre a necessidade de interven\u00e7\u00f5es quando o paciente realizar um TE ou TCPE, como: a identifica\u00e7\u00e3o de eventuais redu\u00e7\u00f5es da aptid\u00e3o f\u00edsica poderia ser decorrente da evolu\u00e7\u00e3o da valvopatia, do sedentarismo ou de ambas as situa\u00e7\u00f5es? Nesse contexto, a pr\u00e1tica regular de exerc\u00edcios f\u00edsicos e a consequente manuten\u00e7\u00e3o ou at\u00e9 melhora da aptid\u00e3o f\u00edsica s\u00e3o importantes para dirimir d\u00favidas no seguimento de pacientes valvopatas.305 Por\u00e9m, o aumento da capacidade funcional dos indiv\u00edduos encaminhados \u00e0 RCV tem sido consistentemente encontrado, 307 justificando o encaminhamento a programas embasados em exerc\u00edcios f\u00edsicos (n\u00edvel de evid\u00eancia C).A participa\u00e7\u00e3o de pacientes valvopatas em programas de RCV ainda \u00e9 objeto de estudo sobre o significado em termos de custo-efetividade. A atua\u00e7\u00e3o da reabilita\u00e7\u00e3o no cen\u00e1rio da valvopatia pode ser subdividida em duas fases: pr\u00e9 e p\u00f3s-interven\u00e7\u00e3o, seja esta cir\u00fargica ou percut\u00e2nea.Pacientes com valvopatias moderadas a graves, na fase pr\u00e9-interven\u00e7\u00e3o, s\u00e3o menos comuns em programas de RCV. O treinamento \u00e9 realizado principalmente em casos assintom\u00e1ticos, nos quais ainda n\u00e3o existe indica\u00e7\u00e3o de corre\u00e7\u00e3o da valvopatia.310A RCV pode ser \u00fatil por manter o paciente fisicamente ativo durante a espera pela futura interven\u00e7\u00e3o. Afinal, o sedentarismo pode deteriorar sua capacidade funcional e, com isso, aumentar o risco de complica\u00e7\u00f5es no p\u00f3s-operat\u00f3rio, principalmente quando a interven\u00e7\u00e3o \u00e9 realizada em idosos com m\u00faltiplas comorbidades e fragilidade. Al\u00e9m disso, o monitoramento realizado durante as sess\u00f5es supervisionadas da RCV pode ser \u00fatil para observar mudan\u00e7as na sintomatologia e aptid\u00e3o f\u00edsica, as quais podem indicar uma poss\u00edvel progress\u00e3o da valvopatia e sugerir a necessidade de reavalia\u00e7\u00f5es m\u00e9dicas.Pacientes na fase p\u00f3s-interven\u00e7\u00e3o s\u00e3o mais comuns em programas de RCV, nos quais o exerc\u00edcio estruturado e sob supervis\u00e3o \u00e9 \u00fatil para a observa\u00e7\u00e3o do comportamento hemodin\u00e2mico da nova condi\u00e7\u00e3o valvar. A obten\u00e7\u00e3o de informa\u00e7\u00f5es relativas \u00e0s respostas ao exerc\u00edcio f\u00edsico pode ajudar o m\u00e9dico assistente em rela\u00e7\u00e3o \u00e0 necessidade de ajustes farmacol\u00f3gicos e/ou revis\u00f5es da fun\u00e7\u00e3o valvar. Al\u00e9m disso, a pr\u00e1tica supervisionada dos exerc\u00edcios confere maior seguran\u00e7a ao paciente para retornar \u00e0s suas atividades di\u00e1rias, de lazer e esporte.310 A troca de informa\u00e7\u00f5es entre o m\u00e9dico assistente e o da reabilita\u00e7\u00e3o configura-se como a melhor estrat\u00e9gia para defini\u00e7\u00e3o do momento mais prop\u00edcio para o encaminhamento, e a avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o tem um papel fundamental na consolida\u00e7\u00e3o dessa decis\u00e3o compartilhada.Apesar de n\u00e3o existir nenhum prazo de tempo consensualmente definido para o encaminhamento \u00e0 RCV no cen\u00e1rio da valvopatia, quanto mais precocemente o paciente iniciar os exerc\u00edcios, menores ser\u00e3o os preju\u00edzos relacionados \u00e0 inatividade f\u00edsica. A avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o sempre ter\u00e1 como pilares b\u00e1sicos e fundamentais a anamnese, o exame f\u00edsico e a avalia\u00e7\u00e3o dos exames complementares. A hist\u00f3ria cl\u00ednica deve contemplar: tempo de interna\u00e7\u00e3o; complica\u00e7\u00f5es relacionadas ao procedimento, como derrame pleural, peric\u00e1rdico, mediastinite e infec\u00e7\u00f5es; tipo e tamanho da pr\u00f3tese utilizada; t\u00e9cnica cir\u00fargica; e se houve CRVM associada, al\u00e9m de outras informa\u00e7\u00f5es cl\u00ednicas que possam ser pertinentes, relativas a outras comorbidades.No exame f\u00edsico, as auscultas card\u00edaca e pulmonar s\u00e3o importantes. Al\u00e9m disso, aten\u00e7\u00e3o deve ser dada \u00e0 cicatriz cir\u00fargica, com verifica\u00e7\u00e3o de sinais de inflama\u00e7\u00e3o e infec\u00e7\u00e3o, instabilidade do esterno e dor ou desconforto \u00e0 palpa\u00e7\u00e3o. Caso tenha sido realizada revasculariza\u00e7\u00e3o concomitante, deve-se observar a regi\u00e3o da safenectomia e/ou da retirada da art\u00e9ria radial. Em casos de procedimentos percut\u00e2neos, verifica-se a via de acesso em busca de sinais de complica\u00e7\u00f5es vasculares perif\u00e9ricas.311 A avalia\u00e7\u00e3o laboratorial da coagula\u00e7\u00e3o \u00e9 relevante nos pacientes que receberam pr\u00f3teses valvares e iniciaram uso de anticoagulantes. A adequa\u00e7\u00e3o do n\u00edvel de anticoagula\u00e7\u00e3o \u00e9 importante na preven\u00e7\u00e3o de complica\u00e7\u00f5es.A busca por anemia no exame f\u00edsico e na avalia\u00e7\u00e3o dos exames complementares \u00e9 importante nos pacientes p\u00f3s-interven\u00e7\u00e3o, pois \u00e9 uma situa\u00e7\u00e3o frequente e pode impactar negativamente na capacidade funcional. 312O ECG de repouso deve ser realizado para avaliar a ocorr\u00eancia de arritmias e dist\u00farbios do ritmo e da condu\u00e7\u00e3o. O exame mais comumente utilizado na avalia\u00e7\u00e3o das valvopatias \u00e9 o ecocardiograma com Doppler, que possibilita a avalia\u00e7\u00e3o da fun\u00e7\u00e3o ventricular e de di\u00e2metros cavit\u00e1rios, a mensura\u00e7\u00e3o de gradientes transvalvares, a estimativa da press\u00e3o sist\u00f3lica da art\u00e9ria pulmonar e as medidas dos fluxos, o que d\u00e1 uma vis\u00e3o ampla do funcionamento do aparelho valvar e da fun\u00e7\u00e3o card\u00edaca em repouso. Sendo assim, o ecocardiograma deve ser realizado antes do in\u00edcio da RCV para avaliar o risco de complica\u00e7\u00f5es nos exerc\u00edcios. 316 Esses exames, principalmente o TCPE, fornecem informa\u00e7\u00f5es relativas \u00e0 aptid\u00e3o aer\u00f3bica e \u00e0 repercuss\u00e3o hemodin\u00e2mica da les\u00e3o valvar, que pode estar subestimada pela avalia\u00e7\u00e3o em repouso. Al\u00e9m disso, identifica par\u00e2metros que s\u00e3o utilizados para guiar a prescri\u00e7\u00e3o dos limites de intensidade e restri\u00e7\u00f5es causadas pela valvopatia. Na indisponibilidade do TCPE ou TE, a utiliza\u00e7\u00e3o de outros testes funcionais, como o teste de caminhada de 6 minutos e o teste de degraus, deve ser considerada. 320A avalia\u00e7\u00e3o da capacidade funcional pelo TCPE ou TE \u00e9 de import\u00e2ncia \u00edmpar na an\u00e1lise complementar. 321\u00c9 importante ressaltar que o TCPE ou TE em pacientes com les\u00f5es esten\u00f3ticas configuram situa\u00e7\u00f5es de maior risco. Por esta raz\u00e3o, devem ser realizados somente por m\u00e9dicos com experi\u00eancia nesse tipo de avalia\u00e7\u00e3o e em servi\u00e7o com retaguarda de seguran\u00e7a. 323Al\u00e9m da indica\u00e7\u00e3o dos testes funcionais na avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o da RCV, sua utiliza\u00e7\u00e3o tamb\u00e9m \u00e9 adequada para esclarecer d\u00favidas em rela\u00e7\u00e3o \u00e0 sintomatologia de pacientes valvopatas na fase pr\u00e9-interven\u00e7\u00e3o. A associa\u00e7\u00e3o com o ecocardiograma ajuda a avaliar a resposta em esfor\u00e7o f\u00edsico do gradiente transvalvar e da press\u00e3o sist\u00f3lica de art\u00e9ria pulmonar, principalmente quando h\u00e1 discrep\u00e2ncia entre os achados do ecocardiograma em repouso e os sinais e sintomas cl\u00ednicos. 324 Em virtude do elevado risco cir\u00fargico, tais pacientes t\u00eam sido submetidos a procedimentos percut\u00e2neos das valvas a\u00f3rtica 325 e mitral. 326 Nessa situa\u00e7\u00e3o, a RCV pode ser considerada antes da interven\u00e7\u00e3o, com o objetivo de diminuir as taxas de complica\u00e7\u00f5es, o tempo de interna\u00e7\u00e3o, a mortalidade e a morbidade relacionadas \u00e0 s\u00edndrome da fragilidade. 327 Ap\u00f3s a realiza\u00e7\u00e3o da interven\u00e7\u00e3o, a RCV permite monitorar e otimizar os resultados do procedimento em todos os seus aspectos. 331Outra quest\u00e3o relevante \u00e9 a avalia\u00e7\u00e3o de pacientes idosos, que frequentemente s\u00e3o acometidos pelas doen\u00e7as valvares e apresentam uma alta preval\u00eancia de fatores de risco e comorbidades. 332A utiliza\u00e7\u00e3o de instrumentos de avalia\u00e7\u00e3o da s\u00edndrome de fragilidade ainda \u00e9 objeto de discuss\u00e3o na literatura, sem um consenso de qual o melhor protocolo para avaliar os resultados da RCV. A avalia\u00e7\u00e3o deve incluir testes objetivos e instrumentos de abordagem do risco em v\u00e1rios dom\u00ednios: mobilidade, massa e for\u00e7a musculares, independ\u00eancia nas atividades da vida di\u00e1ria, fun\u00e7\u00e3o cognitiva, nutri\u00e7\u00e3o, ansiedade e depress\u00e3o. 334 A evid\u00eancia cient\u00edfica \u00e9 escassa quanto ao impacto do exerc\u00edcio f\u00edsico regular na progress\u00e3o da doen\u00e7a valvar e de suas complica\u00e7\u00f5es. Portanto, as recomenda\u00e7\u00f5es s\u00e3o fundamentadas em opini\u00f5es de especialistas (n\u00edvel de evid\u00eancia C).Nesta se\u00e7\u00e3o, s\u00e3o abordadas as orienta\u00e7\u00f5es e recomenda\u00e7\u00f5es para exerc\u00edcios em pacientes com les\u00f5es valvares de grau moderado ou grave, visto que n\u00e3o h\u00e1 restri\u00e7\u00e3o para a pr\u00e1tica nos casos de les\u00f5es leves. A participa\u00e7\u00e3o em esportes competitivos deve observar as publica\u00e7\u00f5es espec\u00edficas sobre o assunto. Agudamente, o exerc\u00edcio provoca um aumento do t\u00f4nus adren\u00e9rgico e da carga hemodin\u00e2mica imposta ao sistema cardiovascular, o que causa preocupa\u00e7\u00e3o com rela\u00e7\u00e3o aos potenciais efeitos delet\u00e9rios cardiovasculares nos pacientes com valvopatias, tais como progress\u00e3o de aortopatias, deteriora\u00e7\u00e3o funcional, hipertens\u00e3o pulmonar, remodelamento card\u00edaco, isquemia mioc\u00e1rdica e arritmias.Pacientes com doen\u00e7as valvares que iniciar\u00e3o um programa de RCV devem ser submetidos a um teste de esfor\u00e7o para avalia\u00e7\u00e3o e prescri\u00e7\u00e3o dos exerc\u00edcios. A Para pacientes sintom\u00e1ticos, sem indica\u00e7\u00e3o de corre\u00e7\u00e3o cir\u00fargica ou que n\u00e3o apresentam as caracter\u00edsticas descritas na Em pacientes que foram submetidos \u00e0 corre\u00e7\u00e3o cir\u00fargica da valvopatia, os limites de intensidade da prescri\u00e7\u00e3o depender\u00e3o da doen\u00e7a de base, do resultado do procedimento, da presen\u00e7a de les\u00f5es residuais, da fun\u00e7\u00e3o ventricular e da resposta ao teste de exerc\u00edcio. Sendo assim, cada caso deve ser analisado individualmente e os limites definidos pela avalia\u00e7\u00e3o m\u00e9dica e pelos resultados dos exames complementares realizados.Esta se\u00e7\u00e3o se destina \u00e0s particularidades sobre os dispositivos implant\u00e1veis: marcapasso card\u00edaco (MP) e cardioversor desfibrilador implant\u00e1vel (CDI). O MP \u00e9 indicado em fun\u00e7\u00e3o de anormalidades el\u00e9tricas, que podem ser isolada de grau avan\u00e7ado) ou associada a cardiopatias estruturais. O CDI \u00e9 indicado para a preven\u00e7\u00e3o prim\u00e1ria ou secund\u00e1ria de MS, em pacientes com doen\u00e7as el\u00e9tricas e/ou cardiopatias graves. Dependendo da cardiopatia presente, devem ser consideradas as recomenda\u00e7\u00f5es sobre a RCV abordadas anteriormente.336 Os profissionais de sa\u00fade tamb\u00e9m compartilham desses receios, 337 o que pode reduzir as orienta\u00e7\u00f5es para a pr\u00e1tica de exerc\u00edcios. Por\u00e9m, tem sido demonstrado que o exerc\u00edcio f\u00edsico \u00e9 seguro e n\u00e3o est\u00e1 associado ao aumento do risco de choques ou de outros eventos adversos. 342 Al\u00e9m disso, n\u00e3o t\u00eam sido observadas complica\u00e7\u00f5es relativas ao CDI, mesmo em atletas competitivos. 344Uma das preocupa\u00e7\u00f5es nos exerc\u00edcios f\u00edsicos em portadores de MP ou CDI \u00e9 relativa ao risco de complica\u00e7\u00f5es com o dispositivo, especialmente em atividades com chances de colis\u00e3o corporal. Nos portadores de CDI, h\u00e1 o receio de choques, o que pode levar a modifica\u00e7\u00f5es comportamentais nos pacientes, com redu\u00e7\u00e3o da atividade f\u00edsica di\u00e1ria e participa\u00e7\u00e3o em exerc\u00edcios de moderada intensidade. Entretanto, \u00e9 de fundamental import\u00e2ncia, para a adequada libera\u00e7\u00e3o dos exerc\u00edcios, que se conhe\u00e7am o motivo do implante e os par\u00e2metros de programa\u00e7\u00e3o do dispositivo, que dever\u00e3o ser investigados na avalia\u00e7\u00e3o pr\u00e9-participa\u00e7\u00e3o.342 que englobou 14 estudos com um total de 2.681 pacientes portadores de CDI comprovou efeito ben\u00e9fico do exerc\u00edcio f\u00edsico na capacidade funcional desses indiv\u00edduos, com um aumento m\u00e9dio de VO 2 de 2,4 ml.kg - 1 .min - 1 . Em outra meta-an\u00e1lise, com cinco estudos randomizados e um n\u00e3o randomizado em pacientes com IC e CDI, 341 o resultado na capacidade f\u00edsica foi semelhante, com aumento no VO 2 pico de 1,98 ml.kg - 1 .min - 1 em rela\u00e7\u00e3o ao grupo controle.Uma meta-an\u00e1lise 342 Sendo assim, apesar dos receios previamente descritos, o treinamento f\u00edsico n\u00e3o se associou a aumento dos choques pelo CDI e se mostrou seguro.Quanto \u00e0s terapias por CDI e treinamento f\u00edsico, uma das meta-an\u00e1lises n\u00e3o encontrou diferen\u00e7as significativas. O percentual de choques associados aos exerc\u00edcio variou de 0 a 20% entre os estudos, com uma m\u00e9dia de 2,2%, similar ao percentual de choques em um per\u00edodo de seguimento n\u00e3o relacionado ao exerc\u00edcio. 345Outra meta-an\u00e1lise relatou menor probabilidade de choques ao longo do seguimento nos pacientes participantes da RCV em rela\u00e7\u00e3o aos controles, corroborando o resultado anterior de um estudo observacional, que relatou maior incid\u00eancia de choques pelo CDI em pacientes que n\u00e3o participavam de programas de RCV. 346 Al\u00e9m disso, os exerc\u00edcios poderiam reduzir a arritmogenicidade mioc\u00e1rdica, em fun\u00e7\u00e3o do remodelamento e da menor excitabilidade simp\u00e1tica. 347Uma das poss\u00edveis explica\u00e7\u00f5es para a menor incid\u00eancia de arritmias e choques nos pacientes em RCV seria a melhora da capacidade f\u00edsica, pois j\u00e1 foi previamente documentado que maior aptid\u00e3o f\u00edsica est\u00e1 associada a menor incid\u00eancia de arritmias. 348Em um estudo nacional com 10 anos de acompanhamento, que contou com 150 pacientes com CDI em programa de RCV, submetidos a TCPE ou TE para prescri\u00e7\u00e3o do treinamento, ocorreram apenas tr\u00eas eventos de choques apropriados, o que refor\u00e7a a seguran\u00e7a das avalia\u00e7\u00f5es e da RCV nesses indiv\u00edduos. O exerc\u00edcio f\u00edsico pode e deve ser indicado desde que a condi\u00e7\u00e3o cl\u00ednica do paciente seja est\u00e1vel e o tratamento cl\u00ednico, otimizado. Al\u00e9m dos benef\u00edcios potenciais na cardiopatia de base, a RCV contribui para o aumento da capacidade f\u00edsica e pode atuar na redu\u00e7\u00e3o das arritmias e nos choques pelo CDI .burst .Nos portadores de dispositivos implant\u00e1veis h\u00e1 necessidade de conhecer o motivo do implante, a fun\u00e7\u00e3o ventricular, a presen\u00e7a de arritmias e, principalmente, os par\u00e2metros de ajuste do dispositivo. Dentre os ajustes do MP, \u00e9 importante saber o modo de programa\u00e7\u00e3o, os limites programados de FC, o tipo e a adapta\u00e7\u00e3o do sensor de frequ\u00eancia. Nos pacientes com CDI, \u00e9 fundamental obter informa\u00e7\u00f5es relativas \u00e0s FC programadas para terapias de choque ou Al\u00e9m da avalia\u00e7\u00e3o cl\u00ednica habitual, a avalia\u00e7\u00e3o ao esfor\u00e7o \u00e9 de suma import\u00e2ncia, sendo ideal a realiza\u00e7\u00e3o do TCPE ou TE para determina\u00e7\u00e3o da capacidade funcional e an\u00e1lise do comportamento do dispositivo em esfor\u00e7o. Entretanto, a impossibilidade de realiza\u00e7\u00e3o desses exames n\u00e3o dever\u00e1 ser um fator limitante para a pr\u00e1tica de exerc\u00edcios. Nesses casos, o monitoramento das sess\u00f5es poder\u00e1 dar ind\u00edcios da necessidade de adequa\u00e7\u00f5es na programa\u00e7\u00e3o do dispositivo, geralmente em rela\u00e7\u00e3o aos ajustes da FC m\u00e1xima programada e da resposta do sensor.350 Em virtude das altera\u00e7\u00f5es do tra\u00e7ado, causadas pelo comando artificial, pode haver erro na determina\u00e7\u00e3o autom\u00e1tica da FC, tanto pela an\u00e1lise eletrocardiogr\u00e1fica quanto pelos frequenc\u00edmetros. Sendo assim, \u00e9 importante aten\u00e7\u00e3o a esses poss\u00edveis erros, com verifica\u00e7\u00e3o manual, se necess\u00e1ria.Durante as sess\u00f5es da RCV, poder\u00e1 ser utilizado o monitoramento eletrocardiogr\u00e1fico, que pode ser feito com o uso de sistemas de telemetria. Dispositivos para controle da FC, como os cardiofrequenc\u00edmetros, tamb\u00e9m podem ser usados para monitoramento durante as sess\u00f5es de RCV desses pacientes. burst ). Esse cuidado \u00e9 especialmente importante em indiv\u00edduos jovens, que podem ter FC elevada no treinamento. Em pacientes mais idosos, com IC e uso de altas doses de betabloqueadores, a FC pico observada no TE ou TCPE costuma estar abaixo da FC de terapia do CDI.Na prescri\u00e7\u00e3o e defini\u00e7\u00e3o de limites de intensidade para o treinamento f\u00edsico aer\u00f3bico, deve-se ter conhecimento da programa\u00e7\u00e3o do CDI e limitar a intensidade a 10 a 20 bpm abaixo da FC programada para a terap\u00eautica Resposta cronotr\u00f3pica sinusal normal ou deprimida. MP sem atua\u00e7\u00e3o (inibido). A resposta cronotr\u00f3pica em esfor\u00e7o \u00e9 mediada pelo ritmo sinusal e pode estar normal ou deprimida . A condu\u00e7\u00e3o ventricular ocorre pela via pr\u00f3pria, e o MP n\u00e3o atua no esfor\u00e7o. Em alguns casos, ele pode atuar em repouso e em cargas iniciais, com comando atrial e/ou ventricular. Por\u00e9m, no esfor\u00e7o, o MP se inibe, predominando as respostas sinusais e a condu\u00e7\u00e3o ventricular pela via pr\u00f3pria. Nesse tipo de resposta ao esfor\u00e7o, a prescri\u00e7\u00e3o de intensidade segue as rotinas habituais e n\u00e3o \u00e9 influenciada pela presen\u00e7a do MP.2) Resposta cronotr\u00f3pica sinusal normal ou deprimida. MP com comando ventricular no esfor\u00e7o. A resposta cronotr\u00f3pica em esfor\u00e7o \u00e9 mediada pelo ritmo sinusal, que \u00e9 percebida pelo MP com subsequente comando ventricular, de modo sincronizado e de acordo com os intervalos AV programados. Nesse caso, se a programa\u00e7\u00e3o do limite m\u00e1ximo de resposta de FC do MP for adequada \u00e0 resposta sinusal do paciente, n\u00e3o haver\u00e1 problema para a prescri\u00e7\u00e3o de intensidade por FC, pois o ventr\u00edculo estar\u00e1 pareado com a atividade sinusal. Por\u00e9m, se a FC m\u00e1xima programada do MP for inferior \u00e0 resposta sinusal do paciente, em moderada a alta carga haver\u00e1 uma perda do pareamento da atividade ventricular com a sinusal. Ent\u00e3o, o MP bloquear\u00e1 alguns est\u00edmulos sinusais por meio de um Wenckebach mediado pelo dispositivo ou eletr\u00f4nico, 352 de modo a manter a FC ventricular dentro do limite programado, havendo um plat\u00f4 na resposta cronotr\u00f3pica ao esfor\u00e7o. Nessas situa\u00e7\u00f5es, a perda do pareamento do ritmo sinusal com a frequ\u00eancia ventricular limitar\u00e1 a utiliza\u00e7\u00e3o da FC para controle de intensidade. A prescri\u00e7\u00e3o dever\u00e1 ser feita por cargas relativas e/ou sensa\u00e7\u00e3o subjetiva de esfor\u00e7o.Wenckebach eletr\u00f4nico, \u00e9 necess\u00e1ria a extrema aten\u00e7\u00e3o no TCPE ou TE, ap\u00f3s iniciar a sua ocorr\u00eancia. \u00c9 fundamental ter a informa\u00e7\u00e3o precisa de qual \u00e9 a FC atrial em que o MP inicia o bloqueio 2:1, pois nela o comando ventricular ser\u00e1 na propor\u00e7\u00e3o 2:1, podendo ocorrer queda s\u00fabita da FC em esfor\u00e7o, a qual pode ser sintom\u00e1tica, por redu\u00e7\u00e3o abrupta do d\u00e9bito card\u00edaco. Sendo assim, a menos que as FC programadas de Wenckebach eletr\u00f4nico e de bloqueio 2:1 sejam muito distantes, a FC do Wenckebach eletr\u00f4nico poder\u00e1 tornar-se um limite para o TCPE ou TE e para a prescri\u00e7\u00e3o dos exerc\u00edcios.No caso do Nesses casos, deve-se considerar e discutir com o m\u00e9dico assistente a reprograma\u00e7\u00e3o do MP, para melhor pareamento com a resposta sinusal do paciente. Outra op\u00e7\u00e3o, a depender do quadro cl\u00ednico, \u00e9 a otimiza\u00e7\u00e3o de medica\u00e7\u00f5es cronotr\u00f3picas negativas, como os betabloqueadores. Com isso, a menor resposta sinusal poder\u00e1 evitar a ocorr\u00eancia descrita.3) Resposta cronotr\u00f3pica mediada pelo MP e fixa, com aus\u00eancia de sensor. Alguns pacientes podem n\u00e3o ter atividade sinusal, como na fibrila\u00e7\u00e3o atrial. Nesses casos, em indiv\u00edduos com bloqueio AV completo, haver\u00e1 total depend\u00eancia de comando ventricular pelo MP. Se n\u00e3o houver sensor, ou se este estiver desativado, haver\u00e1 aus\u00eancia de resposta cronotr\u00f3pica ao esfor\u00e7o e o MP ter\u00e1 FC fixa. Esse tipo de MP ou programa\u00e7\u00e3o \u00e9 muito raro atualmente e tal resposta limita completamente a utiliza\u00e7\u00e3o da FC na prescri\u00e7\u00e3o, que deve se basear na determina\u00e7\u00e3o da intensidade por cargas e/ou pela sensa\u00e7\u00e3o subjetiva do esfor\u00e7o.4) Resposta cronotr\u00f3pica mediada pelo MP com presen\u00e7a de sensor. Nos pacientes com fibrila\u00e7\u00e3o atrial e bloqueio AV, conforme descrito anteriormente, mas com o sensor do MP presente e ativado, haver\u00e1 depend\u00eancia do comando ventricular, mas a ativa\u00e7\u00e3o do sensor no esfor\u00e7o conduzir\u00e1 a uma resposta cronotr\u00f3pica mediada pelo MP. Em pacientes com ritmo sinusal, mas com grande d\u00e9ficit cronotr\u00f3pico, por doen\u00e7a do n\u00f3 sinusal e/ou efeito medicamentoso, poder\u00e1 ocorrer resposta cronotr\u00f3pica ao esfor\u00e7o tamb\u00e9m mediada pelo sensor do MP, com comando atrial seguido ou n\u00e3o de comando ventricular.A velocidade e magnitude da resposta do sensor ao esfor\u00e7o s\u00e3o program\u00e1veis, com possibilidades de ajustes do limiar da ativa\u00e7\u00e3o do sensor, da velocidade de incremento da FC ao esfor\u00e7o e sua redu\u00e7\u00e3o na recupera\u00e7\u00e3o, bem como do limite m\u00e1ximo da FC do sensor. Na realiza\u00e7\u00e3o do TCPE ou TE, poder\u00e1 ser verificada a adequa\u00e7\u00e3o da resposta, com identifica\u00e7\u00e3o de poss\u00edveis necessidades de reprograma\u00e7\u00f5es do MP, que devem ser discutidas com o m\u00e9dico assistente.Nesses casos, como a resposta cronotr\u00f3pica ser\u00e1 mediada artificialmente pelo dispositivo, a prescri\u00e7\u00e3o de intensidade de exerc\u00edcios por FC poder\u00e1 ser imprecisa. Sendo assim, a utiliza\u00e7\u00e3o das cargas relativas e/ou a percep\u00e7\u00e3o do esfor\u00e7o ser\u00e3o preferenciais.Dispositivos com sensores do tipo aceler\u00f4metro e detec\u00e7\u00e3o do movimento axial, que s\u00e3o os mais usuais, t\u00eam boa resposta ao esfor\u00e7o na esteira, caminhada ou corrida. Por\u00e9m, na bicicleta estacion\u00e1ria n\u00e3o h\u00e1 movimento vertical e o sensor n\u00e3o ativa ou \u00e9 pouco ativado. Com isso, h\u00e1 menor resposta cronotr\u00f3pica no erg\u00f4metro, que pode variar de acordo com a resposta individual do paciente.A pr\u00e1tica de exerc\u00edcios resistidos \u00e9 importante na RCV em diversas cardiopatias. Entretanto, ap\u00f3s o implante do dispositivo, alguns cuidados s\u00e3o necess\u00e1rios at\u00e9 a completa cicatriza\u00e7\u00e3o, a fim de evitar les\u00e3o vascular, deslocamento do gerador e fratura de eletrodos. Recomenda-se, por exemplo, cautela ao executar exerc\u00edcios com pesos e eleva\u00e7\u00e3o excessiva dos MMSS durante as primeiras seis semanas ap\u00f3s o implante. Al\u00e9m disso, movimentos repetitivos e intensos com o membro relacionado ao implante do marcapasso devem ser evitados.353Entretanto, tais orienta\u00e7\u00f5es est\u00e3o mais ligadas a pacientes envolvidos com esportes, sendo improv\u00e1veis no caso de exerc\u00edcios realizados em centros de RCV. Em um estudo com mobiliza\u00e7\u00e3o precoce e supervisionada da cintura escapular ap\u00f3s o implante imediato de MP, n\u00e3o foram observadas complica\u00e7\u00f5es ao dispositivo. 356 Entretanto, tamb\u00e9m \u00e9 crescente o uso de dispositivos eletr\u00f4nicos nesses pacientes , o que causa preocupa\u00e7\u00e3o quanto ao uso da ENM, pela possibilidade de interfer\u00eancia eletromagn\u00e9tica.A utiliza\u00e7\u00e3o da eletroestimula\u00e7\u00e3o neuromuscular (ENM) em pacientes com IC tem sido difundida, principalmente naqueles impossibilitados de praticar exerc\u00edcios f\u00edsicos pela gravidade cl\u00ednica. A ENM pode melhorar a capacidade aer\u00f3bica, a for\u00e7a muscular e a \u00e1rea transversa da musculatura do quadr\u00edceps, demonstrando ser uma efetiva op\u00e7\u00e3o de exerc\u00edcio passivo nessa popula\u00e7\u00e3o. 349 demonstrou que a ENM na musculatura de quadr\u00edceps parece ser segura e vi\u00e1vel em pacientes com IC e CDI com sensores bipolares. Entretanto, a pr\u00f3pria revis\u00e3o ressalta que o n\u00famero de estudos e pacientes avaliados \u00e9 muito pequeno para conclus\u00f5es mais abrangentes e conclui que o uso pode ser feito se forem satisfeitas as seguintes condi\u00e7\u00f5es:Uma revis\u00e3o sistem\u00e1tica Se os riscos individuais tiverem sido exclu\u00eddos antes de iniciar a ENM.Se o uso da ENM for realizado apenas nos m\u00fasculos de quadr\u00edceps e gl\u00fateos.Se o tratamento for regularmente supervisionado por um m\u00e9dico e o dispositivo for avaliado ap\u00f3s o uso da ENM.Portanto, no momento, apesar de parecer ser segura a utiliza\u00e7\u00e3o da ENM em portadores de dispositivos com sensores bipolares, quando realizada em musculaturas distantes do implante, ainda h\u00e1 necessidade de estudos com maior n\u00famero de pacientes, para que haja possibilidade de uso amplo sem avalia\u00e7\u00e3o detalhada do dispositivo.358 Nesse contexto, destaca-se a doen\u00e7a arterial obstrutiva perif\u00e9rica (DAOP) de MMII, que, em seu est\u00e1gio mais grave, a isquemia cr\u00edtica, apresenta elevado risco de eventos cardiovasculares, amputa\u00e7\u00e3o de MMII e morte. A isquemia cr\u00edtica dos MMII, com o crescimento de fatores de risco, tais como idade, diabetes e tabagismo, tem aumentado a sua preval\u00eancia e acomete, atualmente, cerca de 2 milh\u00f5es de indiv\u00edduos somente nos EUA. 359O AVC tem sido justamente tratado como doen\u00e7a grave e de grande repercuss\u00e3o em sa\u00fade p\u00fablica. Por\u00e9m, outras doen\u00e7as arteriais perif\u00e9ricas tamb\u00e9m s\u00e3o muito prevalentes e apresentam grande morbimortalidade, embora n\u00e3o tenham sido devidamente abordadas, prejudicando a preven\u00e7\u00e3o, o diagn\u00f3stico e o efetivo tratamento. 361 O ITB tem sido recomendado como recurso diagn\u00f3stico a ser usado anteriormente \u00e0 realiza\u00e7\u00e3o de m\u00e9todos de imagem. 362 Testes funcionais em esfor\u00e7o podem ser necess\u00e1rios para auxiliar no diagn\u00f3stico, especialmente quando o ITB for maior que 0,91, e tamb\u00e9m para classifica\u00e7\u00e3o funcional e prescri\u00e7\u00e3o de exerc\u00edcios na RCV.A presen\u00e7a de DAOP \u00e9 suspeitada quando h\u00e1 dor em MMII ao esfor\u00e7o, sem aparente etiologia ortop\u00e9dica, e o \u00edndice tornozelo-braquial (ITB) \u00e9 menor que 0,90 em repouso. A caminhada pode ser avaliada por meio de testes de campo, que possibilitam o diagn\u00f3stico de claudica\u00e7\u00e3o intermitente, com determina\u00e7\u00e3o das dist\u00e2ncias percorridas para o in\u00edcio da sintomatologia e para o surgimento da total limita\u00e7\u00e3o funcional (claudica\u00e7\u00e3o absoluta).363 Outro estudo relatou notas de corte menores, sendo sugerida DAOP quando h\u00e1 redu\u00e7\u00e3o do ITB p\u00f3s-exerc\u00edcio acima de 18,5% e diminui\u00e7\u00e3o da press\u00e3o p\u00f3s-exerc\u00edcio maior que 15 mmHg. 364Em esteira, tem sido proposta a utiliza\u00e7\u00e3o diagn\u00f3stica de teste funcional em esfor\u00e7o, com medida do ITB em repouso e ap\u00f3s exerc\u00edcio. A presen\u00e7a de DAOP \u00e9 sugerida quando ocorre redu\u00e7\u00e3o do ITB p\u00f3s-exerc\u00edcio superior a 20% em rela\u00e7\u00e3o ao repouso, ou diminui\u00e7\u00e3o da press\u00e3o p\u00f3s-exerc\u00edcio maior que 30 mmHg em rela\u00e7\u00e3o ao repouso. 363Considerando o risco cardiovascular global desses pacientes, o tratamento cl\u00ednico otimizado deve sempre ser institu\u00eddo. Al\u00e9m disso, a interrup\u00e7\u00e3o do tabagismo e a terapia farmacol\u00f3gica com estatinas e antiagregantes plaquet\u00e1rios devem ser consideradas, bem como o adequado controle glic\u00eamico e press\u00f3rico. Em rela\u00e7\u00e3o ao uso do Cilostazol, n\u00e3o h\u00e1 um consenso nas diretrizes de sociedades m\u00e9dicas. 365 As atividades f\u00edsicas realizadas sob supervis\u00e3o direta t\u00eam se mostrado mais efetivas do que sem supervis\u00e3o. 366Em pacientes sintom\u00e1ticos, os exerc\u00edcios t\u00eam potencial para influenciar na morbimortalidade, com redu\u00e7\u00e3o dos sintomas, melhora da qualidade de vida e aumento da dist\u00e2ncia m\u00e1xima caminhada . 365 As367Em 14 ensaios cl\u00ednicos (1.002 participantes), com interven\u00e7\u00e3o entre 6 semanas e 12 meses, a caminhada livre de dor aumentou cerca de 180 metros a mais no treinamento sob supervis\u00e3o direta, quando comparado ao treinamento sob supervis\u00e3o indireta. O treinamento f\u00edsico tem se mostrado seguro. Na maioria dos estudos s\u00e3o realizados exerc\u00edcios de caminhadas, com indu\u00e7\u00e3o do sintoma de claudica\u00e7\u00e3o, em programas com dura\u00e7\u00e3o m\u00ednima de 3 meses e, pelo menos, tr\u00eas sess\u00f5es semanais. 368 embora aquele sob supervis\u00e3o indireta (RCV domiciliar) tenha se mostrado uma boa alternativa, com efeitos positivos sobre a qualidade de vida e a toler\u00e2ncia \u00e0 caminhada, sendo significativamente superior \u00e0 mera recomenda\u00e7\u00e3o para caminhar. 370Nos paciente com DAOP, o treinamento sob supervis\u00e3o direta tem sido superior em termos de custo-efetividade, 371 Cabe ainda ressaltar que exerc\u00edcios f\u00edsicos n\u00e3o podem ser realizados por pacientes com isquemia cr\u00edtica, mas devem ser considerados o mais breve poss\u00edvel ap\u00f3s tratamento intervencionista com sucesso. 373Quando a caminhada n\u00e3o puder ser realizada, outros tipos de atividades, como ciclismo, exerc\u00edcios resistidos e erg\u00f4metro de MMSS, t\u00eam se mostrado efetivos. 374 A interven\u00e7\u00e3o endovascular e a cirurgia aberta t\u00eam comprovadamente se mostrado eficazes para o al\u00edvio de sintomas, aumento da dist\u00e2ncia caminhada e melhora da qualidade de vida. Al\u00e9m disso, est\u00e3o indicadas quando, ap\u00f3s a realiza\u00e7\u00e3o do tratamento cl\u00ednico pleno ou otimizado (exerc\u00edcios f\u00edsicos e tratamento farmacol\u00f3gico otimizado), persistirem sintomas graves que influenciem negativamente na vida di\u00e1ria.Uma revis\u00e3o sistem\u00e1tica de 12 ensaios cl\u00ednicos, com um total de 1.548 pacientes, comparando os claudicantes em tratamento farmacol\u00f3gico (em treinamento f\u00edsico), os com interven\u00e7\u00e3o endovascular e os com cirurgia aberta, mostrou que todas as alternativas proporcionaram aumento da dist\u00e2ncia caminhada, redu\u00e7\u00e3o de sintomas e melhora da qualidade de vida. stent . 375 Entretanto, ap\u00f3s 18 meses de seguimento, os benef\u00edcios funcionais e na qualidade de vida foram equivalentes nos grupos com treinamento ou com revasculariza\u00e7\u00e3o e, em ambos os casos, foram superiores aos do grupo que realizou somente tratamento farmacol\u00f3gico. 376Em um ensaio cl\u00ednico randomizado com 111 pacientes com DAOP aortoil\u00edaca e seguimento de 6 meses, foi evidenciado que o aumento do tempo de exerc\u00edcio no TE incremental foi maior no grupo que realizou exerc\u00edcios supervisionados do que no que realizou revasculariza\u00e7\u00e3o com vers us treinamento f\u00edsico supervisionado) ou utilizaram uma abordagem que n\u00e3o permitia a inclus\u00e3o e a compara\u00e7\u00e3o direta de todos os tratamentos dispon\u00edveis da claudica\u00e7\u00e3o intermitente. 377V\u00e1rios ensaios cl\u00ednicos compararam a efic\u00e1cia e efetividade do exerc\u00edcio f\u00edsico supervisionado, a angioplastia e o tratamento cl\u00ednico otimizado, utilizando uma infinidade de desenhos diferentes. A maioria dos ensaios consistia em dois bra\u00e7os de tratamento. Revis\u00f5es sistem\u00e1ticas j\u00e1 citadas sugeriram que o exerc\u00edcio f\u00edsico supervisionado pode ser superior ao tratamento farmacol\u00f3gico otimizado ou \u00e0 angioplastia. Essas meta-an\u00e1lises, no entanto, inclu\u00edram estudos com compara\u00e7\u00f5es diretas entre dois bra\u00e7os de tratamento espec\u00edficos . As compara\u00e7\u00f5es foram realizadas entre tratamento cl\u00ednico otimizado (n = 688), treinamento f\u00edsico supervisionado (n = 1.189), angioplastia (n = 511) e angioplastia mais treinamento f\u00edsico supervisionado (n = 395). O seguimento m\u00e9dio foi de 12 meses. Comparados ao tratamento medicamentoso otimizado isoladamente, a angioplastia e o treinamento f\u00edsico supervisionado superaram todas as outras estrat\u00e9gias terap\u00eauticas, havendo ganho de dist\u00e2ncia m\u00e1xima de caminhada de 290 m ou ganho proporcional de 141% , com per\u00edodo m\u00e9dio de acompanhamento de 12 meses. 378O treinamento f\u00edsico supervisionado isoladamente e a angioplastia associada ao treinamento f\u00edsico supervisionado novamente superaram as demais modalidades de tratamento, com ganho de dist\u00e2ncia m\u00e1xima de caminhada de 110 m ou incremento proporcional de 66% . O treinamento f\u00edsico supervisionado, com aumento de dist\u00e2ncia m\u00e1xima de caminhada de 180 m (IC 95%: 130 a 230 m) e ganho proporcional de 87% (IC 95%: 63 a 111%) foi superior \u00e0 angioplastia isolada, mas inferior ao treinamento f\u00edsico supervisionado associado \u00e0 angioplastia, no quesito dist\u00e2ncia m\u00e1xima de caminhada. 380 Nesse contexto, o treinamento f\u00edsico supervisionado e a angioplastia s\u00e3o fundamentais para melhorar a dist\u00e2ncia de caminhada e a qualidade de vida. Essa recente meta-an\u00e1lise citada sugere fortemente que o treinamento f\u00edsico supervisionado, associado \u00e0 angioplastia, deve fazer parte do tratamento de primeira linha, sempre no contexto da terapia medicamentosa otimizada. A oferta de angioplastia sem treinamento f\u00edsico otimizado deve ser evitada sempre que poss\u00edvel. 378 Por\u00e9m, frequentemente, os centros de tratamento da DAOP oferecem primeiramente a angioplastia, devido \u00e0 car\u00eancia de centros voltados ao treinamento f\u00edsico supervisionado. N\u00e3o se pode negligenciar que o treinamento f\u00edsico supervisionado enfrenta resist\u00eancia por parte do pr\u00f3prio paciente, provocando pouca ades\u00e3o ao tratamento, o que justifica, em parte, a conduta majorit\u00e1ria pelo tratamento percut\u00e2neo. 381Esses estudos de revis\u00e3o t\u00eam implica\u00e7\u00f5es importantes para a pr\u00e1tica cl\u00ednica. Isso porque todos os pacientes com claudica\u00e7\u00e3o intermitente devem receber tratamento cl\u00ednico otimizado, tendo em vista as evid\u00eancias que demonstram redu\u00e7\u00e3o de eventos cardiovasculares futuros e melhora de desfechos relacionados aos membros. 382Entretanto, esses estudos recentes, que investigaram as modalidades de tratamento da DAOP sintom\u00e1tica com treinamento f\u00edsico isolado ou associado \u00e0 angioplastia, t\u00eam demonstrando os benef\u00edcios da combina\u00e7\u00e3o dos tratamentos, o que pode aumentar a probabilidade de que a RCV se torne cada vez mais difundida e acess\u00edvel. Sendo assim, al\u00e9m do tratamento cl\u00ednico otimizado, a angioplastia combinada ao treinamento f\u00edsico supervisionado parece ser a estrat\u00e9gia ideal para tratamento inicial dos pacientes com claudica\u00e7\u00e3o intermitente, tanto para melhorar a m\u00e1xima dist\u00e2ncia de caminhada, como para a qualidade de vida. No entanto, os dados dessas \u00faltimas revis\u00f5es s\u00e3o incapazes de afirmar se primeiramente deve ser proposto o treinamento f\u00edsico supervisionado e, posteriormente, a angioplastia, ou vice-versa. Development:Department of Exercise Testing, Sports Exercise, Nuclear Cardiology, and Cardiovascular Rehabilitation of the Brazilian Society of Cardiology (SBC)Norms and Guidelines Council:Brivaldo Markman Filho, Antonio Carlos Sobral Sousa, Aurora Felice Castro Issa, Bruno Ramos Nascimento, Harry Correa Filho, Marcelo Luiz Campos VieiraNorms and Guidelines Coordinator:Brivaldo Markman FilhoGuideline Coordinators:Tales de Carvalho e Mauricio MilaniContent1. Introduction 9461.1. Strengths (Grades) of Recommendation 9471.2. Levels of Evidence 9472. Structure of a Cardiovascular Rehabilitation Program 9472.1. Staffing and Individual Responsibilities 9472.1.1. Primary Physician 9472.1.2. Lead Physician of Cardiovascular Rehabilitation Program 9472.1.3. Other Health Care Workers 9472.1.4. Physical Therapists and Physical Educators 9472.1.5. Nurses 9472.2. Physical Infrastructure of a Rehabilitation Service 9482.2.1. General Aspects 9482.2.2. Fitness and Exercise Equipment 9482.2.2.1. Aerobic Exercise 9482.2.2.2. Strength Training 9482.2.2.3. Other Exercise Modalities 9482.2.3. Monitoring 9482.2.4. Safety 9483. Phases of Cardiovascular Rehabilitation and Risk Stratification 9493.1. High Clinical Risk 9493.2. Intermediate Clinical Risk 9503.3. Low Clinical Risk 9514. Cost-Effectiveness of Cardiovascular Rehabilitation 9515. Home-Based Cardiovascular Rehabilitation 9526. Integration of Cardiovascular Rehabilitation into Optimized Clinical Care of Cardiovascular Diseases 9536.1. General Guidance for Increasing Physical Activity and Engagement in Physical Exercise 9536.2. Hypertension 9556.2.1. Therapeutic Benefits of Physical Exercise 9556.2.2. Indications for Physical Exercise in Hypertension 9556.2.3. Pre-Exercise Evaluation 9566.2.4. Special Considerations for the Prescription and Follow-Up of Physical Exercise Programs 9566.3. Stable Coronary Artery Disease after an Acute Event or Revascularization 9576.3.1. Therapeutic Benefits of Physical Exercise 9576.3.2. When Is Rehabilitation Indicated? 9576.3.3. Pre-Exercise Evaluation and Exercise Prescription 9586.3.4. Special Considerations for the Prescription and Follow-Up of Physical Exercise Programs 9596.3.4.1. Refractory Angina 9596.3.4.2. Exercise Training with Myocardial Ischemia 9596.3.4.3. Drug Adjustments in Response to Physical Exercise 9596.4. Heart Failure 9606.4.1. Pre-Exercise Evaluation and Exercise Prescription 9606.4.2. Final Considerations on Heart Failure 9626.5. Heart Transplantation 9626.5.1. Benefits of Physical Exercise 9626.5.2. Pre-Exercise Evaluation and Unique Features 9636.5.3. Exercise Prescription 9636.5.4. Home-Based Cardiovascular Rehabilitation 9636.5.5. Recommendations 9646.6. Cardiomyopathies 9646.6.1. Hypertrophic Cardiomyopathy 9646.6.1.1. Therapeutic Benefits of Physical Exercise 9656.6.1.2. When Is Physical Exercise Indicated? 9666.6.1.3. Pre-Exercise Evaluation 9666.6.1.4. Unique Features in the Prescription and Follow-Up of Physical Exercise Programs 9666.6.2. Myocarditis 9676.6.3. Other Cardiomyopathies 9676.6.3.1. Arrhythmogenic Right Ventricular Cardiomyopathy 9676.6.3.2. Noncompaction Cardiomyopathy 9686.7. Valvular Heart Disease 9686.7.1. Pre-Intervention Phase 9686.7.2. Post-Intervention Phase 9686.7.3. Pre-Exercise Evaluation 9696.7.4. Special Considerations for the Prescription and Follow-Up of Physical Exercise Programs 9696.8. Patients with Artificial Pacemakers or Implantable Cardioverter-Defibrillators 9706.8.1. Therapeutic Benefits of Physical Exercise 9706.8.2. When Is Cardiovascular Rehabilitation Indicated? 9716.8.3. Pre-Exercise Evaluation 9716.8.4. Special Considerations for the Prescription and Follow-Up of Physical Exercise Programs 9716.8.5. Resistance Training 9736.8.6. Neuromuscular Electrical Stimulation 9736.9. Peripheral Arterial Occlusive Disease 973References 9751 reductions in hospitalization rate, 2 and significant gains in quality of life, 2 as consistently documented in the literature, including in meta-analyses of randomized clinical trials. These effects justify the consensual, emphatic recommendation of CVR by major medical societies worldwide. 6It is common sense\u2014and has been scientifically proven \u2013 that physical activity helps to preserve and restore the health of both body and mind. The favorable effects of cardiovascular rehabilitation (CVR), with an emphasis on physical exercise, include significant reductions in cardiovascular and overall morbidity and mortality, 8 Conversely, higher levels of physical activity are positively associated with better quality of life and longer life expectancy. 13 In addition, there is a strong, inverse association of the various components of physical fitness with all-cause mortality and with occurrence of adverse cardiovascular events: the lower the level of physical fitness, the higher the mortality rate. 21Sedentary behavior, which is highly prevalent in Brazil and elsewhere, is strongly associated with cardiovascular disease (CVD) and early mortality. 21Therefore, the main objective of CVR, with an emphasis on physical exercise, is to improve the various components of physical fitness, both aerobic and non-aerobic . This requires a combination of different exercise modalities and types of training. In this view, beyond rehabilitation, CVR aims to provide the highest achievable level of physical fitness \u2013 in order to reduce the risk of further cardiovascular events \u2013 and to promote all of the other benefits derived from regular physical exercise. 23 CVR is underutilized worldwide. In Brazil, considering the size of the country and the diversity of its population, several barriers limit access to RCV, 25 such as a scarcity of structured services, limited urban mobility, and high rates of violence in cities. 27 Within this context, so-called home-based cardiovascular rehabilitation (HBCR) programs, in which most sessions take place in the patient\u2019s home under indirect supervision, can supplement or serve as an alternative to traditional programs in which training sessions are always carried out under direct supervision.However, despite its documented benefits and excellent cost-effectiveness, 31 this guideline will exclusively address interventions based on physical exercise. The strength (or grade) of recommendation will always be proportional to the level of evidence available, as explained below.As in previous documents on this topic published by the Brazilian Society of Cardiology, Grade I: there is conclusive evidence, or, failing that, a consensus that the procedure is safe and useful/effective;Grade II: there is conflicting evidence and/or divergent opinions on the safety and utility/effectiveness of the procedure:\u2013 Grade IIA: weight of the evidence/opinion is in favor of the procedure. Most experts approve;\u2013 Grade IIB: safety and utility/effectiveness are less well established, with no predominance of opinions in favor.Grade III: there is evidence and/or expert consensus that the procedure is not useful/effective and, in some cases, can even be harmful.Level A: data obtained from multiple, large, concordant randomized studies and/or robust meta-analyses of randomized clinical studies;Level B: data obtained from a less robust meta-analysis, based on a single randomized trial or on non-randomized studies;Level C: data obtained from consensus expert opinions.32The makeup of CVR teams must be adjusted to its objectives, target audience, and availability of human and material resources, taking into account regional characteristics, the modality of rehabilitation (direct or indirect supervision), and the site or setting . A multidisciplinary CVR team is usually composed of physicians, physical educators, physical therapists, and nurses, but may also include other professionals, such as dietitians, psychologists, and social workers. 31CVR is an integral part of the optimized clinical treatment of patients with stable CVD. Thus, it is essential that the CVR team and the patient\u2019s primary physician work in an integrated manner. When referring a patient for rehabilitation, primary clinicians must be aware of its indications and potential benefits and should carry out the necessary pre-exercise evaluation. As this integration will involve frequent progress reports, potential needs for adjustment of drug therapy, awareness of complications and intercurrent events, etc., it is very important that mechanisms be established to facilitate communication between the patient\u2019s primary physician and the CVR team. 34The lead physician coordinates all medical activities. In Brazil, this role usually falls to the general coordinator of the CVR program. He or she must have in-depth subject knowledge of CVR and be trained to act in cardiovascular emergencies. Some of the main activities of this position include:31Perform pre-exercise evaluation, including cardiopulmonary exercise testing as needed, to provide inputs for the initial prescription of CVR training sessions. Train the CVR team to identify high-risk situations and provide appropriate care in emergencies.Establish restrictions and set limits for the exercise prescription.Lead and interact with other team members, to optimize the quality and safety of exercise prescription.Schedule follow-up evaluations, always in coordination with the patient\u2019s primary physician.31Like physicians, the other members of the team, when carrying out their respective duties, must follow the program\u2019s rules and guidelines as well as the formal recommendations of their respective professional associations. Physical therapists and physical educators are directly involved in the prescription and supervision of physical exercise, within the targets and limits defined by the physician as a result of the pre-exercise evaluation and follow-ups. They must have specific knowledge of CVD and exercise physiology and training in basic life support, including the use of an automated external defibrillator. Such training must be periodically refreshed to ensure continued competence. In addition to their direct role during CVR sessions, these professionals can provide patient guidance and contribute to other lifestyle measures aimed at adopting healthy habits.In a CVR program, nurses and other nursing professionals can assist in clinical evaluation and obtain and provide information on the patient\u2019s medical status, including through contact with family members. Nurses can also be in charge of blood glucose measurements and blood pressure (BP) checks before and/or during exercise sessions. In case of complications or intercurrent events, nurses can provide direct care, assist the physician and administer medications. Nurses must, of course, also be trained in basic life support, including the use of an automated external defibrillator.2.2. Physical Infrastructure of a Rehabilitation Service29A CVR program can be run out of various types of facilities, depending on its objectives and on the available resources. Most often, CVR programs are carried out indoors, in air-conditioned environments, although exercise sessions can be held in outdoor venues such as running tracks, courts, gymnasiums, parks, or public recreation spaces. 2 to a several hundred square meters. Proper changing rooms and restrooms are essential. To minimize the risk of accidental falls, slip-resistant flooring is mandatory. 29Indoor venues should be adequately sized and appointed, with dimensions and characteristics varying according to local resources and service capacity. The space should be large enough for patients to exercise in, ideally with a ceiling height not less than 2.5 meters. It should also be properly lit, well ventilated, and climate-controlled so as to maintain a temperature of 22\u201325\u00b0C and a relative humidity of 40\u201365% during sessions. The exercise area per se, not considering changing rooms, restrooms, and the reception area or waiting room, varies greatly \u2013 from 20 m 2.2.2.1. Aerobic Exercise29The most commonly used aerobic exercise equipment are treadmills and stationary bicycles (cycle ergometers), but upper-body ergometers, rowing machines, cross-country ski machines, and elliptical trainers can be used, among countless others. Treadmills must be electric, with a nominal capacity of at least 100 kg body weight, front and side supports for the hands, and a safety key. They must also allow individualized adjustments of speed and slope over a wide range. Cycle ergometers can be mechanical or electromagnetically braked. There are specific models for the upper body, and even some models which allow all four limbs to be exercised simultaneously. Conventional (lower body) models may be upright or recumbent. Ideally, the cycle ergometer should provide a readout of cadence or speed and, most importantly, power (in watts). Some cycle ergometers allow the user to program the intensity directly in watts, so that the resistance of the pedals increases when the cadence decreases and vice versa.Rowing machines, ski machines, and elliptical trainers can be particularly useful for patients with a lower degree of functional limitation or who have had previous experience with such equipment. These devices provide the advantage of allowing simultaneous exercise of the upper and lower limbs.2.2.2.2. Strength TrainingSeveral types of equipment can be used for strength training. Bodyweight exercises, which require no equipment at all, are often sufficient in the most debilitated patients. One representative and functional example is rising from a seated position, which requires only a chair or bench.29Ropes or straps, firmly attached to the ceiling or high on a wall, can also be used to facilitate a wide range and variety of bodyweight exercises. Free weights, dumbbells, or ankle weights are often adopted in CVR programs, as they allow patients to execute a wide range of movements and provide appropriate stimulation of different muscle groups. Specific devices consisting of weights connected to cables and pulleys, known as cable machines or stack machines, can also be used. Other useful equipment includes workout bars, weighted exercise balls , stability balls , and elastic bands or tubes with varying degrees of resistance. During all exercises, attention must be paid to proper posture and execution of the prescribed movements, so as to prevent musculoskeletal injuries. Attention when handling exercise equipment is also important to prevent accidents and potential injury.2.2.2.3. Other Exercise ModalitiesWith a view to overall health, considering heart disease and associated conditions, patients may benefit from or even require other types of exercise, such as isometric handgrip training, inspiratory muscle training, and exercises designed to improve balance and flexibility.Several modalities are available for patient monitoring, including heart monitors, mobile applications for monitoring heart rate (HR), glucometers, pulse oximeters, and conventional devices such as sphygmomanometers and stethoscopes. Depending on the clinical complexity and the risk of unfavorable cardiovascular events, continuous electrocardiographic monitoring (at rest or during exercise) may be convenient. This can be achieved by conventional ECG (connected directly to the patient) or by telemetry systems. Rapid access to monitoring equipment is of fundamental importance for proper detection and subsequent management of potential cardiovascular events.Although serious cardiovascular events \u2013 such as cardiac arrest, which, in most adults, results from ventricular fibrillation or pulseless ventricular tachycardia \u2013 are extremely uncommon during CVR, it is essential that all programs have a plan in place to respond appropriately to these events if they do occur. Therefore, a defibrillator is mandatory safety equipment. Other basic and advanced life support supplies must also be available, such as laryngoscopes, orotracheal tubes of various sizes, masks, a bag-valve-mask manual resuscitator, and supplemental oxygen.36For more detailed guidance on techniques, equipment, and recommended drugs, readers are advised to check subject-specific guidelines. 31Traditionally, CVR is divided into time-bound phases, with phase 1 occurring in hospital and phases 2 to 4 in the outpatient setting. In the early days of CVR, phase 1 was intended for recovery after acute myocardial infarction (AMI) or coronary artery bypass surgery (CABG). Subsequently, in the context of what is now known as cardiopulmonary and metabolic rehabilitation, phase 1 was expanded to include hospitalized patients who underwent percutaneous coronary intervention (PCI), valve replacement or repair surgery, procedures for congenital heart disease, and heart transplantation (HTx), in addition to those with heart failure (HF), coronary artery disease (CAD), diabetes, hypertension, and chronic lung and kidney disease . Therefore, CVR should begin immediately once the patient is considered clinically stable as a result of clinical and/or interventional treatment. 31In phase 1 of CVR, the aim is for the patient to be discharged from the hospital in the best possible physical and psychological condition and with guidance to pursue a healthy lifestyle, especially with regard to physical exercise. A combination of low-intensity physical exercise, techniques for stress control, and education on risk factors and heart disease is recommended. The team must be composed of at least one physician, one physical therapist, and one nurse. All should be trained specifically in CVR, but full-time dedication to the rehabilitation program is not required; team members are free to perform other duties in the hospital. Upon discharge from hospital, patients must be referred to the outpatient phases of CVR. Phase 2 begins immediately after hospital discharge and lasts, on average, 3 months. Phase 3 usually lasts 3 to 6 months, and phase 4 is quite prolonged. In all phases, the goal is to obtain progressive benefits from RCV or at least maintain any gains obtained.31A strict division of CVR into time-bound phases may fail to take into account that some very symptomatic, debilitated patients with severe heart disease will remain in long-term \u201cphase 2\u201d rehabilitation, as they continue to require direct supervision of physical exercise, while other low-risk patients may be fit for phase 3 or even phase 4 programs straight away, and are thus candidates for home-based CVR . Therefore, stratification of clinical risk is recommended to enable a more rational use of CVR programs, with individualized targeting of phases and modalities. In this context, high-risk patients, those with less physical capacity, and those most symptomatic should participate in supervised sessions indefinitely, while those at lower risk, with greater physical capacity and fewer or less severe symptoms can engage in a wider range of more intense exercises without direct supervision .37 while the cut-off points for this classification are based on expert opinion (level C evidence), and can thus be modified according to the experience of the CVR team and the discretion and judgment of the clinician in response to the pre-exercise evaluation and subsequent evaluations , and cardiopulmonary exercise test (CPET) or treadmill exercise test (TMET).High-risk patients will often need immediate or short-term medical attention during CVR . Therefore, they require closer monitoring by the team, which must be able to identify signs and symptoms of high-risk events and act immediately if such an event arises, providing basic and advanced life support, including with use of a manual or automated external defibrillator as necessary. It is preferable that this equipment be present in the room at all times. The medical team must be readily available on site to attend to the patient in the event of any serious complications.It should be noted that the best way to prevent cardiovascular events during a rehabilitation program, and especially after events and interventions, is to conduct systematic pre-exercise evaluation and revaluations.The exercise program must be individualized in terms of intensity, duration, frequency, training modality, and progression, according to the functional testing performed at the start of the program and subsequently. Proper measurement of HR and BP at rest and during exercise are mandatory; measurement of oxygen saturation, capillary blood glucose, and electrocardiographic monitoring should also be available.The rehabilitation program should also include an educational program aimed at lifestyle modification, with an emphasis on dietary re-education and strategies for smoking cessation, if necessary. It is essential that patients acquire knowledge about their illness and learn to self-monitor, both while exercising and in terms of red-flag signs and symptoms which may signal unstable or high-risk clinical situations.The clinical characteristics of patients who would initially be classified as high clinical risk (presence of at least one such feature) are:Hospitalization due to recent (< 8\u201312 weeks) cardiovascular events: AMI or unstable angina; surgical or percutaneous revascularization; complex arrhythmias; cardiac arrest; acute decompensated HF.Chronic heart disease with or without recent cardiovascular events and/or interventions but with significant functional changes on physical exertion, i.e.:2 peak < 60% of predicted for age and sex).\u2013 Low functional capacity on TMET (< 5 MET) or CPET (Weber C/D classification or VO 2 of < 15 ml.kg -1 .min -1 ).\u2013 Signs and symptoms of myocardial ischemia at low loads .Other clinical characteristics of patients at increased risk during physical exercise include dialytic chronic kidney disease (CKD), oxygen desaturation on exertion, and complex ventricular arrhythmias at rest or exertion.Considering that high-risk patients often need frequent readjustment of drug therapy and reassessment and occasionally need advanced intervention (revascularization or other procedures), constant communication between the CVR team and the patient\u2019s primary physician(s) is essential. It is also important to note that some patients who experience intercurrent events during exercise or unfavorable findings on follow-up evaluations may remain in the high-risk classification indefinitely.Patients may have completed previous stages of CVR and been reclassified; may be classified directly into this category despite no previous engagement in exercise; or may have been referred from other exercise programs. The duration of CVR under this classification can also be variable, depending on the clinical status and progress achieved with physical training as demonstrated in follow-up evaluations.Exercises should be supervised by a physical therapist or physical educator, and the service should rely on the coordination of a physician with experience in CVR. Devices for measurement of HR and BP at rest and during exercise are recommended; measurement of oxygen saturation, capillary blood glucose, and electrocardiographic monitoring should also be available as necessary.If there is no on-site physician, one must be readily on call. Basic life support material must be available on site and all team members must be trained in cardiopulmonary resuscitation, including use of an automated external defibrillator, which must also be present on site.It is essential that pre-exercise evaluation be carried out by the CVR team, with appropriate risk stratification. Regular medical follow-up and revaluations as necessary must be carried out to ensure the safety of the exercise regimen.The clinical characteristics of patients who would initially be classified as intermediate risk (presence of at least one such feature) are:Longer than 12 weeks since the latest cardiovascular event or intervention and currently stable clinical condition.Chronic heart disease with some loss of function on exertion:2 peak 60\u201385% of predicted for age and sex).\u2013 Moderate functional capacity on TMET (5\u20137 MET) or CPET .Any other clinical features judged by the physician responsible for pre-exercise evalution to pose intermediate cardiovascular risk on exertion.The main objective of CVR in patients with this risk profile remains the improvement of physical fitness, both aerobic and non-aerobic , as well as superior disease control. The need to promote wellness and improved quality of life, in addition to other procedures that contribute to reducing the risk of clinical complications, such as strategies for smoking cessation, dietary reeducation, and weight control, should all be considered. An emphasis on maintaining and adhering to prescribed drug therapy is also essential to preventing the progression or instability of CVD. Acquisition of knowledge about the disease itself, allowing for better self-management, increases the accuracy in identifying signs and symptoms of disease progression or red flags of unstable clinical situations, which may require interruption of the exercise program and medical revaluation.Patients in this category, after an initial period of guidance and knowledge acquisition about physical exercise and self-monitoring, may be able to adapt to home-based CVR, in which physical exercise is performed under the indirect supervision of team members. Such supervision, as well as adjustments to the exercise prescription or patient education to clarify any questions, should take place during periodic face-to-face or remote follow-up sessions.Patients may have completed previous stages of CVR and been reclassified; may be classified directly into this category despite no previous engagement in CVR; or may have been referred from other physical exercise programs. Training for these patients is a long-term endeavor, aimed at maintaining overall health and achieving the greatest possible gains in physical fitness, with the objective of reaching the highest attainable standard of health.Depending on availability and individual preferences, exercises can be carried out under direct (face-to-face) or remote supervision. However, given their lower clinical risk and reduced need for supervision, patients at this stage are a perfect fit for home-based rehabilitation models, so that the CVR team can devote on-site resources to patients at higher clinical risk.Patients must nevertheless undergo periodic reevaluation by their primary physician and by the CVR team, including CPET or TMET. In principle, the time between follow-up revaluations should not exceed 12 months. The purpose of follow-up is to readjust the exercise prescription and identify any deterioration of the underlying disease or red flags for clinical decompensation or cardiovascular events, thus allowing preemptive adjustment of drug therapy and/or surgical or percutaneous intervention as needed.Patients receiving home-based programs should be periodically reevaluated and receive guidance on exercise. These occasions are advised to serve as opportunities for participation in supervised exercise sessions, especially for less-experienced patients, as well as for readjustment of the exercise prescription as needed and to answer any questions. Periodic remote assessment by the CVR team , at least once every 6 months, is recommended to encourage continued adherence to the physical exercise program.The clinical characteristics of stage 4 patients are :Longer than 6 months since the latest cardiovascular event or intervention and currently stable clinical condition.Chronic heart disease with little or no loss of function on exertion.Patients in this classification usually exhibit the following:2 peak > 85% of predicted for age and sex).\u2013 Good functional capacity on TMET (> 7 MET) or CPET are considered to have excellent cost-effectiveness, whereas those that require investments of USD 20,000\u201340,000/LYS are acceptable and those requiring investments of USD > 40,000/LYS are unacceptable.Assessment of cost-effectiveness, which is done through a combined analysis of clinical consequences (effectiveness) and health-system expenditures, is essential in evaluating the relevance of large-scale implementation of a given treatment. 23 published the first systematic review of studies on the cost-effectiveness of CVR as a secondary prevention strategy in patients with CAD and HF. In a 2018 evaluation of studies published after 2001, 54 the cost-effectiveness ratio was very similar to that described by Papadakis; the addition of CVR to conventional treatment resulted in an increase in expenditure of USD 2,555 to USD 23,598 per LYS.According to 1985\u20132004 data, CVR can be considered an intervention with an excellent cost-effectiveness ratio, as its addition to conventional treatment resulted in an increase in expenditures from USD 2,193 to USD 28,193 per LYS. In 2005, Papadakis et al. 55 there is a lack of data on the cost-effectiveness of CVR in these settings. 56 Most of the information available comes from high-income nations, such as the United States, Canada, and European countries, which hinders extrapolation of results to the Brazilian reality. However, those few studies which are available from lower-income nations show a similar trend. In Brazil, the addition of rehabilitation to conventional treatment of patients with HF resulted in an increase in expenditures of USD 21,169 per LYS. 57It is worth mentioning that, although more than 75% of CVD deaths occur in low- and middle-income countries, 59 In fact, most Brazilian states \u2013 including most capitals and large cities \u2013 lack even a single cardiac rehabilitation service.Despite the clear clinical and economic benefits of CVR, the percentage of eligible patients who effectively receive this type of care is far short of desired levels. According to international data, only around 30% of patients attend a CVR program; in Brazil, this number is estimated to be well below 15%. 62 in addition to advances in technology that now allow remote monitoring, the use of HBCR has been expanded to patients with a higher risk profile.As a result, the use of home-based models has grown. Initially, concerns about the safety of physical exercise meant that HBCR was intended only for low-risk patients. However, with growing evidence of noninferiority in terms of safety and similar clinical benefits in relation to the conventional strategy, 60 who compared the effects of a 3-month program of either model in low- and moderate-risk CAD patients after an acute coronary event. Although the group of patients who attended the traditional program performed a higher volume of exercise, there was no difference in increase in functional capacity or quality of life between the two groups. Jolly et al. 62 compared cardiovascular risk outcomes between patients undergoing traditional and home-based rehabilitation for longer periods, with 6, 12, and 24 months of follow-up, and also observed no differences.Recent studies show that HBCR has effectiveness similar to traditional CVR, as demonstrated by Ades et al., 61 also found no significant differences between conventional and home-based rehabilitation across a series of outcomes in the short term (3 to 12 months) or long term (up to 24 months).A recent systematic review by Anderson et al. of studies enrolling patients with a history of AMI, CABG, or HF 64 This major research gap precludes comparison of the two models in terms of cost-effectiveness. 67Thus, HBCR programs should be considered as a strategy to facilitate access, adherence, and wider use of rehabilitation. Despite the aforementioned evidence of noninferiority in outcomes, comparatively few studies have demonstrated that the cost of HBCR is comparable to that of traditional CVR programs. Given the facts, it is unsustainable that countries of all income levels \u2013 and, most worryingly, lower-incomes \u2013 continue to provide high-cost therapeutic interventions massively, without stricter indications and criteria, while they continue to neglect the highly effective, economically viable, and readily applicable intervention that is CVR. There is an urgent need for public health policies to expand the availability of, participation in, and adherence of eligible patients to both traditional and home-based CVR programs.Finally, considering the relevance of CVR given its broad clinical benefit and excellent cost-effectiveness, strategies must be implemented to change medical culture and stances toward it and facilitate the dissemination of structured rehabilitation programs. In this context, it is particularly relevant that specialty cardiology services offer CVR to their patients both during hospitalization and after discharge. The availability of a CVR service should be considered as a mandatory prerequisite for a medical institution to be recognized or accredited as having excellence in cardiology.68 which, compounded by the scarcity of referral to CVR programs and the limited availability of services, lead to very low levels of actual participation in supervised exercise programs. In this context, programs of indirectly supervised exercise carried out in the home, known as home-based cardiovascular rehabilitation (HBCR), are an attractive alternative or supplement to conventional, on-site CVR. Given its inherently greater availability, HBCR should be considered the main modality of CVR intervention when it comes to public health strategies, aiming at mass engagement of the CVD population in rehabilitation programs.There are several barriers to patient access and adherence to conventional CVR, 61 of 23 studies including 2,890 patients with heart disease compared the effects of conventional CVR and HBCR. No differences were found in mortality, physical capacity, or quality of life. Therefore, the decision to participate in conventional (on-site) or home-based programs depends on the availability of services and patients\u2019 individual preference.A Cochrane review HBCR is understood as the practice of physical exercise without face-to-face supervision, but with guidance and follow-up from the CVR team. It is thus also known as semi-supervised rehabilitation, or rehabilitation under indirect or remote supervision. The indications and objectives of HBCR are the same as those of conventional CVR; the same care is required regarding pre-exercise evaluation and exercise prescription. Most sessions are held under indirect supervision, but participation in some on-site classes, especially at the start of the program, is of fundamental importance to ensure that patients understand the exercise prescription, consolidate guidance and clarity doubts. Exercise can be done at home, in parks, on public roads, gyms, sports centers, and health clubs, among others, with patients self-monitoring and following the guidance received.Therefore, in order to achieve HBCR as a viable population strategy, it is first necessary to expand the availability and capacity of conventional CVR programs, in order to enable initial evaluation, guidance, exercise prescription, and follow-up of home sessions (with periodic reassessment for any adjustments). Thus, the home strategy must be aligned with that of conventional CVR. The two modalities may be used in parallel, including patients with different risk profiles, or in sequence, with the same patient engaging in conventional or home-based CVR depending on clinical status.Just as in conventional CVR, the first step of HBCR is referral by the primary physician, followed by evaluation by the rehabilitation physician and other team members, ideally including a stress test (CPET or TMET) or other physical fitness tests. After the pre-exercise evaluation, patients defined as high risk can be prioritized for conventional, face-to-face CVR. Those at lower risk, who are capable of self-monitoring and according to individual preference, can be routed to the HBCR component of the program. After receiving instruction on the prescribed exercises, patients perform the sessions on their own. The exercises may be documented in printouts or spreadsheets, with the aid of resources such as cardiac monitors, pedometers, or fitness trackers. Smartphone apps can mediate this exchange of information between patients and the health care team.In some cases, a combined CVR program \u2013 with both on-site and home-based sessions \u2013 may be an option for moderate-risk patients who are still learning to self-monitor or find it difficult to attend face-to-face sessions due to social issues or reduced mobility. The proportion of on-site versus home-based sessions will vary according to the patient's clinical characteristics and the logistics and infrastructure of the CVR service.The overall focus is always to make patients more physically active; a reduction in sedentary behavior and its harmful consequences is the imperative. It is essential that all available resources \u2013 whether alone or in combination, whether informal physical activity, home rehabilitation, or conventional CVR \u2013 be deployed toward this goal.70CVR must be integrated with full clinical treatment of CVD, which consists of a synergistic combination of structured lifestyle changes and optimized drug therapy. For instance, in patients with stable coronary disease \u2013 even those with moderate and severe ischemia \u2013 interventional treatments have not been shown to be superior in reducing major outcomes . 73 In this context, it may be necessary to adjust current drugs or prescribe additional agents prior to the start of the physical exercise program. Once CVR has been initiated and adequate adherence has been achieved, some patients may require dose reduction or even discontinuation of some drugs due to adaptations to physical training, e.g., patients who develop hypotension, marked bradycardia, or symptomatic hypoglycemia. 75To increase the efficacy and safety of CVR, it is important that drug therapy of CVD be properly optimized. The aim is to increase exercise tolerance and thus facilitate engagement in physical exercise while reducing the risk of further events. 76 Therefore, for health promotion and CVD prevention, medical guidelines have recommended the practice of moderate-intensity physical activity for at least 150 minutes per week or high-intensity for at least 75 minutes per week (grade 1B recommendation). 83 Engagement in more than 300 minutes/week of moderate- to high-intensity exercise can confer additional benefit, as has already been demonstrated in patients with CAD. 84There is an association between sedentary lifestyle , and higher all-cause and cardiovascular mortality. According to the results of individual evaluation, the exercise prescription may vary in terms of type , modality and duration; weekly frequency and intensity should also be considered and 3.Sedentary patients should start exercising at the lower limit of the exercise prescription and progress gradually over the following weeks. Progression should initially be based on the duration of each session and, later, on exercise intensity. Physically active patients can perform exercises at a more intense level from the outset, aiming at a minimum duration of 75 minutes divided into two or more weekly sessions.Resistance training of localized muscle groups (whether strength or endurance training) has proven quite beneficial for overall health and for the cardiovascular and musculoskeletal systems, and is particularly important \u2013 in fact, essential \u2013 in patients with sarcopenia and/or osteopenia. Resistance training should be performed at least twice a week, favoring the large muscle groups of the upper limbs, lower limbs, and core. Exercise can be done against the individual\u2019s own body weight or using implements such as free weights, ankle weights, bands, or weight machines. The load (or weight) for each exercise or movement must be individually adjusted, and particular attention should be paid to proper posture and technique.Several protocols for resistance training are available, with variations in parameters such as the number of exercises per session , the number of sets per exercise , and the number of repetitions per set (6 to 20). The intensity of resistance training can be adjusted according to the relative intensity of the maximum force, and can be expressed as a function of the maximum load that can be borne during a single repetition (one repetition maximum test or 1RM). Light intensity would be up to 30% of 1RM; medium intensity, between 30 and 60\u201370% of 1RM; and high intensity, above 60\u201370% of 1RM. Resistance exercises may also be prescribed subjectively, on the basis of perceived exertion alone see .A practical approach is the variable repetition method, which aims to perform a range of repetitions . If the patient is unable to perform the movement correctly for the minimum number of repetitions prescribed, the applied load is too high. On the other hand, if the patient reaches the maximum prescribed number of repetitions easily, the load is insufficient. Thus, the load will be adjusted so that training takes place within the proposed range of repetitions. This method can be applied to a wide range of localized exercises and can be altered as the patient progresses; the repetition ranges can also be modified depending on the desired objective .Flexibility exercises can also provide musculoskeletal benefits, improve health-related quality of life, and prevent falls in the elderly. By facilitating and increasing the efficiency of joint movement, they reduce oxygen demand during motion, thus enhancing cardiovascular performance. The aim of these exercises is to reach the maximum range of motion (point of mild discomfort) and remain static for 10 to 30 seconds.86Depending on the age group, clinical condition, and objectives of the exercise program for a given patient, other types of exercise can be included in the prescription, such as motor coordination and balance exercises. The countless benefits of more playful, social-based forms of exercise, such as dance and other modalities, should also be considered. 87 Such adherence can also provide clinical and functional inputs that enable adequate counseling as to whether and how patients can resume sexual activity, based on the KiTOMI model, which was proposed by Brazilian authors in 2016. 88 In addition, periodic reevaluation is essential to encourage commitment and measure the progress and benefits obtained.Assessment of aerobic and non-aerobic physical fitness enables a more individualized exercise prescription, with the objective of achieving the best outcomes while minimizing hazard to the patient through proper risk stratification and a thorough search for possible abnormalities. In general, the initial evaluation is based on a thorough history, physical examination, and ECG. More detailed assessments should be individualized to include CPET or TMET, anthropometric measurements, and evaluation of muscle strength/endurance and flexibility. The initial evaluation allows quantification of the patient\u2019s functional deficit in relation to the desired level of function, as well as goal setting. Even those patients with poor baseline physical fitness can benefit from adequate adherence to a supervised exercise program. Finally, it is important to establish a systematic follow-up revaluation scheme which, in addition to encouraging patient commitment, will allow measurement of the progress and benefits obtained and yield reports to support treatment adjustments. Therefore, these reports should always be sent to the patient\u2019s primary physician, who is obviously an integral part of full clinical treatment.89 Worldwide, the number of patients with HTN rose from 594 million in 1975 to 1.13 billion in 2015, with growth largely credited to underdeveloped and developing countries. 90 Considering that most cases of HTN are lifestyle-related, with sedentary behaviors as a prominent cause, the importance of physical exercise is clear, alongside other behavioral measures and drug therapy as indicated. 72Hypertension (HTN) remains one of the leading risk factors for the development of CAD, HF, CKD, and ischemic or hemorrhagic stroke, representing a huge social and economic challenge to global public health. 91 The regular practice of physical exercise has a therapeutic effect on the physiological restructuring of these systems, reducing oxidative stress and inflammation, correcting baroreflex dysfunction, increasing vagal tone, decreasing sympathetic activity, reversing hypertrophic arteriolar remodeling in exercised tissues, and reducing peripheral vascular resistance, with a consequent decrease in BP levels similar, or even superior, to that provided by drug therapy. 93HTN has a complex, multifactorial pathophysiology involving structural and physiological modifications, particularly to the vasculature , kidneys , and nervous system . 94 All of these structural changes in the vessel wall, which occur in both arteries and arterioles, increase the overall stiffness of the vasculature, with a consequent increase in pulse wave velocity, pulse pressure \u2013 the difference between systolic BP (SBP) and diastolic BP (DBP) \u2013 and hydrostatic pressure in the capillaries. These structural imbalances are compounded by endothelial dysfunction, with an increase in vasoconstrictive compounds, inflammatory mediators, and oxidizing agents, to the detriment of synthesis of vasodilating and antioxidant compounds. 95In vascular tissue, HTN is characterized by disorganization of smooth muscle cells, increased collagen deposition, and a decreased elastin/collagen ratio, in addition to the formation of abnormal elastic fiber and internal elastic lamina with a smaller fenestrated area. 98 Experimental studies 94 in spontaneously hypertensive rats have demonstrated reorganization of all vascular structures of the aorta after implementation of a period of aerobic exercise. Aerobic training promotes vascular adaptations in the conductance arteries , arterioles , and capillaries, stimulating angiogenesis. 100Physical exercise, by increasing the tangential stress derived from the friction of blood flow on the endothelial surface of the vessel wall (commonly described by the term shear stress) positively stimulates the endothelial tissue, increasing production of antioxidant enzymes and vasodilating agents, in addition to decreasing the action of free radicals, pro-inflammatory cytokines, adhesion molecules, and vasoconstricting agents, thus restoring the balance of endothelial function. 101 The antihypertensive effect of exercise is comparable to that of medication, 102 and both can be additive, occasionally requiring adjustments of drug dosage.Thus, physical exercise has multifactorial effects on HTN, and is considered a key intervention to mitigate the burden of the disease and its comorbidities. The greatest evidence of benefit in BP reductions among hypertensive patients is for aerobic physical exercise, as corroborated in a meta-analysis by Cornelissen et al. which showed an average SBP reduction of 8.3 mmHg and DBP reduction of 5.2 mmHg as a result of aerobic exercise.103 ) is to preserve or increase muscle mass, strength, and endurance, factors that decrease the relative intensity needed to perform the activities of daily living, with consequent damping of the blood pressure response to exertion. Furthermore, resistance training may also promote improvement in baroreflex sensitivity. 104The goal of resistance training exercises and shown significant effects in reducing BP levels. 109 Physical fitness, measured objectively through graded stress tests, attenuates the increase in BP with age and prevents the development of HTN. In a cohort of 20-to-90-year-old men who were followed for 3 to 28 years, greater physical fitness decreased the rate of BP increase over time and delayed the onset of HTN. 110 Epidemiological studies have revealed that both level of physical activity and cardiorespiratory fitness are inversely associated with hypertension. 112Higher levels of physical activity have been associated with a decrease in the risk of developing HTN. With the advent of electronic activity trackers and ambulatory BP monitoring, it has become increasingly feasible to conduct studies that correlate physical activity with BP. 112 In addition, the continuous practice of physical activities can be beneficial for both the prevention and the treatment of hypertension, further reducing cardiovascular morbidity and mortality. Demonstrating this, active individuals have up to a 30% lower risk of developing hypertension than sedentary ones, 111 and increasing daily physical activity significantly reduces BP. 113Large randomized controlled trials and meta-analyses have confirmed that regular exercise can reduce BP levels. 114 as it is the most prevalent of the cardiovascular risk factors and one of the leading factors contributing to mortality worldwide. 115 Survival is lower among people who spend most of their time sitting than in those who spend little time in this position. 116 Television viewing time is directly associated with high BP levels and cardiovascular morbidity and mortality; 117 therefore, to reduce time spent in the seated position, standing for at least 5 minutes for every 30 minutes spent sitting is recommended as a valid preventive measure. Physical exercise is indicated for all patients with HTN or ventilatory thresholds (CPET) to determine the target training zone.A CPET or TMET should be performed during pre-exercise evaluation, especially if there is suspicion of heart disease, target organ damage, or presence of three or more risk factors. The exercise recommendation for hypertensive patients, is similar to that proposed for the general population: at least 150 minutes per week (five 30-minute sessions) of moderate-to-intense aerobic activity. In addition, two to three resistance training sessions per week are advisable. For greater benefit, absent any contraindications, patients may gradually increase their engagement towards a goal of 300 min/week of moderate aerobic exercise or 150 min/week of intense aerobic exercise.37 or to reduce training intensity until better BP control is achieved. In supervised CVR programs, these recommendations are flexible and can be adjusted individually at the discretion of the rehabilitation physician and according to the BP response observed during the stress test and exercise sessions. During exercise, it is recommended that BP remain below 220/105 mmHg. If BP exceeds this level, the session should be halted or the load reduced, and adjustment of drug therapy should be considered. 37During training, it is important that BP be assessed at rest and in exertion. Patients with a resting BP higher than 160/100 mmHg or with target organ damage are advised to optimize antihypertensive therapy for better BP control before starting or resuming exercise, 120 This acute effect of physical training can cause symptomatic hypotension once the session ends, which usually improves with rest and hydration. Patients on alpha blockers, beta blockers, calcium channel blockers, and vasodilators may be at increased risk of post-exercise hypotension, and thus require special attention during the cooldown period. If post-exercise hypotension becomes recurrent, which usually results from an add-on antihypertensive effect of training, the need for dose adjustments or even discontinuation of medications must be considered.BP must be measured after each exercise session, and is commonly found to be lower than before the start of activities. In hypertensive patients, the acute antihypertensive effect of a single session tends to be greater with more intense levels of aerobic exercise. 121 Although such effects need to be reproduced in further studies, exercise in warm water appears to be appropriate for patients with resistant hypertension.There is little data regarding the effect of exercise in patients with resistant hypertension, which is characterized by BP above target despite the use of three or more antihypertensive medications. In these patients, who require closer monitoring, a randomized, single-center clinical trial showed that exercising in warm water (30 to 32\u00baC) resulted in a pronounced reduction in BP (36/12 mmHg) after 3 months. 124 The underlying mechanisms of stable CAD include atherosclerotic obstruction of the epicardial vessels, microvascular disease, and coronary spasm, either alone or in combination. 5 Clinically, the most common manifestation of stable CAD is angina pectoris, which is characterized by reversible episodes of chest pain due to myocardial ischemia, resulting from the imbalance between myocardial oxygen supply and consumption, usually triggered by physical exertion or emotional stress, which resolve with rest or the administration of fast-acting nitrates. 5Cardiovascular disease (CVD), led by coronary artery disease (CAD), is responsible for the majority of deaths in the adult population worldwide. 125 Nonetheless, full clinical treatment is essential, including optimization of drug therapy and regular physical exercise, in addition to other behavioral changes to address smoking, diet, and body composition. Elective revascularization may also be indicated in patients with stable CAD, depending on their symptoms and cardiovascular risk. 5 However, it is worth noting that, in stable patients, even those with angina, exclusively clinical treatment has not been shown to be inferior to treatment with the addition of an interventional approach. 127Stable CAD has a good prognosis, with annual mortality estimated at 1.5% and a nonfatal infarction incidence of 1.4%. 131Development of an acute coronary syndrome, with AMI or unstable angina, is associated with increased cardiovascular risk and may require adjustment of drug therapy plus urgent surgical or percutaneous revascularization. 136 improvement of cardiorespiratory functional capacity, 136 and improvement in myocardial perfusion imaging. 140 These benefits persist as long as regular physical exercise is maintained, which contributes to improvement in quality of life 146 and reduction of hospitalization and mortality from cardiovascular causes. 148The short- and long-term beneficial effects of regular physical exercise in patients with stable CAD have been demonstrated in the scientific literature. During the first 8 to 12 weeks of CVR, there is a marked increase in ischemic threshold, 144 Myocardial perfusion increases due to an improved endothelium-dependent vasodilator response 151 and increased recruitment of collateral vessels during exercise, 152 which is reflected in the reduction of ST segment depression during exercise. 137 It is also notable that the combination of physical training and a low-fat diet can influence the progression of atherosclerotic plaque. 153In patients with stable CAD, different mechanisms explain the increase in ischemic threshold, which gradually allows physical activity at higher loads. Reduction of the double product at submaximal loads is associated, among other mechanisms, with an improvement in cardiac autonomic modulation. 1 of 63 studies involving 14,486 patients aged 47\u201371 years revealed that CVR reduced cardiovascular mortality by 26% and overall hospitalization rates by 18%, with additional improvement in quality of life. In this population, CVR should be encouraged whenever possible.CVR is an adjunctive therapy that is also effective after acute coronary events and surgical or percutaneous revascularization. A systematic review and meta-analysis 154 Other similar studies reported reductions in cardiac or all-cause mortality on the order of 8\u201334% for each MET of improvement in cardiorespiratory fitness. 156The improvement in cardiorespiratory fitness is one of the factors responsible for reduction of all-cause mortality after CVR. In a cohort of 5,641 CVR patients in Canada, every 1 MET increase in cardiorespiratory capacity was found to decrease all-cause mortality by 25%. 2 peak, 27% improvement in quality of life, and 20% reduction in cardiac events, including fewer AMIs and fewer hospitalizations, were observed in patients who underwent CVR after angioplasty when compared to those who remained sedentary. 157In addition, CVR provides an add-on effect to reduce cardiovascular events after coronary angioplasty, as demonstrated in the ETICA trial (Exercise Training Intervention After Coronary Angioplasty). A 26% increase in VO 146CVR is indicated in all cases of CAD . It is Despite increasingly early interventional treatments and decreased length of hospital stay after acute coronary syndromes, it is not uncommon for patients to begin rehabilitation only after outpatient follow-up with their primary physician, which may mean 15 days or longer after the event. Early initiation of CVR is possible, and can have a direct, positive influence on adherence and on the degree of clinical benefit achieved after the acute event.163 others report neutral 139 or even positive effects 164 on this process. A systematic review and meta-analysis 159 carried out to answer this question found that the changes observed in ventricular function, ventricular diameter, and functional capacity were directly related to the timing of CVR initiation. The greatest benefits in ventricular remodeling and functional capacity were obtained when programs were started in the acute phase (6 hours to 7 days) after the event, declining when initiated 7\u201328 days after the event and even further after 29 days, at which time the positive effect on ventricular remodeling was progressively lost. It is important to note that there was no difference in events between the initial training phases and that the sample studied was primarily composed of young men, which highlights the need for further studies, especially in other populations (such as older adults and women). For every 1-week delay in initiation of CVR after an AMI, an additional 1 month of training may be necessary to achieve similar benefits in end-systolic volume and left ventricular ejection fraction (LVEF). 160One of the greatest concerns of early CVR refers to the effect of physical training on the ventricular remodeling process. While some authors report negative effects, 168 Therefore, political, social, and structural changes \u2013 as well as a shift in medical culture \u2013 are needed to change this scenario.Although widely endorsed by the medical literature for their beneficial effects and cost-effectiveness, CVR programs are only attended by a minority of eligible patients. Multiple barriers can explain this, such as a lack of programs, difficulty in accessing existing services, few referrals, and poor urban mobility; women, the elderly, and ethnic minorities are most affected. Both in patients with stable CAD and after a coronary event and/or revascularization, risk stratification for CVR is essential and should be based on a targeted clinical evaluation focused on detailed knowledge of the patient\u2019s CVD and treatment history, whether clinical or interventional. Presence of symptoms, ventricular function, functional capacity, arrhythmias, and the possibility of residual ischemia all aid in stratification and should queried during the initial assessment. Ideally, this evaluation should be carried out by a CVR team member (rehabilitation physician).The profile of a patient referred to CVR can vary widely, from individuals receiving elective treatment to patients with a complicated acute coronary syndrome and history of prolonged hospitalization. A broader assessment, including nutritional, psychological and musculoskeletal issues, should be part of the clinical history and examination, as these factors can directly impact the CVR process. In patients who have undergone percutaneous or surgical revascularization, examination of the arterial puncture site or surgical wound should always be performed. Any abnormalities or complex medical needs identified during the pre-exercise evaluation must be relayed to the CVR team members who will be involved in the patient\u2019s exercise sessions.During pre-exercise evaluation for CVR, the purpose of functional tests is to gain better insight into functional capacity, identification of residual ischemia and stress-induced arrhythmias. Myocardial ischemia on exertion is identified by detection of symptoms such as angina pectoris and/or by ECG changes. The ischemic threshold can be identified during TMET by the onset of these clinical and/or ECG changes and expressed as the load and/or HR at which ischemia initiate. This information is essential to guide the exercise prescription.When used for exercise prescription purposes, TMET should be performed on all of the patient\u2019s usual medications, especially those that may affect HR. This is important to reproduce the conditions that will be present during training sessions. If patients on beta blockers have their dose adjusted during rehabilitation, ideally a new TMET would be performed for adjustment of the exercise prescription. If this is impossible, a subjective (perceived exertion) test may assist in adjusting the prescription until a new test can be performed.In some cases, patients entering CVR may have clinical limitations precluding a maximal exercise test. In these patients, the initial exercise prescription can be guided by a submaximal test, and a maximal test performed once there has been sufficient clinical improvement and/or optimization of drug therapy. Considering the possibility of serious error due to marked individual variation in chronotropic response, formulas that consider age as a parameter to define peak HR should never be used. The error is even greater in patients who are on beta blockers.131 The target exercise intensity can also be determined by subjective assessment of breathing; moderate-intensity activity leaves the patient only slightly short of breath, but still able to speak complete sentences without interruption and on arbitrary limitation of HR during training, i.e., the use of a resting HR + 20 bpm for patients who have had an acute coronary syndrome or resting HR + 30 bpm for those who have undergone elective surgical or interventional revascularization. tion see .When a TMET is performed, the intensity of the prescribed exercise should lie between 40 and 80% of HR reserve (Karvonen formula: [Peak HR \u2013 resting HR] x percent intensity + resting HR). In such cases, the initial exercise prescription usually targets the lower limit of HR, progressing from there according to the patient\u2019s clinical course and improvement in functional capacity. Most patients will be prescribed an exercise intensity between 50 and 70% of their HR reserve. Those that are more functionally limited or have significant ventricular dysfunction may be prescribed at lower intensities (40\u201360%), while those who were previously active and still retain better functional capacity may be prescribed at a higher and wider range (50\u201380%). Percentages of peak HR can also be used, with moderate-intensity exercise corresponding to 70\u201385% of HR peak see .169 When there is an occurrence of an early plateau of oxygen pulse or, particularly, a drop during exertion, the exercise prescription should be limited to loads below that alterations. Thus, CPET is considered the gold-standard assessment method to support exercise prescription and should be used whenever it is available. 171 In patients who complete a CPET, the prescribed exercise intensity should lie between the ventilatory thresholds and increase progressively from there.CPET, by allowing analysis of the oxygen pulse response, provides increased sensitivity and specificity for the diagnosis of myocardial ischemia. Regarding the volume of exercise, at least 150 minutes/week are recommended, distributed across 3 to 5 sessions. Depending on tolerance, adaptations to training, and individual preferences, as well as consideration of clinical status, this volume may be increased to 300 minutes or more per week.For resistance training, the gold-standard method to determine the optimal intensity is 1RM testing. However, in practice, many rehabilitation programs do not perform this test due to time constraints or clinical limitations, such as in patients who have undergone CABG and may thus be limited not only by sternotomy but also by saphenectomy. In such cases, subjective perceived exertion is a practical and useful alternative.173In patients who have undergone sternotomy, upper body exercise should be limited to low intensity and performed with restricted loads for 5 to 8 weeks. Exercises involving the full range of motion of the arms may be allowed after this period if there is no sternal instability, although recent and ongoing studies are evaluating the safety of earlier exercise after CABG. Patients should always be instructed on how to correctly perform movement and breathing, avoiding the Valsalva maneuver. The interval between series can range between 45 s and 1 min, depending on the load applied and the patient\u2019s tolerance.6.3.4.1. Refractory Angina175 Such patients are generally not referred to CVR programs due to fear of adverse events during physical training, although rehabilitation has already been considered a feasible and safe possibility for these patients. 175Refractory angina is defined as disabling angina of over 3 months\u2019 duration, despite optimized clinical treatment, with documentation of myocardial ischemia in a patient who is not considered eligible for percutaneous and/or surgical coronary intervention. 178 A single controlled study has evaluated CVR in patients with refractory angina. The study randomized 42 subjects to a CVR exercise program or usual care for 8 weeks. Patients in the CVR group were prescribed training to a target HR between 60 and 75% of HR reserve (for those with preserved ventricular function) or between 40 and 60% of HR reserve (when LVEF was <40%). Patients in the rehabilitation group increased their total distance on the shuttle walk test by 50 m, with no change in severity or frequency of angina. There were no adverse events in either group. 161The objective of therapeutic interventions in this setting is to improve quality of life and facilitate performance of the activities of daily living. 162 will evaluate the safety and efficacy of a 12-week supervised exercise program in patients with refractory angina, carried out in a hospital environment with continuous ECG monitoring. Exercise prescription is individualized, on the basis of CPET findings and the ischemia and/or angina threshold. To date, 42 patients have been included, and no exercise-emergent cardiovascular events or hospitalizations related have been documented. Serum levels of high-sensitivity troponin T, a known predictor of worse prognosis, 179 did not change in 32 patients who completed a 40-minute acute aerobic exercise session (at the ischemia threshold) at the time of study enrollment ( unpublished data ).An ongoing Brazilian randomized trial 180In patients with refractory angina and a low ischemic threshold, administration of rapid-acting nitrates before the start of each physical training session can help prolong the duration of training and even allow exercise at higher intensities. 6.3.4.2. Exercise Training with Myocardial Ischemia182 because changes in contractility and perfusion defects precede clinical and electrocardiographic ischemic changes. 184Traditionally, there is a recommendation that physical exercises in patients with CAD be performed below the clinical and electrocardiographic ischemic threshold; however, this can be difficult to control. Previous studies have shown that physical exercise, even when prescribed according to literature recommendations, can trigger scintigraphic perfusion defects which are not demonstrable on ECG and do not trigger angina, 185 Other authors demonstrated in a small series of patients with CAD that, after 6 weeks of training, repetitive ischemic stimuli did not result in significant damage, myocardial dysfunction, or arrhythmias. 187The functional significance of ischemic defects visible only on myocardial perfusion imaging is still unclear, but some studies of training above the ischemic threshold have been carried out. In one study of a single 20-minute training session conducted above the ischemic threshold, no evidence of acute myocardial damage was identified. 188 Additionally, there is evidence of the superiority of combined aerobic and resistance training as opposed to aerobic training alone in patients with CAD. 189Therefore, there is evidence to suggest that interval training, a modality that has proven to be safe and effective in improving physical fitness, endothelial function, and left ventricular function with results superior to those obtained with moderate-intensity continuous training (MICT), may be feasible in patients with stable CAD. 6.3.4.3. Drug Adjustments in Response to Physical ExercisePatients with stable CAD usually rely on medications for symptom relief, reduction of ischemia, improvement of endothelial function, stabilization of atherosclerotic plaque, control of risk factors, and maintenance of adequate hemodynamics. For example, high SBP and/or HR levels (increased double product) lead to clinical deterioration. On the other hand, systolic hypotension and bradycardia produce reduced cardiac output, which can cause abnormalities due to a drop in coronary flow.In CVR programs, particular attention should be paid to improving the angina threshold before training begins, which allows greater tolerance to the progression of exercise intensity and, thus, facilitates achievement of the desired beneficial effects. Therefore, the optimization of drug therapy is essential for a safe and effective CVR.190 These adaptations can reduce the need for antianginal and antihypertensive agents. It is the role of the rehabilitation physician to discuss adjustments of drug therapy with the patient\u2019s primary physician as necessary.Patients engaging in RCV can present a number of physiological adaptations to exercise, including modulation of the autonomic nervous system and reduced HR at baseline and on exertion. Together, these adaptations improve endothelial function and BP reduction, reduce afterload, and improve the diastolic function of the heart. 191 impairment of vasoreactivity, peripheral endothelial dysfunction, 192 and chronic inflammation. 193 In this context, physical exercise has been established as a safe therapeutic strategy that mitigates the effects of progressive physical deconditioning due to the natural course of the disease. 194Chronic HF is a complex, multisystem syndrome that features dyspnea and progressive exercise intolerance are its core symptoms. Despite recent advances in drug therapy, which have reduced once very high morbidity and mortality rates, symptoms tend to persist, compromising patient quality of life. There is consistent evidence that reduced levels of physical activity in HF trigger a vicious circle that contributes to increasing symptoms and exercise intolerance, secondary to a reduction in functional capacity, producing negative psychological effects, 197 A single large, multicenter randomized trial, HF-ACTION, 198 revealed a modest but nonsignificant reduction in primary outcomes , as well as major benefits in quality of life and a reduction in the rate of HF hospitalization. As a weakness of the study, poor adherence to exercise probably impaired the effectiveness of the intervention, a hypothesis that was confirmed later in another study, which demonstrated that adherence is a determining factor for obtaining medium-term benefits. 199Small randomized studies, systematic reviews, and meta-analyzes have consistently demonstrated that regular physical training is safe, increases exercise tolerance, improves quality of life, and reduces hospitalizations in HF. 2 on physical training in patients with HF, which analyzed 33 randomized studies including 4,740 patients (with a predominance of reduced ejection fraction), there was a trend toward reductions in all-cause mortality in the physical exercise group at 1 year of follow-up. Compared to controls, the physical training group had a lower rate of HF hospitalization and improved quality of life. Regarding benefits in women with HF, the available studies suggest that a positive impact equivalent to that seen in men. 200In a systematic review 201 Nevertheless, additional studies are needed to confirm these initial results before a stronger recommendation can be issued.For patients with advanced symptoms (NYHA class IV), data are still insufficient to indicate specific exercise programs. A single Brazilian randomized trial tested a daily exercise program on a cycle ergometer combined with noninvasive ventilation. The study evaluated patients hospitalized for acute decompensated HF, and found functional benefits and reduced length of stay. 2 peak (measured by CPET), 203 quality of life, 204 and diastolic function (as assessed by echocardiography). 205In HF with preserved LVEF, there is recent evidence from small randomized studies and a systematic review showing benefits in VO 207In light of this evidence, exercise-based CVR is recommended in HF regardlPhysical exercise alone should not be prescribed for patients with clinically unstable HF, with a clinical picture suggestive of acute myocarditis, or in the presence of acute systemic infection (Class IIIC).Internationally, CVR programs are implemented in a wide range of formats, using different exercise modalities alone or in combination. The exercises can include aerobic training (moderate- and/or high-intensity), localized resistance training, and respiratory muscle training .208 Functional tests should be performed while the patient is on his or her prescribed medications, to mimic the conditions of actual training.Before starting the training program, it is essential that the patient be clinically stable and on optimized drug therapy; ideally, a functional assessment should be performed, preferably with CPET or a TMET. If the aforementioned functional tests are unavailable, the 6-minute walk test can serve as a parameter for monitoring functional gains. The recommended aerobic training can be MICT, which corresponds to the HR zone delimited by the ventilatory thresholds of CPET, or, in the case of a TMET, to the zone located between 60 and 80% of peak HR or 50 and 70% of reserve HR. Patients with more severe disease and greater functional limitations may start at the lower end of the prescription. Intensity can progress up to the upper limit as training advances.Recently, the use of high-intensity aerobic exercises performed in an interval mode \u2013 known as high-intensity interval training, or HIIT \u2013 has become popular. In this modality, more intense periods of exercise alternate with periods of passive or active recovery, which allows a greater total duration of high-intensity exercise and, consequently, can increase stimuli for central and peripheral physiological adaptations.209 demonstrated that HIIT was superior to MICT in promoting improvement in functional capacity and in different cardiovascular parameters. Subsequently, other clinical trials were carried out and meta-analyzed. The superiority of HIIT over MICT in terms of effects on functional capacity was confirmed in a meta-analysis. 210 The largest multicenter study published to date, Smartex-HF, 211 compared MICT versus HIIT. The authors found similar benefit, with no superiority of one modality over the other in any respect. Therefore, the choice of protocol will depend on team experience, clinical conditions, physical capacity, and patient preferences.In HF patients with reduced LVEF, Wisl\u00f8ff et al. 212 One consists of 4 min of high-intensity exercise (90 to 95% of maximum HR) alternating with 3 min of low-intensity exercise (70% of maximum HR). 209 Protocols with much shorter durations of high-intensity load (30 or 90s) have already been described, and the tolerance to different HIIT protocols may vary according to patient preference and physical capacity. 213 Therefore, the use of this modality will depend on the patient\u2019s clinical picture and choices, as well as on the experience and preferences of the CVR team.In addition, HIIT protocols can vary widely; several have been described. 214 These exercised can be prescribed as percentages of maximum voluntary contraction or according to subjective perceived exertion. The recommended loads and repetitions may vary according to the patient\u2019s functional limitations and must be individualized, progressing as rehabilitation itself progresses.The addition of localized muscle resistance exercises to aerobic training has been suggested as a means of obtaining additional benefit. 215 In a meta-analysis by Smart et al. 216 which evaluated 11 studies including 287 participants with HF, 148 of whom underwent inspiratory muscle training (IMT) compared with 139 sedentary controls, significant gains in VO 2 peak, distance walked in the 6-minute test, quality of life, PImax, and VE/VCO 2 slope were observed. Thus, IMT provided gains in cardiorespiratory fitness and quality of life of a similar magnitude to those obtained with conventional training, and should be considered a valid alternative for severely deconditioned and debilitated HF patients, perhaps as a bridge to conventional physical exercise.The addition of breathing exercises has been recommended for patients with respiratory muscle weakness. 217Given the variety of benefits observed, it is essential that patients with HF perform physical exercises regularly. Ideally, this should be done in the context of a CVR program, with an individualized prescription combining moderate- and/or high-intensity aerobic training, localized muscle resistance exercises, and respiratory muscle training, depending on the patient\u2019s clinical condition and functional limitations, and according to patient preferences and staff experience. In addition, there are valid alternatives even for very debilitated and severely deconditioned patients. Heart transplantation (HTx) is the treatment of choice for patients with refractory HF, whose symptoms remain severe despite use of the entire pharmacotherapeutic arsenal and surgical procedures as indicated.218In recent years, there have been significant advances in HTx, with the emergence of new surgical techniques and the development of more efficient immunosuppressants. In Brazil, there has been substantial growth in the number of procedures, which had been stagnant since 2015, with a rate of 1.7 transplants per million population (pmp). In 2019, the rate grew 17.6%, reaching 2 transplants pmp, very close to the target set for the year (2.1 pmp). In 2018, 357 procedures were performed, and by March 2019, 104 hearts had been transplanted in Brazil. 220 Recipients are able to return to work and lead normal lives with minimal or no symptoms. 221 The survival rate is estimated at 90% at 1 year and around 70% at 5 years. 222HTx aims to improve quality of life, as well as survival, in this population. 2 peak is still reduced when compared to that of healthy, age-matched individuals. 224 Among other factors, this can be explained by: 1) in the immediate post-transplant period, the allograft is devoid of sympathetic and parasympathetic innervation (autonomic denervation), causing an increase in resting HR, attenuating its natural elevation in response to exercise, and impairing recovery after exertion 225 ; 2 ) patients often exhibit skeletal muscle dysfunction (sometimes to the point of cachexia), in which immunosuppressive therapy and pre-transplant HF play prominent roles 226 ; and 3) impairment of vascular and diastolic function. 227 During the acute phase of exercise, the increase in cardiac output of HTx recipients depends fundamentally on the Frank\u2013Starling mechanism, i.e., on increase in venous return, inotropy, chronotropy, and reduction in afterload. 229 In addition, there is an increase in the concentrations of circulating catecholamines, 227 which decrease slowly after the end of exercise, explaining the slow recovery of HR in these patients. 230Although HTx significantly improves patients\u2019 functional capacity, VO 231 and these patients may develop HTN, diabetes mellitus, and CAD. 232 Conversely, physical exercise is known to be an excellent therapy for management of these chronic diseases 233 and is effective in optimizing autonomic control. 234Immunosuppression may predispose HTx recipients to a higher risk of other complications, 2 peak and improvements in hemodynamic control, muscle strength, and bone mineral density, 236 thus improving prognosis. 19 Although there are countless possibilities for training prescription, the recommended method remains aerobic exercise, which can be performed continuously or, in specific cases, at intervals and at different intensities, 170 combined with resistance training whenever possible. 6Physical training after HTx contributes to an increase in VO 237 the investigators found that, 46 months after HTx, patients who underwent aerobic training had a functional capacity and chronotropic function similar to those of healthy individuals. Previous studies had already demonstrated the safety of physical training in this population. 240In a pioneering study by Richard et al., 234 An average increase in VO 2 peak of 2.5 ml.kg - 1 .min - 1 was observed in those who received training versus those allocated to usual care. Rosenbaum et al. 241 assessed the relationship between early participation in a CVR program after HTx and found that the number of sessions performed in the first 90 days was directly associated with better 10-year survival.A Cochrane meta-analysis of nine randomized clinical trials, including 284 patients, compared the effect of physical training to usual care in the post-HTx setting. 242 described significant improvements in VO 2 peak of HTx recipients, with an average increase of 3.1 ml.kg - 1 .min - 1 after 12 weeks of combined training (resistance and aerobic). Kobashigawa et al. 243 studied 27 patients after HTx who received a combination of aerobic, resistance, and flexibility training for 6 months versus a control group. The duration and intensity of the aerobic exercise sessions had a goal of at least 30 minutes of continuous, moderate exercise on a cycle ergometer. The intervention group showed an average increase of 4.4 ml - 1 .kg - 1 .min in VO 2 peak, versus 1.9 ml.kg - 1 .min - 1 in the control group. These data provide valuable information on the importance of both types of training for this population.Haykowsky et al. 244 found a greater effect of HIIT compared to MICT on VO 2 peak, with an additional gain of 2.3 ml.kg - 1 .min - 1 and superior improvement in quality of life. One meta-analysis 233 included three randomized controlled trials that compared HIIT (intense blocks: 80 to 100% of VO 2 peak or 85 to 95% of peak HR) to usual care. Post-HTx patients randomized to HIIT showed an increase in VO 2 peak of 4.45 ml.kg - 1 .min - 1 after the intervention period, which ranged from 8 to 12 weeks of three to five weekly sessions.Regarding high-intensity training in patients after HTx, the number of studies is still small, but the results obtained have been encouraging. In a crossover study, Dall et al. 224 evaluated the effects of a HIIT program compared to a control group in 43 HTx recipients. The authors evaluated the progression of allograft vasculopathy, assessed by intravascular ultrasound, and found less progression of atheromatous plaque in the HIIT group. However, additional studies are still needed to elucidate these benefits. 245Nytr\u00f8en et al. 235 were the first to study the effect of resistance training on glucocorticoid-induced myopathy in HTx recipients. One group received training and was compared with a control group. After 6 months, although both groups had increased muscle strength in the quadriceps and lumbar extensors, the increase was up to six times greater in the training group.Some well-known common adverse effects of the use of glucocorticoids after HTx are muscle atrophy and weakness. In 1998, Braith et al. 236Resistance training also appears to have a major therapeutic effect on bone metabolism. After HTx, patients commonly experience significant bone loss at femur head and mineral total bone loss. In one study, patients were enrolled for resistance training 2 months after HTx, and training was shown to be able to restore bone mineral density to pre-transplant levels. 246 The physical therapist and/or physical educator will prescribe, administer, supervise, and guide exercise, following the safety limits recommended by the physician on the basis of the pre-exercise evaluation. 247HTx recipients must undergo a thorough history, physical examination, 12-lead resting ECG, color Doppler echocardiogram, and other tests at the discretion of the CVR team. Ideally, a functional stress test should be performed, preferably CPET, which is the gold-standard method for assessing functional capacity in this patient population. The stress test must be performed by a trained physician; it evaluate the cardiopulmonary and metabolic responses to increasing exercise and yields several variables that have an impact on the clinical examination and the exercise prescription. 170 When even this is unavailable or otherwise impossible, the 6-minute walk test can assist in clinical assessment, in addition to providing a parameter for comparing functional capacity during training. 249The impossibility of performing CPET should not be considered an impediment to exercises; if CPET is not available, a TMET is suggested. 170 In patients who have undergone HTx, most studies evaluated the effect of MICT.Aerobic exercise is most recommended, with supplemental resistance training, starting on the 6th week after HTx. Different training methodologies have been studied in isolation and have proven effective in promoting cardiovascular health in individuals undergoing CVR. 250 In this sense, programs that include interval training (even HIIT) have demonstrated good outcomes. 233 However, an optimized and safe exercise prescription requires proper individualization of each component of the training session. 170Depending on the patient\u2019s clinical condition, the intensity of aerobic exercise may be gradually increased from moderate to high over the course of training, in order to optimize adaptation and obtain greater benefit, as exercise intensity is directly associated with the magnitude of cardiovascular adaptations. 170 However, as recent HTx recipients will exhibit a compromised chronotropic response, 251 prescriptions based on percent of peak HR or threshold HR will not be useful during the first training sessions, although they may be used once there has been improvement in autonomic response. 224 Continuous assessment of the HR response to exercise and during recovery is thus extremely important. When CPET is available, the prescription of aerobic exercise can be based on the ventilatory thresholds or on established percentages of VO 2 peak. Another simple and feasible strategy is assessment of subjective perceived exertion using the Borg scale. 252 The multidisciplinary team must be firmly committed to educating the patient regarding the various levels of perceived exertion and the symptoms of which they should be aware. 6The determination of target training zones is advised as a means of optimizing the exercise prescription. 253 This test has been validated in active older adults and proved to be reasonably reliable in providing information about lower limb strength, and is now widely used in rehabilitation centers and scientific research on a wide range of clinical conditions. 256In addition to the evaluation and prescription of aerobic exercises, resistance exercises are essential. The methods traditionally used for pre-exercise assessment and exercise prescription are 1RM load tests. However, the use of these protocols after HTx \u2013 especially after a recent procedure \u2013 may be inappropriate, and clinical investigations on the safety of these tests in this specific patient population are still lacking. An alternative evaluation method is the 30-second sit-to-stand test. Resistance exercises may also be prescribed subjectively, on the basis of perceived exertion alone. The variable repetition method may be used, whereby the aim of the user is not a set number but a range of repetitions . If the patient is unable to perform at the lower end of the range, the applied load is too high; if the patient can execute the maximum number of repetitions with ease, the load is too light. Thus, the load can be adjusted so that training takes place within the proposed range of repetitions.258 This clearly demonstrates the importance of patient supervision throughout training, should any intercurrent event arise that warrants discontinuation of the exercise session. In view of the foregoing, some authors suggest that patients should not perform physical exercise when receiving pulse steroid therapy and on the days of myocardial biopsies. 170During training, particular attention should be paid to any complications or intercurrent events, such as infections related to the transplant procedure. A. survey found that 36% of HTx recipients are hospitalized within the first year after transplantation, and 61% at 4 years. 1 and are recommended as an alternative to traditional CVR in low-risk patients. 71Previous studies have shown that HBCR programs are safe and effective, 259 conducted a prospective, randomized study to evaluate the effect of a home exercise program for 2 months in 37 patients after HTx. The control group maintained their usual lifestyle throughout the study period. Individuals in the intervention group performed an exercise program at least three times a week which included a 5-min warm-up, upper limb and lower limb strength training, 15 to 20 min of aerobic exercise at an intensity of 60 to 70% of VO 2 peak, and a 5-min cooldown period. To ensure proper exercise performance at home, an initial period of direct supervision was enforced. At the end of the 2-month period, patients had improved muscle strength and endurance, fatigue index, and quality of life . CPET revealed an increase in workload, but with no change in VO 2 peak, probably due to the short follow-up period or the less-intense methodology used to guide the training prescription.Wu et al. 260 with an equivalent aerobic training protocol but a longer duration (five times a week for 6 months), documented improvements in VO 2 peak, workload, and BP in individuals after HTx. In addition, there were signs of cardiac sympathetic reinnervation and restoration of arterial sensitivity to autonomic modulation, with no changes in the control group.Another study, 2 peak. 261Even beyond 5 years after HTx, HBCR can still improve functional capacity, as demonstrated by a study in which 21 patients were instructed to follow a home-based physical training program consisting of cycle ergometer exercises for 1 year. Nine patients served as controls. To ensure adequate control, patients received a smart card programmed for a 6-min warm-up and 20 min at a constant workload, with load adjustment according to the exercise prescription and HR monitoring. At the end of 12 months, there was a modest improvement in VO 262 compared the effects of home-based and hospital-based exercise programs on exercise capacity and chronotropic variables in 28 patients after HTx and observed significant improvements in VO 2 peak and HR reserve only in the traditional CVR group. However, new studies, with the inclusion of a larger number of patients, are necessary to better elucidate this superiority of the hospital-based program observed in this study.Karapolat et al. Based on the evidence reviewed above, physical training has an unequivocal beneficial effect after HTx, is safe and feasible, and can be performed in the hospital or home environment . HoweveCVR should be started 6 to 8 weeks after the HTx, with referral at hospital discharge. In selected cases and after careful evaluation by the CVR team, rehabilitation may begin earlier. As in any post-sternotomy situation, special care must be taken not to prescribe exercises that might overload the chest muscles and lead to sternal traction, especially in the first 90 days after transplant.The ideal prescription will include exercises that promote different components of physical fitness, always maintaining an emphasis on specific recommendations for each condition. After HTx, just as in other indications for CVR, aerobic exercise should be the main component of training sessions, supplemented by resistance and flexibility training within an individualized, periodically reassessed program. Sessions should always start with a warm-up period and end with a controlled cooldown period. This strategy aims not only to warm up the skeletal muscles but also to provide an adequate time for adjustments of HR and BP to exercise, as the exertion response in these patients is affected by denervation of the heart, especially in the early stages of the training program after transplant.Aerobic exercise may consist of walking or cycling, whether indoors (using treadmills and/or cycle ergometers) or outdoors. A weekly frequency of three to five sessions, each lasting 20 to 40 minutes, is recommended. The frequency and duration of these sessions will be adjusted according to the patient\u2019s preexisting condition and should progress over time with training. Control of exercise intensity is essential; given the larger evidence base, MICT (between the first and second ventilatory thresholds) is recommended, with a perceived exertion no greater than 11 to 13 on the modified Borg Scale. In selected cases, interval training can be adopted to add variety and, potentially, to enhance functional gain.Resistance training is essential, especially in the early phase after transplantation. Many HTx recipients had longstanding HF, endured prolonged hospitalizations, and have been exposed to massive surgical stress. In this regard, a resistance exercise program can be particularly useful. At the start of training, activities performed against body weight alone are considered sufficient for these patients. Over time, elastic bands, dumbbells, ankle weights, and weight machines can be added to the training program. Greater than usual care is needed during upper body exercises, considering that corticosteroid therapy may make for slower healing of the thoracotomy scar.265Further information and examples of training protocols for these patients are available elsewhere. This section will address hypertrophic cardiomyopathy (HCM), myocarditis, and other cardiomyopathies. The indications for CVR in this setting are listed in 271 It is the most common hereditary heart disease in the general population, caused by a range of mutations in genes which encode the cardiac sarcomere proteins. 268 HCM has a characteristically heterogeneous clinical expression, with unique pathophysiological changes and a variable natural history. Up to 10% of cases are caused by other genetic disorders, including hereditary metabolic and neuromuscular disorders, chromosomal abnormalities, and genetic syndromes. 272 Some patients have other disorders that can mimic HCM, such as amyloidosis. 273Hypertrophic cardiomyopathy (HCM) is a disease characterized by left ventricular hypertrophy, usually without dilatation of the ventricular chambers, in the absence of another cardiac or systemic disease capable of explaining the magnitude of hypertrophy observed. 268 However, this estimate appears to differ in clinical practice, which allows us to infer that a portion of the affected individuals are asymptomatic. Various patterns of asymmetric hypertrophy of the left ventricle are commonly seen in HCM, and there may be different phenotypes in first-degree relatives. Typically, one or more regions of the left ventricle exhibit increased wall thickness when compared to others; transitions and variations in thickness may occur in adjacent or noncontiguous areas. However, although asymmetric septal hypertrophy is the most common finding, there is no \u201cclassic\u201d HCM pattern, and virtually all possible patterns of left ventricular hypertrophy can occur. Hypertrophy may even be absent in genetically affected individuals, in what is known as a negative phenotype.The population-wide prevalence is estimated at around 0.2% or 1 in 500. 274 Notably, only a small subgroup of patients with HCM experience significant complications and premature death; these complications can occur due to obstruction of the left ventricular outflow tract, HF with diastolic and/or systolic dysfunction and sudden cardiac death (SCD), or cardiac arrhythmias . 275 In HCM, SCD can occur at any age, although it is most common in adolescents and young adults; therefore, identification of individuals at the highest risk is an essential component of the pre-exercise evaluation, especially in patients who may want to engage in competitive sports. 276Several multicenter retrospective and observational cohort studies, conducted in different populations, have elucidated the natural history and clinical course of HCM. Recent studies have reported an annual mortality of around 1%, much lower than in older surveys. 279 Therefore, encouraging a healthy lifestyle for HCM patients is essential to reducing the overall risk of morbidity.In many cases, SCD can be the first manifestation of the disease; indeed, it occurs most commonly in those without warning symptoms and who had not been diagnosed prior to the event. Nevertheless, most patients with HCM have a normal or near-normal life expectancy, with mortality usually attributable to other causes, some even of non-cardiovascular etiology. 6.6.1.1. Therapeutic Benefits of Physical Exercise19 In patients with obstructive and minimally symptomatic HCM, an association of mortality with aerobic fitness has also been observed. 281 Patients with a VO 2 peak below 18 ml.kg - 1 .min - 1 on CPET had higher mortality and were more symptomatic compared to those who achieved values equal to or greater than this threshold. A VO 2 peak below 60% of predicted was associated with worse 4-year survival (as low as 60%). 280In the general population, cardiorespiratory fitness is a determinant of the risk of cardiovascular and all-cause mortality. 271 Evidence does suggest that high-intensity physical training could accelerate these changes, but this is still a controversial topic. However, it is well established that the increase in myocardial fibrosis is associated with a lower VO 2 peak in this population. 282Myocardial fibrosis and myofibrillar derangement may underlie the increased risk of SCD in HCM, as these structural changes act as a substrate for fatal arrhythmias. 281 When there is a reduction in VO 2 peak, physical exercise can help increase functional capacity.Therefore, assessing aerobic fitness \u2013 preferably through CPET \u2013 is essential in patients with HCM. 2 peak after 16 weeks of a moderate-training intervention (+1.35 ml.kg - 1 .min - 1 or < 0.5 MET). 266 Another prospective, non-randomized study included 20 patients with HCM and found a significant increase in treadmill test duration, as well as in estimated functional capacity (+2.5 MET). 267 In this study, patients completed a CVR program which consisted of 60-minute sessions of moderate to vigorous exercise, performed on a treadmill or cycle ergometer, twice a week. The intensity of exercise progressed from 50 to 85% of HR reserve, which resulted in a gradual increase in conditioning and may have minimized the risk of adverse events, such as exercise-induced arrhythmias. Serious adverse events, such as death, aborted SCD, implantable cardioverter-defibrillator (ICD) activation, or sustained ventricular tachycardia, did not occur in any of these studies. 267To date, only one randomized controlled trial has examined the effect of physical training on patients with HCM (RESET-HCM). This study, which included 136 patients, demonstrated an increase in VO 6.6.1.2. When Is Physical Exercise Indicated?281The intensity of exercise which patients with HCM can be cleared to do still represents a major challenge. If, on the one hand, intense physical exercise can be harmful, with an increased risk of potentially fatal arrhythmias, on the other hand, excessive restrictions on physical activity lead to deconditioning and can have negative effects on health and quality of life; they may even increase cardiovascular risk, given the well-established association between physical fitness and mortality. 283In its official position statement on management of HCM, the American Heart Association discourages patients with the disease from engaging in competitive sports of moderate to vigorous intensity see . This l284Exercise-triggered arrhythmias (in the short term) and adverse myocardial remodeling (in the long term) are the most fearsome side effects of exercise in HCM. The fear of SCD during sport extends to non-competitive athletic activities, although there is a clear lack of evidence about the safety of exercise in this patient profile. However, it should be emphasized that this risk of exercise is theoretical, and that recommendations to limit physical activity have been advocated with caution, based solely on the opinion of experts, and are not supported by more robust evidence. Thus, patients with HCM receive little guidance regarding the best dose or amount of physical activity to maintain general health and well-being; instead, greater focus is placed on restrictions on physical activities. As a result, more than 50% of patients with HCM do not achieve the minimum recommended physical activity target due to the belief that they are unable to exercise and/or that physical activity can worsen their disease.Therefore, a balanced approach seems to be most appropriate, and extremes should be avoided , as both could increase cardiovascular risk.New evidence suggests a positive effect of moderate physical exercise in selected patients with HCM, with individualized risk assessment and exercise prescription. It is noteworthy that the evidence suggests benefits of MICT, while other modalities need further studies.However, the presence of any of the following could be considered major contraindications to the practice of exercise: history of aborted SCD in the absence of an ICD; history of syncope on exertion; exercise-induced ventricular tachycardia; increased exercise pressure gradient (greater than 50 mmHg); and abnormal BP response to exertion.6.6.1.3. Pre-Exercise EvaluationClearance to begin exercising must be based on the pre-exercise evaluation, including a thorough history, physical examination, and 12-lead ECG.285 Electrocardiographic changes may precede structurally detectable disease for some years, which makes ECG extremely important in this scenario. 269 Only a minority of patients with HCM present with a normal ECG \u2013 usually those without any other phenotypic manifestations (positive genotype/negative phenotype).A large proportion of individuals with HCM are asymptomatic or oligosymptomatic; clinical suspicion is raised only by changes on resting ECG, which is abnormal in up to 95% of patients with the disease. 286Echocardiography remains the most widely used modality for diagnosis of HCM. Magnetic resonance imaging (MRI) is usually reserved for cases in which echocardiography is in conclusive, or to assess more localized hypertrophy . In young athletes, distinguishing physiological hypertrophy (\u201cathlete\u2019s heart\u201d) from the pathological hypertrophy of HCM is a challenge. This is because, most athletes with HCM exhibit an asymmetric pattern of left ventricular hypertrophy, as do sedentary individuals with the condition. In contrast, those with physiological left ventricular hypertrophy show a more homogeneous, symmetrical distribution of wall thickness, with only minor differences between contiguous segments and a symmetrical pattern of left ventricular hypertrophy. 287Exercise testing is always recommended in these patients prior to the start of CVR, whether to assess functional capacity or to detect abnormal BP responses and signs of increased dynamic obstruction of the outflow tract with exertion. For better detection of outlet tract obstruction during exercise, a combination of imaging (echocardiography) with stress testing is the gold standard and should be encouraged whenever possible. Patients with no obstruction at rest can present significant gradients on exertion, and thus be reclassified in relation to prognosis. 2 peak, a parameter with documented prognostic value. 281 In addition, determining ventilatory thresholds contributes to a more individualized exercise prescription.When available, CPET is advised instead of TMET, as it allows direct measurement of VO 6.6.1.4. Special Considerations for the Prescription and Follow-Up of Physical Exercise ProgramsSome particularities of exercise in patients with HCM should be noted:So-called \u201cexplosive\u201d activities , with the potential for rapid acceleration and deceleration, should be avoided.Activities with steady, constant energy consumption are preferred.Exercise in adverse environmental conditions, including extreme heat or cold, should be avoided, as there is an increased risk of exacerbating exercise-induced physiological changes.Training programs that aim competitivity, or achievement of higher levels of fitness and excellence, should be avoided, as they usually motivate patients to strive beyond safe limits.Intense static (isometric) exercises, such as weight lifting, should be avoided, as there is an increased risk of left ventricular outflow tract obstruction due to the intense Valsalva maneuver involved.Resistance training with low loads and a greater number of repetitions is considered safe for patients with CVD, although there is no solid evidence for patients with HCM.Some notes on drug therapy are warranted. Beta blockers and calcium channel blockers may be indicated in the treatment of HCM. As these medications attenuate the HR response to exercise, patients may experience a very reduced chronotropic response to exertion, which can cause increased exercise intolerance, suggesting a need for dosage adjustment. Excess diuretic use can be harmful because it increases the gradient of the outflow tract. Therefore, these agents should be used with caution. Like diuretics, exercise-induced dehydration can raise the outflow tract gradient; therefore, adequate hydration during training is of paramount importance.The pathogenesis of myocarditis consists of three phases: acute myocardial injury, usually of viral etiology; host immune response; and recovery, or transition to fibrosis and dilated cardiomyopathy. Clinically, there is no clear distinction between these phases. The initial insult can cause acute myocardial damage, with impairment of contractility mediated by cytokines produced by the local inflammatory process. This acute inflammation may progress, in the late phase, to extensive fibrosis, which can cause ventricular dilatation and dysfunction.283Acute myocarditis should be suspected when the following criteria are present: A clinical syndrome of acute HF, angina-type chest pain, or myopericarditis of less than 3 months\u2019 durationUnexplained rise in serum troponinECG changes suggestive of myocardial ischemiaGlobal or regional wall motion abnormalities and/or pericardial effusion on echocardiographyCharacteristic changes in tissue signal on T2- or T1-weighted MRI, as well as late gadolinium enhancement.290The participation of myocarditis patients in CVR programs after resolution of the acute phase has been the subject of very little study. There is no published research on the safety and effectiveness of this intervention. However, reports of CVR in this patient population have demonstrated benefits in quality of life and physical fitness, especially when there is functional impairment, even after improvement of the acute condition and optimization of drug therapy. 283 After this evaluation, selected cases may initiate moderate CVR, aiming at the general benefits obtained by patients with HF.Before starting any exercise practice, patients with a history of myocarditis should undergo echocardiography, 24-hour Holter monitoring, and an exercise test no less than 3 to 6 months after the acute phase has resolved. 269In sports, it is reasonable for athletes to return to their normal training routine only if they achieve: return of systolic function to normal values; markers of myocardial necrosis and inflammation within normal range; and absence of clinically significant arrhythmias on both Holter monitoring and an exercise test. It is noteworthy that the clinical significance of persistent late gadolinium enhancement on MRI in post-myocarditis patients whose clinical symptoms have resolved remains unknown. Thus, it seems reasonable that those with small areas of enhancement and without significant arrhythmias on Holter monitoring and exercise testing can return to sports, provided that clinical monitoring is continued. In chronic cases, in which ventricular dysfunction persists throughout the follow-up period, the patient should follow the general recommendations for CVR as described for chronic HF see .6.6.3.1. Arrhythmogenic Right Ventricular Cardiomyopathy291Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited disease that is associated with SCD in young adults and athletes. Pathologically, myocytes are lost and replaced with fibroadipose tissue, especially in the myocardium of the right ventricle, although isolated left ventricular or biventricular involvement may also occur. 292 In individuals with positive genotypes, an increased risk of arrhythmias with exercise has also been confirmed. Ventricular tachyarrhythmias and SCD events in this condition usually occur during exertion, including sports and endurance exercise, with an increased risk of tachycardia, ventricular fibrillation, and HF. 293There is evidence, in an experimental animal model, that exercise increases penetrance and risk of arrhythmias in patients with traditional ARVC mutations. 270 The reduction in exercise intensity was associated with a substantial decrease in the risk of ventricular tachyarrhythmias or death, especially in patients without a detected desmosomal mutation and with an ICD for primary prevention. 294 Therefore, the scientific evidence suggests that participation in sports and intense exercise are associated with early onset of symptoms and an increased risk of ventricular arrhythmias and major events in patients with ARVC. Therefore, these patients must be disqualified from participation in sport. 276It has been shown that individuals with ARVC who are involved in competitive sports experience a higher incidence of ventricular tachyarrhythmias and SCD, in addition to earlier symptom onset, compared with those who participated only in light physical activity and those who were sedentary. 21Regarding participation in CVR programs, there is no scientific data to indicate or suggest any benefits of physical exercise for patients with ARVC. On the other hand, keeping them sedentary, which contributes to low physical fitness, may also be inappropriate, as there is a general association of low physical fitness with mortality. 270 Thus, it can be assumed that participation in a supervised CVR program, restricted to exercise of light to moderate intensity, could not be harmful. Depending on other individual clinical characteristics, such as the presence of cardiovascular risk factors, physical exercises could be prescribed to control these conditions.In a small observational study of patients with ARVC, there was no difference in mortality rate between inactive individuals and those who performed only recreational physical activities. Therefore, the inclusion of a patient with ARVC in CVR programs should only be carried out after a thorough pre-exercise evaluation and rigorous evaluation of the risk-benefit balance of physical exercises. Options should be discussed with the patient, exposing the absence of proven benefits versus the potential risks of physical inactivity and low physical fitness. It is then up to the patient to choose according to their own personal preferences.In the context of CVR, extrapolating findings from athletes, a restriction on higher training intensities is also suggested. Patients with ARVC could thus perform supervised physical exercises of light to moderate intensity.6.6.3.2. Noncompaction Cardiomyopathy296Noncompaction cardiomyopathy (NCM) is a heart disease that occurs due to embryonic interruption of myocardial compaction. It is characterized by segmental thickening of the left ventricular walls, consisting of two layers: a compacted epicardial one and an endocardial one with marked trabeculation and deep intratrabecular recesses, where spaces are filled by blood flow. 297 Clinically, it can be asymptomatic or present with symptoms of HF, ventricular and/or atrial arrhythmias, pre-excitation, thromboembolic events, or SCD. There are no universally accepted criteria for morphological diagnosis; however, a ratio between noncompacted/compacted myocardium greater than 2.1:1 at the end of systole on echocardiography or 2.3:1 at the end of systole on MRI has become the most widely accepted proposed criterion. 298Its incidence and prevalence are uncertain, ranging from 0.02 to 0.05% according to some echocardiographic records. 300 In recent studies, athletes have shown a high prevalence of increased ventricular trabeculation when compared to a control group (18.3 versus 7%). It is believed that the increase in ventricular trabeculation or the presence of isolated echocardiographic criteria for cardiomyopathy is probably of little significance, and may be part of the spectrum of athlete\u2019s heart. 301 Therefore, not all athletes with isolated ventricular compaction are diagnosed with NCM. Therefore, functional parameters (such as ejection fraction) must also be considered to guide management. 301It is not yet established how physical training can influence NCM, nor is the frequency of development of noncompaction morphology in the population known. To date, there is no evidence from studies of CVR or training in NCM. Therefore, patients with left ventricular dysfunction should follow the same exercise recommendations as those with chronic HF see .304Patients with valvular heart disease represent a very heterogeneous group with major variability in terms of age, etiology, affected valves, and severity of involvement, whether due to stenosis, regurgitation/insufficiency, or mixed lesions. However, most valvular heart diseases share a common feature in their clinical manifestations induced by exertion, which include chest pain, dyspnea, and/or functional limitations. The severity of these symptoms in patients with severe valvular heart disease can be used as one of several criteria to indicate surgical or percutaneous intervention. In addition, the identification of reduced aerobic fitness, as documented by CPET or TMET, is also a criteria used to define whether interventions are indicated. One major issue in the clinical follow-up of patients with valvular heart disease is the prolonged natural history of these conditions. The onset and progression of symptoms and functional limitations is often slow, which may lead patients to spontaneously reduce their engagement in physical activity due to symptoms on exertion. This sedentary lifestyle can contribute to further reductions in aerobic physical fitness and worsen symptoms.Thus, doubts may arise regarding the clinical management and need for interventions when the patient undergoes a CPET or TMET, namely: are limitations in physical fitness identified on exercise testing a result of progressive valvular heart disease, a sedentary lifestyle, or both? In this context, the regular practice of physical exercise and the consequent maintenance or even improvement of physical fitness are important to elucidate these questions in the follow-up of patients with valvular heart disease.305 However, increases in the functional capacity of individuals referred for CVR have been demonstrated consistently, 307 which justifies referral to exercise-programs (level of evidence C).Participation of these patients in CVR programs has been the subject of a single cost-effectiveness study. Rehabilitation in the setting of valvular heart disease can be subdivided into two phases: pre- and post-intervention .Patients with moderate to severe valvular heart disease in the pre-intervention phase are rarely enrolled in CVR programs. Training is carried out mainly in asymptomatic cases, in whom there is still no indication for valve repair or correction.310CVR can be useful to keep the patient physically active while waiting for future intervention; after all, a sedentary lifestyle can deteriorate functional capacity and, thus, increase the risk of postoperative complications, especially when the intervention is performed in older adults with multiple comorbidities and established frailty. In addition, monitoring during supervised CVR sessions can be useful to observe changes in symptoms and physical fitness, which can indicate progression of valvular heart disease and suggest the need for medical reevaluation.Post-intervention patients are more common in CVR programs, as structured and supervised exercise is a useful means of observing the hemodynamic behavior of a patient\u2019s new (or newly repaired) valve. Information on a patient\u2019s response to physical exercise can help their primary physician adjust drug therapy and/or review valve function. In addition, supervised exercise provides a greater measure of safety for the patient to return to his or her activities of daily living, leisure, and sports.310 The exchange of information between the patient\u2019s primary physician and the rehabilitation physician is the best strategy for defining the optimal timing of referral, and the pre-exercise evaluation has a fundamental role in consolidating this shared decision.Although there is no consensually defined time limit for referral to CVR in the setting of valvular heart disease, the earlier the patient starts exercise, the less function he or she will lose from inactivity. The pre-exercise evaluation should always consist of a thorough history, physical examination, and evaluation of laboratory tests and imaging. The clinical history must include: length of hospital stay; complications related to the procedure, such as pleural or pericardial effusion, mediastinitis, and infections; type and size of prosthetic valve; surgical technique; and whether CABG was performed concomitantly, in addition to other clinical information that may be relevant regarding other comorbidities.On physical examination, cardiac and pulmonary auscultation are particularly important. In addition, attention should be paid to the surgical scar, which should be examined for signs of inflammation and infection, sternal instability, and pain or discomfort on palpation. If concomitant revascularization was performed, the saphenectomy and/or radial artery donor site must be examined. If valve repair or replacement was performed percutaneously, the access site should be checked for signs of peripheral vascular complications.311 Laboratory evaluation of coagulation is relevant in patients who received a mechanical valve and were started on anticoagulants. Achieving the correct level of anticoagulation is important in preventing complications.It is important that the clinician look for signs of anemia on physical examination and laboratory tests, because this is a common complication and can have a negative impact on functional capacity. 312A resting ECG should be obtained to check for any arrhythmias and disturbances in rhythm or conduction. The most commonly used imaging modality in the evaluation of valvular heart disease is Doppler echocardiography, which allows assessment of ventricular function and cavity dimensions, measurement of transvalvular pressure gradients, estimation of pulmonary artery systolic pressure, and measurement of blood flow, which provides a good overview of valve function and cardiac function at rest. Echocardiography should always be performed before the start of a CVR program, to assess the risk of exercise-related complications. 316 These tests, especially CPET, provide extremely useful information regarding aerobic fitness and the hemodynamic repercussions of valvular heart disease, which may be underestimated by assessments performed at rest. In addition, treadmill tests identify parameters that are used to guide exercise prescription and restrictions. When TMET and CPET are unavailable, the use of functional tests, such as the 6-minute walk test and the step test, should be considered. 320It is important to evaluate functional capacity by CPET or TMET. 321It is important to emphasize that CPET and TMET pose greater risk in patients with stenotic lesions; therefore, they should only be carried out by experienced physicians and in a safe setting with the necessary infrastructure to respond in case of emergency. 323Functional tests are indicated not only in pre-exercise evaluation, but also to elucidate any doubts regarding the symptoms of patients in the pre-intervention phase of valvular heart disease. The combination of functional tests with echocardiography helps assess the response of the transvalvular pressure gradient and pulmonary artery systolic pressure to exertion, especially when there is a discrepancy between echocardiogram findings at rest and clinical signs and symptoms. 324 Due to their high surgical risk, such patients are now considered candidates for percutaneous repair or replacement of the aortic 325 and mitral valves. 326 In this scenario, CVR can be considered before the intervention, with the aim of decreasing complication rates, length of hospital stay, and mortality and morbidity associated with the frailty syndrome. 327 After the intervention, CVR then provides an opportunity for monitoring and optimization of the outcomes of the procedure in all its aspects. 331Another relevant issue is the evaluation of elderly patients, who are frequently affected by valvular heart disease and have a high prevalence of risk factors and comorbidities. 332The use of frailty syndrome assessment instruments is still a controversial subject in the literature; there is no consensus regarding the best protocol to assess CVR outcomes. The assessment should include objective tests and instruments to address risk in several domains: mobility, muscle mass and strength, independence in activities of daily living, cognitive function, nutrition, anxiety, and depression. 334 Scientific evidence is scarce as to the impact of regular exercise on the progression of valvular heart disease and its complications; therefore, recommendations are based on expert opinion alone (level of evidence C).This section will only address guidelines and recommendations for exercise in patients with moderate or severe valvular heart disease, as there are no restrictions to exercise in patients with mild involvement. Participation in competitive sports should follow the recommendations of the specific literature on the subject. Acutely, exercise causes an increase in adrenergic tone and in the hemodynamic load imposed on the cardiovascular system, which raises concerns regarding the potential for deleterious cardiovascular effects in patients with valvular heart disease, including progression of aortic disease, functional deterioration, pulmonary hypertension, cardiac remodeling, myocardial ischemia, and arrhythmias.Patients with valvular heart disease who will start a CVR program must undergo a stress test to guide exercise prescription. For symptomatic patients in whom surgical correction is not indicated or who do not have the characteristics described in In patients who have undergone surgical correction of valvular heart disease, the exercise prescription will depend on the underlying disease, the outcome of the procedure, the presence of residual lesions, ventricular function, and the response to the exercise test (TMET or CPET). Therefore, each case must be assessed individually, and the limits of exercise prescription defined by the pre-exercise assessment and the results of physical examination and any other tests performed.This section describes particulars involving implantable devices: artificial pacemakers and implantable cardioverter-defibrillators (ICD). Artificial pacemakers are indicated in the management of electrical abnormalities, which may be isolated \u2013 sick sinus syndrome, advanced atrioventricular (AV) block \u2013 or associated with structural heart diseases. ICDs are indicated for the primary or secondary prevention of SCD in patients with severe electrical and/or structural heart disease. Depending on the underlying heart condition, the recommendations on CVR described elsewhere in this guideline apply.336 Health care providers also share these fears, 337 which may limit their exercise prescribing practices. However, studies have shown that physical exercise is safe and is not associated with an increased risk of shocks or other adverse events. 342 In addition, ICD-related complications have not been observed even in competitive athletes. 344One of the main concerns of physical exercise in patients with an artificial pacemaker or ICD is the risk of device-related complications, especially in high-impact activities. In patients with ICDs, there is the added fear of inadvertent activation, which can cause behavioral changes, such as reduced physical activity and participation in moderate-intensity exercise. Nevertheless, before clearing a patient for exercise, the clinician must be aware of the reasons for device placement and become familiar the device\u2019s programming parameters and settings, ideally during the pre-exercise evaluation.342 of 14 studies enrolling 2,681 patients with ICDs showed a beneficial effect of physical exercise on functional capacity in this population, with an average increase in VO 2 of 2.4 ml.kg - 1 .min - 1 . In another meta-analysis, which included five randomized trials and one nonrandomized study in patients with HF and ICDs, 341 a similar improvement in physical capacity was observed, with an increase in VO 2 peak of 1.98 ml.kg - 1 .min - 1 in relation to the control group.A meta-analysis 342 Thus, despite widespread fear of this phenomenon, physical training was not associated with increased ICD activation and proved safe.As for the concern of inadvertent ICD activation during physical training, one meta-analysis found no significant differences. The rate of exercise-associated shocks ranged from 0 to 20% across studies, with an average of 2.2%, similar to the rate of shocks during an exercise-free follow-up period. 345Another meta-analysis actually reported a lower likelihood of shocks during follow-up in patients participating in CVR compared to controls, corroborating the previous result of an observational study, which reported a higher incidence of ICD activation in patients who did not participate in CVR programs. 346 In addition, exercise could reduce myocardial arrhythmogenicity due to remodeling and reduction of sympathetic excitability. 347One possible explanation or the lower incidence of arrhythmias and shocks in patients undergoing CVR would be the improvement of physical capacity, as it has been previously documented that greater physical fitness is associated with a lower incidence of arrhythmia. 348 This provides additional evidence of the safety of stress testing and CVR in this population.In a nationwide study with 10 years of follow-up which included 150 patients with ICDs in a CVR program, all of which completed a CPET or TMET to support exercise prescription, there were only three shock events and all were appropriate. Physical exercise can and should be indicated as long as the patient\u2019s clinical condition is stable and clinical treatment is optimized. In addition to the potential beneficial effects on underlying heart disease, CVR increases physical fitness and can help reduce the incidence of arrhythmias and, consequently, of ICD activation .In patients with implantable devices, the clinician must become familiar with the reason for implant placement, the patient\u2019s ventricular function, whether any arrhythmias are present and, particularly, the device settings and parameters. For patients with an artificial pacemaker, this means understanding the programming mode, the set HR limits, and the type and response of the activity sensor. In patients with an ICD, essential information includes the HR threshold which has been set to trigger shock or burst therapies.In addition to the standard clinical examination, pre-exercise evaluation is of paramount importance in these patients. Ideally, a CPET or TMET should be performed to determine functional capacity and analyze the behavior of the device during exertion. However, the impossibility of performing CPET or TMET should not prevent the practice of physical exercise. In these cases, monitoring during sessions may reveal a need for device reprogramming, usually of maximum HR and sensor response settings.350 However, due to changes in the ECG tracing caused by artificial pacing, automated HR measurement both by ECG telemetry systems and by cardiac monitors may be erroneous. The team should be aware of this potential for error and measure HR manually as needed.During CVR sessions, continuous ECG monitoring can be achieved with the use of telemetry systems. HR control devices, such as regular cardiac monitors, can also be used for monitoring CVR sessions. When prescribing and defining intensity limits for aerobic physical training, one should be aware of ICD programming and limit the intensity accordingly to 10\u201320 bpm below the HR set to trigger therapy (shock or burst). This is especially important in young individuals who experience elevated HR during training. In older patients with HF who are on high-dose beta blockers, the peak HR observed during CPET or TMET is usually below the threshold that triggers ICD therapy.351Patients with an artificial pacemaker may have different chronotropic responses observed on CPET or TMET, which will impact the prescription of aerobic exercise. In addition, the individual\u2019s own pace, type of pacemaker, and presence of a rate sensor will influence the HR response to exertion and, consequently, the exercise prescription. The four possible types of artificial pacemaker response to exertion are as follows:1) Normal or depressed sinus-node chronotropic response. Pacemaker inhibited (not triggered). The chronotropic response to stress is mediated by sinus rhythm and may be normal or depressed (due to sinus node dysfunction and/or drug effect). Ventricular conduction occurs via the own pathway, and the pacemaker is not triggered on exertion. In some cases, it can be triggered at rest and during initial loads, with atrial and/or ventricular pacing. However, during exertion, the pacemaker is inhibited, with a predominance of sinus responses and ventricular conduction via the own pathway. In this type of response to exertion, the intensity of exercise prescription should be based on the usual concerns and is entirely unaffected by the presence of an artificial pacemaker.2) Normal or depressed sinus-node chronotropic response. Pacemaker triggered (activity-initiated ventricular pacing). The chronotropic response to exertion is mediated by the sinus rhythm. Sinus activity is sensed by the pacemaker and triggers synchronized ventricular pacing according to preset paced atrioventricular intervals. In this case, if the maximum pacemaker response limit has been set appropriately for the patient\u2019s sinus response, the exercise prescription may be HR-based, as the ventricle will be paired with sinus activity. However, if the maximum pacemaker response limit is set lower than the patient\u2019s sinus response, dyssynchrony of ventricular pacing and sinus activity will occur at moderate to high exercise intensity. The pacemaker will then block some sinus stimuli by mimicking AV-node Wenckebach activity, a phenomenon known as \u201celectronic Wenckebach\u201d, 352 in order to keep the ventricular HR within the programmed limit; a plateau in the chronotropic response to exertion will ensue. In this scenario, the loss of synchrony between sinus rhythm and ventricular rate will interfere with the utility of HR to guide exercise intensity. The exercise prescription should instead be based on relative loads and/or subjective perceived exertion.When the electronic Wenckebach phenomenon occurs, extreme care is required to detect it during CPET or TMET. It is essential to obtain precise information on the atrial rate at which the pacemaker will initiate 2:1 block, because as this rate is reached, ventricular pacing will occur at a 2:1 ratio, with the potential for a sudden fall in HR on exertion and an abrupt, symptomatic reduction in cardiac output. Therefore, unless the programmed Wenckebach interval and the 2:1 block rate are quite far apart, the HR which triggers electronic Wenckebach may be used as the upper limit for CPET or TMET, as well as for the exercise prescription.In such cases, pacemaker reprogramming to better match the patient\u2019s sinus response should be considered and discussed with the primary physician. Another option, depending on the clinical picture, is the optimization of drug therapy with negative chronotropic agents (such as beta blockers). A reduced sinus response may prevent the aforementioned event.3) Fixed, pacemaker-mediated chronotropic response (no rate responsive pacing). Some patients may have no sinus activity at all, as in atrial fibrillation. In these cases, individuals with complete AV block will be completely dependent on ventricular pacing. If the pacemaker has no rate responsive pacing, or if the sensor is disabled, there will be no chronotropic response to exertion; the pacemaker will be set to a fixed HR. This type of pacemaker and programming is now exceedingly rare. Nevertheless, in such patients, the HR is useless to guide exercise prescription, which should instead be based on relative loads and/or subjective perceived exertion.4) Pacemaker-mediated chronotropic response (rate responsive pacing). In patients with atrial fibrillation and AV block, as previously described, but whose artificial pacemaker has an active sensor with rate responsive pacing, there will be dependence on ventricular pacing, but activation of the sensor by exertion will lead to a pacemaker-mediated chronotropic response. In patients with sinus rhythm, but with a large chronotropic deficit due to sinus node dysfunction and/or drug effects, a chronotropic response to exertion may also occur, mediated by the pacemaker sensor, with atrial pacing followed or not by ventricular pacing.The speed and magnitude of the rate sensor\u2019s response to exertion are programmable, with the possibility of adjusting the sensor activation threshold, the rate of increase in HR to exertion and the rate of reduction during recovery, and the maximal HR limit for the sensor. TCPE or TMET can be used to verify the adequacy of the response and identifying potential needs for pacemaker reprogramming, which should be discussed with the patient\u2019s primary physician.In such cases, as the chronotropic response will be artificially mediated by the device, HR-based prescription of exercise intensity may be inaccurate. Therefore, the use of relative loads and/or perceived exertion is preferred.Pacemakers with accelerometer sensors and axial motion detection, which are the most common, have a sensitive response to walking or running on a treadmill. However, as there is no vertical movement on a cycle ergometer, the sensor is activated very little or not at all. As a result, there is inferior chronotropic response during ciclo ergometer exercise, which may vary according to the individual response of the patient.Resistance training is an important component of CVR for several heart diseases. However, after implantation of a pacemaker of ICD, some care is required until healing is complete, to prevent vascular injury, displacement of the device, and electrode fracture. For instance, caution is recommended when performing weight training and any exercise which involves raising the arms during the first 6 weeks after the implant procedure. In addition, repetitive and intense movements of the limb ipsilateral to the device should be avoided.353However, such guidelines are geared more at patients involved in sports, and are unlikely to be relevant to the exercises carried out in CVR programs. A study of early, supervised shoulder mobilization immediately after artificial pacemaker implantation did not observe any device complications. 356 However, the use of electronic devices in these patients is also increasing, which raises concerns of the possibility of electromagnetic interference.The use of neuromuscular electrical stimulation (NMES) in patients with HF has become widespread, especially for those unable to exercise due to disease severity. NMES can improve aerobic capacity, muscle strength, and cross-sectional area of the quadriceps muscles, and is an effective passive exercise modality in this population. 349 demonstrated that NMES of the quadriceps muscles appears to be safe and feasible in patients with HF and a bipolar sensing ICD. However, the review itself notes that the number of studies and patients included is too small to allow more comprehensive conclusions, and concludes that NMES can be used provided the following conditions are met:A systematic review If individual hazards have been excluded before starting NMES.If NMES is performed only on the quadriceps and gluteal muscles.If treatment is regularly supervised by a doctor, and the device is analysed after every NMES session.Therefore, at the present time, although NMES seems safe to use in patients with bipolar-sensing implantable devices when performed on muscles far from the implant, there is still a need for studies with a larger number of patients to confirm that use is safe and feasible without the need for repeated, detailed device evaluation after sessions.358 In this context, peripheral arterial occlusive disease (PAOD) of the lower limbs is particularly concerning, as, at its most severe stage , it is associated with a high risk of cardiovascular events, lower limb amputation, and death. With the growth of risk factors such as age, diabetes, and smoking, critical ischemia of the lower limbs has become more prevalent, and currently affects about 2 million individuals in the United States alone. 359Stroke has been correctly viewed and addressed as a serious disease with massive impact on public health. However, peripheral artery disease, which is also highly prevalent worldwide and carries high morbidity and mortality rates, affecting more than 40 million individuals in Europe alone, have not been properly addressed, hindering prevention, diagnosis, and effective treatment. 361 The ABI has been recommended as a diagnostic resource prior to use of imaging. 362 Functional tests during exertion effort may be necessary to establish the diagnosis, especially when the ABI is greater than 0.91, as well as for functional classification and exercise prescription in CVR.The presence of PAOD is suspected when there is pain in the lower limbs on exertion, with no apparent musculoskeletal etiology, and the ankle-brachial index (ABI) is <0.90 at rest. Gait can be assessed by means of field tests, which allow the diagnosis of intermittent claudication and determination of the distance walked until onset of symptoms and until development of total loss of function (absolute claudication).363 Another study reported lower cut-off scores, with PAOD being suggested when there is a greater than 18.5% reduction in ABI and a greater than 15 mmHg decrease in BP after exercise. 364A TMET with measurement of the ABI at rest and after exercise has also been proposed as a diagnostic test. The presence of PAOD is suggested by a greater than 20% reduction in post-exercise ABI as compared to resting values, or a decrease in post-exercise BP greater than 30 mmHg as compared to the resting state. 363Considering the overall cardiovascular risk of these patients, optimized clinical treatment should always be instituted. In addition, smoking cessation and drug therapy with statins and antiplatelet agents must be considered, as well as adequate blood glucose and BP control. Regarding the use of cilostazol, there is no consensus among the guidelines of different medical societies. 365 Physical activities performed under direct supervision have been shown to be more effective than unsupervised exercise. 366 In 14 clinical trials , with an intervention duration from 6 weeks to 12 months, pain-free walking increased about 180 meters more in training under direct supervision when compared to training under indirect supervision. Physical training is safe; in most studies, the sole exercise was walking to claudication, at least three times a week, for at least 3 months. 367In symptomatic patients, exercise has the potential to influence morbidity and mortality, reducing symptoms, improving quality of life, and increasing the maximum walking distance . 365 Ph368 although indirect supervision (HBCR) is a good alternative, with positive effects on quality of life and significantly greater improvement in walking tolerance as compared to a simple recommendation to walk. 370In patients with PAOD, training under direct supervision is also superior in terms of cost-effectiveness, 371 It is also worth noting that physical exercise is contraindicated in patients with critical ischemia, but should be considered as soon as possible after successful interventional treatment. 373When walking is not feasible, other activities, such as cycling, resistance training, and use of an upper body ergometers, have also proven effective. 374 Endovascular intervention and open surgery have proven effective in relieving symptoms, increasing walking distance, and improving quality of life, and are indicated when severe symptoms that negatively influence daily life persist despite full clinical treatment .A systematic review of 12 clinical trials including a total of 1,548 patients compared patients who received drug therapy with physical training, endovascular intervention, and open surgery for treatment of claudication. All modalities increased walking distance, reduced symptoms, and improved quality of life. 375 However, after 18 months of follow-up, the functional and quality of life benefits were equivalent in the exercise and revascularization groups, and, in both cases, were superior to those in the group that received medication alone. 376In a randomized clinical trial of 111 patients with aortoiliac disease, the increase in exercise time on a graded treadmill test was greater in the supervised exercise group than in the stent revascularization group. 377Several clinical trials have compared the efficacy and effectiveness of supervised physical exercise, angioplasty, and optimized medical care, using a multitude of different designs. Most trials consisted of two treatment arms. The aforementioned systematic reviews suggested that supervised physical exercise may be superior to optimized medical care or angioplasty. However, these meta-analyses included head-to-head comparisons between two specific treatment arms or used an approach that did not allow inclusion and direct comparison of all available treatments for intermittent claudication. 378For these reasons, a recent meta-analysis sought to establish comparisons between all available treatments in order to elucidate the best management of patients with symptomatic PAOD. The sample included 2,983 participants with intermittent claudication , 54.5% of whom were male. The comparisons were optimized medical care (n = 688), supervised physical training , angioplasty (n = 511), and angioplasty plus supervised physical training (n = 395). The mean follow-up period was 12 months. Compared with optimized medical care alone, angioplasty and supervised physical training outperformed all other therapeutic strategies, with a 290 m gain in maximum walking distance , corresponding to a proportional gain of 141% , with an average follow-up period of 12 months. 378Supervised physical training alone and angioplasty plus supervised physical training again surpassed the other treatment modalities, with an additional gain of 110 m in maximum walking distance , or a proportional gain of 66% . Supervised physical training alone yielded a 180-m gain in maximum walking distance (95% CI: 130 to 230 m), corresponding to a proportional gain of 87% (95% CI: 63 to 111%); this was higher than with angioplasty alone, but lower than with supervised physical training plus angioplasty, in terms of maximum walking distance. 380 In this context, supervised physical training and angioplasty are essential to improve walking distance and quality of life. This recent meta-analysis cited above strongly suggests that supervised physical training associated with angioplasty should be part of first-line treatment, always in the context of optimized drug therapy. The offer of angioplasty without optimized physical training should be avoided whenever possible. 378 However, DAOP treatment centers often offer angioplasty primarily due to the lack of centers focused on supervised physical training. Furthermore, it cannot be neglected that supervised physical training faces resistance on the part of patients themselves, who are often little adherent to treatment, which partly justifies the majority preference in favor of percutaneous treatment. 381These review studies have important implications for clinical practice. This is because all patients with intermittent claudication should receive optimized clinical treatment, in view of the evidence that shows a reduction in future cardiovascular events and an improvement in limb-related outcomes. 382However, recent studies have demonstrated the benefits of combining treatment modalities for symptomatic PAOD, which may increase the likelihood that CVR will become increasingly widespread and accessible. Thus, in addition to optimized medical care, angioplasty combined with supervised physical training seems to be the ideal strategy for initial treatment of patients with intermittent claudication, both to improve maximum walking distance and to improve quality of life. However, the data from these latest reviews cannot confirm whether supervised physical training should be followed by angioplasty or vice versa."} +{"text": "To compare and assess the immediate analgesic effects of conventional and burst transcutaneous electrical nerve stimulation in patients with chronic low back pain. We conducted a three-arm single-blinded randomized controlled trial. A total of 105 patients with non-specific chronic low back pain aged between 18 and 85 years were randomly assigned into the following groups: Placebo Group , Conventional TENS Group (continuous stimulation at 100Hz for 100\u00b5s with sensory intensity), and Burst TENS Group (stimulation at 100Hz modulated at 2Hz for 100\u00b5s with motor-level intensity). All groups received a single application of transcutaneous electrical nerve stimulation for 30 minutes. The outcomes, namely, pain intensity, quality of pain, and pressure pain threshold were measured by the visual analog scale, McGill pain questionnaire, and algometry, respectively. The patients were evaluated before and immediately after the transcutaneous electrical nerve stimulation application. Pain intensity and quality of pain (McGill pain questionnaire score) significantly decreased (p<0.05) in Intervention Groups . A positive effect was observed in the interventions compared to the Placebo Group in all domains of the McGill pain questionnaire (p<0.05), excepting for the pain intensity. Pressure pain threshold significantly increased (p<0.05) immediately after the transcutaneous electrical nerve stimulation application in both Intervention Groups, but not in the Placebo Group. For significant difference was found during assessment when comparing both Intervetion Group. Both transcutaneous electrical nerve stimulation modes were effective for pain modulation. Moreover, there was an increase in the pressure pain threshold. No significant results were found to indicate the best mode for the treatment of chronic low back pain.Clinical Trial Registration: RBR-59YGRB. The notion of non-specific CLBP is often used to describe this condition because the mechanism of pain is poorly understood. The clinical practice guidelines define CLBP as the absence of red flag symptoms for more serious causes of pain with multifactorial pathogenesis.The orthopaedic section of the American Physical Therapy Association defined chronic low back pain (CLBP) as generalized low back pain, which lasts longer than 3 consecutive months. approximately 34,253 cases of back pain (CID10 M54) were registered in Brazil in 2013, being low back pain (LBP) ranked as the fourth common injury. Low back pain was the most frequent type of back pain, and it was one of the main causes of absence from work. Moreover, a systematic review on the prevalence of LBP in Brazil reported high 1-year prevalence rate (>50%) among adults.According to the Brazilian Statistical Yearbook of Work-Related Accidents, Among the possible physical therapy treatments, electrophysical agents as the transcutaneous electrical nerve stimulation (TENS) are widely used for the management of CLBP as a complement to other therapeutic interventions, particularly for exercising. Transcutaneous electrical nerve stimulation is relatively safe, non-invasive, easy to apply, and it provides pain relief during exercise. Different TENS modalities differing in frequency, amplitude, pulse duration, and waveform can be used in clinical practice, such as the conventional or burst modes.Low back pain overloads all health services and various interventions have been established to treat this such as medication, surgery, patient education, behavioral therapy, and physiotherapy.,6-10 however, the optimal parameters in CLBP treatment are still unknown. When TENS is applied with high frequency, such as in the conventional mode, the physiological intention is to produce A\u03b2 fiber depolarization effect, capable of inhibit transmission of nociceptive information for spinal cord. When TENS is applied at low frequency and strong stimuli, such as burst mode, the physiological intention is depolarizes the fast pain (A\u03b4) fibers capable to activation of descending analgesia. In addition, both TENS activate opioid receptors in the central nervous system, which induces analgesia e.g., such serotonin and M\u03bc or delta opioid receptors, when low or high frequency TENS is applied, respectively.,10Studies have reported that TENS can reduce acute and chronic pain of different etiologies, A systematic review on TENS efficacy in CLBP was conducted in four high-quality randomized controlled clinical trials (585 patients) wherein only three showed pain relief compared to the Placebo Group (PG). Due to the small number of studies, it was not possible to find supporting evidence on the efficacy of this procedure in patients with LBP. Hence, no evidence on the appropriate parameters and stimulation modes was found. Jauregui et al., conducted a meta-analysis and concluded that treatment using TENS resulted in not only reduced pain, but also reduced drug intake among patients with CLBP treated for less than 5 months.Despite the extensive use, there is still no consensus on the actual effectiveness of TENS to individuals with LBP. evaluated TENS use in the management of acute pain caused by multiple etiologies wherein 19 studies were reviewed . The authors were able to assess the analgesic effect of TENS in six studies and found that TENS was 3 times more effective than the placebo. However, they found low to moderate evidence due to the small number of patients and high risk of biased results. An editorial by Johnson et al., suggested that most studies on the use of TENS were inconclusive and of insufficient quality. In addition, a meta-analysis by Resende et al., reported the necessity of more quality studies on the effects of TENS on CLBP patients considering factors, such as stimulation intensity, pain, incapacity and functionality assessment, and time for evaluating results.Johnson et al.,-10 pointed to studies comparing a single TENS parameter with PG or with other therapies. Of the studies cited in these reviews, only one compared three types of TENS , and during one month of treatment no differences were found between groups. However, this study did not describe the exact parameters of application. No studies have compared different parameters of TENS application in CLBPAll systematic reviews,14 most of them only report the long-term effects. However, the main therapeutic goal during a physical therapy session is an immediate analgesic effect-either to enable patients to practice physical exercises or to relieve pain after a therapy session.Although there are several studies on the effect of TENS on LBP,To compare and assess the immediate analgesic effects of conventional and burst transcutaneous electrical nerve stimulation in patients with chronic low back pain.A three-arm single-blinded randomized controlled trial was conducted. The study was approved by the Research Ethics Committee, CAAE: 44642615.2.0000.0102, number 1.145.540, and prospectively registered on ensaiosclinicos.gov.br (RBR-59YGR8). All patients signed a consent form before the study begins.Universidade Federal do Paran\u00e1 (UFPR) physical therapy laboratory and ambulatory of the Hospital das Cl\u00ednicas, Curitiba, PR, Brazil.The present study was conducted in the without irradiation, and with a minimum pain intensity of three on the Numeric Pain Rating Scale (NPRS).The inclusion criteria included patients aging 18-85 years with a primary diagnosis consistent with non-specific LBP (LBP not otherwise specified that lasted more than 3 months)e.g., disc herniation, spinal stenosis, vertebral fracture, infection, back-related tumor, osteoporosis, rheumatoid arthritis, inflammatory processes, pregnancy, and kidney problems); previous surgery in the lumbar region; skin lesions; presence of pacemaker; using medications for relief from LBP 48h before our intervention; and those currently undergoing some treatment for the LBP.The exclusion criteria were as follows patients with specific causes of pain , Conventional TENS Group (CTG) (n=5), and Burst TENS Group (BTG) (n=5). A total of 7 blocks were inserted including 105 participants that were blinded to the intervention. The intervention was conducted by one physiotherapist who was unaware of the object of the study.After the baseline assessment, the participants were randomized using random blocks. The CTG was administered using a continuous stimulation at a high frequency (100Hz) with pulse duration (100\u00b5s), and sensory intensity. The BTG (acupuncture-like TENS was administered in the BTG using a 100Hz modulated at 2Hz (burst mode) and a pulse duration of 100\u03bcs with motor-level intensity. Transcutaneous electrical nerve stimulation was applied only once for 30 minutes, and reassessed was performed immediately following the intervention. The interval between assessment and reassessment was 30 minutes. The procedures for the PG were similar to those of the other groups. However, the current amplitude/intensity did not increase. Participants were informed that they may or may not experience any sensation on the site of application of the electrodes.Electrical stimulation was applied using the same device previously calibrated by the MedMart (Brazil). The participants were placed in a prone position and instructed to relax the musculature while the procedures were performed. Four rectangular surface (90x50mm) electrodes were bilaterally placed at the level of the L3 and L5 spinous process . With this quadripolar placement, the current in each channel crossed the area experiencing the pain. After cleansing the skin with rubbing alcohol 70%, carbon-impregnated silicone rubber, and flexible electrodes were placed using an electroconductive gel and fixed with an adhesive tape.One blinded trained physiotherapist examined all the participants prior to and immediately after the intervention. The measured outcomes were pain intensity, quality of pain, and pressure pain threshold (PPT).The participants were instructed to point out what best represented the intensity of their pain using a 10cm straight line on the NPRS . of the McGill pain questionnaire (MPQ) contains 78 descriptors divided into 4 subclasses of pain quality . The descriptors were divided into 20 subcategories, each containing 4 to 6 words. Patients had to choose one or none of the words in all subcategories, and the numeric index was calculated by summing the number of chosen options, with a maximum value of 20. The Brazilian-Portuguese Long Form-MPQ was cross-culturally adapted to Brazilian Portuguese and clinimetrically tested in a previous study.The Brazilian-Portuguese Long Form2 in diameter and with an application rate of 0.5kgf/cm2/s.The PPT was measured using an algometer with the tip measuring 1cm However, in our study, the single blinded physiotherapist conducted a preliminary intra-observer reliability evaluation. She evaluated 10 individuals to assess the intra-test reliability, with an interval of at least 48 hours. The intra-test (ICC) analysis indicated excellent reliability for the intervention results (ICC=0.99).Pressure algometry results have previously shown good validity. previous marked with a dermatographic pencil: 5cm to the left and right spinous process of the L3 and L5 vertebra. Two more points were used as a control and marked 5cm lateral to the tibial tuberosity in the left and right tibialis anterior muscle. After marking the points, the tip of the algometer was positioned perpendicular to the skin and gradually pressed at a rate of 0.5kgf/cm2/s, starting with zero kgf/cm2/s. The participants were instructed to say \u201cstop\u201d when the pressure of the algometer\u2019s rubber tip caused pain. Then, this value was registered by the researchers and represented the PPT at the time. After 1 minute, two more measurements were performed at each point, and the average was calculated for each point.The PPT was measured bilaterally at the following pointsAll statistical analyses were performed using the SPSS 25.0. Data were presented as mean\u00b1standard deviation of the mean and they were subjected to analysis of the sphericity and homogeneity of variance using the Mauchly\u2019s and Levene\u2019s tests, respectively. The confidence interval was set at 95% for all analyses. To analyze the differences between and within groups, analysis of variance with repeated measures was used for parametric values. For non-parametric data, the Wilcoxon signed-rank test was used to verify the difference within groups and the Kruskal-Wallis test was used to establish the differences between groups. The level of significance was set at p<0.05. a statistical power of 95% (1\u03b2 error probability), an \u03b1 error level probability of 0.05, and a possible sample loss of up to 5% were considered. Considering these calculations, we included 35 patients per group .The program GPower 3.0 was used to calculate the sample size. The statistical power was set at 0.95, a moderate effect size (0.35),A total of 105 patients were evaluated from August 2016 to August 2017. No participants left the study before its completion. The consolidated standards of reporting trials diagram is shown in This study aimed to assess the immediate analgesic effects of Conventional and Burst TENS modes in patients with CLBP, which is assessed by using the NPRS, MPQ, and pressure algometry.To the best of our knowledge, this is the first study which compares the immediate analgesic effect of two different modes of TENS in CLBP. Immediate pain relief is necessary to enable patients to perform rehabilitation exercises, as pain can be a limiting factor or intense pain that may occur following a therapy session.i.e., high frequencies (more than 50Hz) stimulate \u03b4-opioid receptors and low frequencies (less than 10Hz) activate \u00b5-opioid receptors. Since the analgesic pathways are not the same, it was expected that the responses would also be different. However, the difference of responses was not noted. This data corroborates with a study conducted by Cheing et al., wherein TENS was applied in individuals with chronic pain and a decrease in the pain pattern that was verified through the VAS score after a single application. Tousignant-Laflamme et al., compared the outcomes of 1st round of TENS acupuncture applied for different time periods (15 minutes and 30 minutes) in 11 patients with CLBP and found no difference in the VAS score between the groups. However, no comparison with the PG was performed.In the present study, no difference in NPRS results was identified between the TENS Groups. Thus, Conventional and Burst TENS modes were equally effective in providing immediate pain relief. High- and low-frequency TENS promote analgesia through the release of endogenous opioids at the spinal level and rostral ventral medulla. The different stimulation frequencies led to the release of different opioids, evaluated the analgesic effects of TENS and dynamic currents on the PPT of patients with CLBP and found an increase in PPT in the TENS Group. In a trial conducted by Giesbrecht et al., the PPT in LBP patients and healthy individuals was compared. The results revealed that the Group with LBP had a lower PPT, which indicates local sensitization. According to the study, central sensitization was enhanced by the activation of small-diameter primary nociceptors (A\u03b2 fibers), allowing low-intensity stimuli to be perceived as pain and causing long-term changes in the spinal neurons.In the assessment by pressure algometry, positive results were observed, with an increase in the PPT ranging from 8% to 15% in groups treated with TENS compared with a 2% to 5% reduction in the PG. Ebadi et al., assessed generalized hyperalgesia in individuals with LBP with intervertebral disc herniation. In the algometry assessment, patients with LBP had a lower PPT than the Control Group.A study by O\u2019Neill et al., healthy individuals were assessed before, immediately after, and 20 and 60 minutes after TENS application. They were divided into four groups received different frequencies of electrical stimulation and a PG. The hypothenar region was the point used for algometry analysis and the elbow for the electrical stimulation. There were no significant differences at any point of assessment.Furthermore, in a study by Gomes et al.,A trial with 240 healthy subjects assessed the effect of TENS with different frequencies (4-110Hz), intensities (varying between the \u201ctolerable\u201d and \u201cstrong but comfortable\u201d), and site of application . The pulse duration was fixed at 200\u00b5s with an application duration of 30 minutes. The algometer was applied in the first dorsal interosseous muscle. The patients were assessed immediately before the application of the intervention, during the application (every 10 minutes), immediately after the application of the intervention, and at every 10 minutes for the first 60 minutes after the intervention. Therefore, an increase in the PPT caused by the analgesic effects of TENS may be possibly observed, indicating pain improvement.Initially, there was an increase in the PPT when low-frequency stimulation was applied with high intensity maintained for 30 minutes after the stimulation. In contrast, immediate improvement was noted with high-frequency high-intensity stimulation; however, the improvement was not maintained.The main strength of our study is the immediate decrease in the intensity of LBP on application of TENS, which has great relevance in the physical therapists clinical practice. Randomization of participants and comparison with the PG increase the quality of the study and reinforce its applicability. One of the limitations of the study is the lack of assessment after 24 or 48 hours following a single application of the current to verify its effectiveness for pain reduction, and other is that we do not use the Global Perceived Effect Scale that was used to assess the patients\u2019 global perception of improvement and which is important to evaluate the improvements after a physiotherapy program.The two modes of transcutaneous electrical nerve stimulation application, conventional and burst, were effective to modulate pain and increase pain threshold pressure among individuals with chronic low back pain. However, no significant results were found to indicate the best treatment for chronic low back pain. O conceito de DLC n\u00e3o espec\u00edfica \u00e9 geralmente utilizado para descrever esta condi\u00e7\u00e3o devido a falta de entendimento dos mecanismos da dor. As orienta\u00e7\u00f5es cl\u00ednicas definem tais mecanismos como a aus\u00eancia de sintomas de bandeira vermelha para causas mais graves de dor com patog\u00eanese multifactorial.A divis\u00e3o de ortopedia da Associa\u00e7\u00e3o Americana de Fisioterapia define dor lombar cr\u00f4nica (DLC) como dor lombar generalizada com dura\u00e7\u00e3o de mais de 3 meses consecutivos. aproximadamente 34.253 casos de dorsalgia (CID10 M54) foram registrados no Brasil em 2013, sendo a dor lombar (DL) classificada como a quarta les\u00e3o mais comum. Ainda, a lombalgia \u00e9 indicada como tipo de DL mais frequente, e uma das principais causas de aus\u00eancia ao trabalho. Al\u00e9m disso, uma revis\u00e3o sistem\u00e1tica sobre a preval\u00eancia de DL no Brasil relatou elevada taxa de preval\u00eancia de 1 ano (>50%) desta condi\u00e7\u00e3o entre adultos.De acordo com o Anu\u00e1rio Estat\u00edstico Brasileiro de Acidentes do Trabalho, Entre os poss\u00edveis tratamentos fisioterap\u00eauticos, os agentes eletrof\u00edsicos, tais como a estimula\u00e7\u00e3o el\u00e9ctrica transcut\u00e2nea do nervo (TENS), s\u00e3o amplamente utilizados no manejo da DL para complementar outras interven\u00e7\u00f5es terap\u00eauticas, especialmente a pr\u00e1tica de exerc\u00edcios. A estimula\u00e7\u00e3o do nervo el\u00e9trico transcut\u00e2neo \u00e9 relativamente segura, n\u00e3o invasiva, f\u00e1cil de aplicar e promove al\u00edvio da dor durante o exerc\u00edcio. Diferentes modalidades de TENS variando em termos de frequ\u00eancia, amplitude, dura\u00e7\u00e3o do pulso e forma de onda podem ser utilizadas na pr\u00e1tica cl\u00ednica, tais como os modos convencional ou burst.As admiss\u00f5es por DL sobrecarregam todos os servi\u00e7os de sa\u00fade, sendo que v\u00e1rias interven\u00e7\u00f5es t\u00eam sido estabelecidas para tratamento da les\u00e3o, tais como medica\u00e7\u00e3o, cirurgia, educa\u00e7\u00e3o do paciente, terapia comportamental e fisioterapia.,6-10 contudo, os par\u00e2metros ideais de tratamento DLC ainda n\u00e3o s\u00e3o conhecidos. Quando a TENS \u00e9 aplicada com alta frequ\u00eancia, tal como no modo convencional, a inten\u00e7\u00e3o fisiol\u00f3gica \u00e9 produzir A\u03b2 um efeito de despolariza\u00e7\u00e3o da fibra capaz de inibir a transmiss\u00e3o de informa\u00e7\u00e3o nociceptiva para a medula espinal. Quando a TENS \u00e9 aplicada com baixa frequ\u00eancia e por meio de fortes est\u00edmulos, tais como o modo de ruptura, a inten\u00e7\u00e3o fisiol\u00f3gica \u00e9 despolarizar as fibras de dor r\u00e1pida (A\u03b4) capazes de ativar a analgesia descendente. Al\u00e9m disso, ambas as TENSs ativam receptores opioides no sistema nervoso central que induzem a analgesia, por exemplo, de receptores de serotonina e de M\u00b5 ou delta opioide, quando TENS de baixa ou alta frequ\u00eancia \u00e9 aplicada.,10Estudos t\u00eam relatado que a TENS pode reduzir a dor aguda e cr\u00f4nica de diferentes etiologias, Uma revis\u00e3o sistem\u00e1tica sobre a efic\u00e1cia das TENSs em DLC incluindo quatro ensaios cl\u00ednicos controlados randomizados de alta qualidade (585 pacientes) em que observou-se al\u00edvio da dor em apenas tr\u00eas pacientes em compara\u00e7\u00e3o ao Grupo Placebo (GP). Devido aos poucos estudos, n\u00e3o foi poss\u00edvel encontrar evid\u00eancias sobre a efic\u00e1cia deste procedimento em pacientes com DL. Consequentemente, n\u00e3o foram encontradas indicativos sobre os par\u00e2metros e modos de estimula\u00e7\u00e3o adequados. Jauregui et al., conduziram metan\u00e1lise e conclu\u00edram que o tratamento com a TENS resultou n\u00e3o s\u00f3 na redu\u00e7\u00e3o da dor, mas tamb\u00e9m na redu\u00e7\u00e3o do uso de medicamentos entre pacientes com DLC tratados por menos de 5 meses.Apesar da sua ampla utiliza\u00e7\u00e3o, ainda n\u00e3o existe consenso sobre a real efic\u00e1cia das TENSs em indiv\u00edduos com DL. avaliaram o uso de TENS no tratamento da dor aguda causada por m\u00faltiplas etiologias em revis\u00e3o incluindo 19 estudos (1.346 pacientes). Os autores conseguiram avaliar o efeito analg\u00e9sico da TENS em seis estudos e observaram que a TENS era 3 vezes mais eficaz do que o placebo. No entanto, encontraram evid\u00eancia baixa a moderada devido a pequena amostra de pacientes e elevado risco de vi\u00e9s nos resultados. Em editorial Johnson et al., sugeriram que a maioria dos estudos sobre a utiliza\u00e7\u00e3o de TENS eram inconclusivos e de qualidade insuficiente. \u00c9 importante mencionar que a metan\u00e1lise de Resende et al., relatou a necessidade de mais estudos qualitativos sobre os efeitos das TENS em paciente com DLC, que considerem fatores como intensidade da estimula\u00e7\u00e3o, dor, avalia\u00e7\u00e3o da incapacidade e funcionalidade, e tempo da avalia\u00e7\u00e3o dos resultados.Johnson et al.,-10 apontam para estudos que compararam \u00fanico par\u00e2metro da TENS com o GP ou com outras terapias. Dos estudos citados nessas revis\u00f5es, apenas um comparou tr\u00eas tipos de TENS ao longo de um m\u00eas de tratamento e n\u00e3o foram encontradas diferen\u00e7as entre os grupos. Contudo, esse estudo n\u00e3o descreveu os par\u00e2metros exatos de aplica\u00e7\u00e3o. H\u00e1 escassez na literatura de estudos que comparem diferentes par\u00e2metros de aplica\u00e7\u00e3o das TENSs em DLC.Todas as revis\u00f5es sistem\u00e1ticas,14 a maioria deles apenas relata os efeitos de longo prazo. Contudo, o principal objetivo terap\u00eautico durante uma sess\u00e3o de fisioterapia \u00e9 alcan\u00e7ar efeito analg\u00e9sico imediato \u2013 tanto para permitir aos pacientes a pr\u00e1tica de exerc\u00edcios f\u00edsicos quanto para aliviar a dor ap\u00f3s a sess\u00e3o terap\u00eautica.Embora existam v\u00e1rios estudos sobre o efeito da TENS na DL,Comparar e avaliar o efeito analg\u00e9sico da estimula\u00e7\u00e3o el\u00e9trica nervosa transcut\u00e2nea convencional e do burst em pacientes com dor lombar cr\u00f4nica.Foi conduzido ensaio controlado, randomizado, de tr\u00eas bra\u00e7os em um \u00fanico centro. O estudo foi aprovado pelo Comit\u00ea de \u00c9tica em Pesquisa, CAAE: 44642615.2.0000.0102 e n\u00famero 1.145.540, e registrado prospectivamente em ensaiosclinicos.gov.br (RBR-59YGR8). Todos os pacientes assinaram o termo de consentimento antes de iniciar o estudo.Estudo realizado no laborat\u00f3rio de reabilita\u00e7\u00e3o da Universidade Federal do Paran\u00e1 (UFPR) e ambulat\u00f3rio do Hospital das Cl\u00ednicas, Curitiba, PR, Brasil. sem irradia\u00e7\u00e3o e com intensidade de dor m\u00ednima de tr\u00eas na Escala Num\u00e9rica de Dor (END).Os crit\u00e9rios de inclus\u00e3o permitiram o ingresso de pacientes com idade entre 18 e 85 anos, com diagn\u00f3stico prim\u00e1rio consistente de DL n\u00e3o espec\u00edfica (DL n\u00e3o especificada com dura\u00e7\u00e3o de n\u00e3o mais de 3 meses),Os crit\u00e9rios de exclus\u00e3o compreenderam paciente com causa de dor espec\u00edfica ; cirurgia anterior na regi\u00e3o lombar, les\u00f5es na pele, presen\u00e7a de marcapasso, utilizando medicamento para melhora da DL 48 horas antes da interven\u00e7\u00e3o, e \u00e0queles durante a realiza\u00e7\u00e3o do estavam realizando algum tipo de tratamento para DL. Cada bloco continha 15 participantes distribu\u00eddos randomicamente: GP (n=5), Grupo TENS Convencional (GTC) (n=5) e Grupo TENS Burst (GTB) (n=5). Foram inseridos sete blocos, totalizando 105 participantes eram \u201ccegos\u201d a interven\u00e7\u00e3o. O fisioterapeuta que conduziu a interven\u00e7\u00e3o desconhecia os objetivos do estudo.Ap\u00f3s a avalia\u00e7\u00e3o por linha de base, os pacientes foram randomizados utilizando t\u00e9cnica de bloco de randomiza\u00e7\u00e3o. O GTC foi administrado utilizando uma simula\u00e7\u00e3o cont\u00ednua em alta frequ\u00eancia (100Hz) com dura\u00e7\u00e3o da pulsa\u00e7\u00e3o (100\u00b5s) e intensidade sensorial. O GTB (TENS acupuntura) foi administrado adotando 100Hz modulado em 2Hz (modo burst) e com dura\u00e7\u00e3o da pulsa\u00e7\u00e3o de 100\u03bcs com intensidade de n\u00edvel motor. O TENS foi aplicado por apenas 30 minutos, e reavalia\u00e7\u00e3o foi feita imediatamente ap\u00f3s a interven\u00e7\u00e3o. O tempo de intervalo entre avalia\u00e7\u00e3o e reavalia\u00e7\u00e3o foi de 30 minutos. Os procedimentos para GP foram similares aqueles de outros grupos, por\u00e9m a amplitude/intensidade atual n\u00e3o foi aumentada. Os pacientes foram informados que poderiam ou n\u00e3o sentir qualquer sensa\u00e7\u00e3o no local da aplica\u00e7\u00e3o dos eletrodos.Estimula\u00e7\u00e3o el\u00e9trica foi aplicada utilizando o mesmo dispositivo anteriormente calibrado por MedMart (Brasil). Os participantes foram colocados em posi\u00e7\u00e3o prona e instru\u00eddos para relaxar a musculatura enquanto os procedimentos eram realizados. Quatro eletrodos retangulares de superf\u00edcie (90x50mm) foram bilateralmente posicionados no n\u00edvel L3 e L5 processo espinhoso . Com posicionamento quadripolar, a corrente em cada canal cruzava a \u00e1rea dolorosa. Ap\u00f3s limpeza da pele com \u00e1lcool 70%, eletrodos de borracha de silicone impregnada com carbono e flex\u00edveis foram posicionado com fita adesiva e utilizando gel eletrocondutor.Um fisioterapeuta treinado examinou de forma cega todos os participantes antes e imediatamente ap\u00f3s a interven\u00e7\u00e3o. Os resultados medidos foram dor intensa, qualidade da dor, e limiar de dor \u00e0 press\u00e3o (LDP).Os participantes foram instru\u00eddos a fazer marca\u00e7\u00e3o no ponto que melhor representava a intensidade das dores utilizando linha reta de 10cm na END . do question\u00e1rio de dor de McGill (MPQ) para l\u00edngua portuguesa cont\u00e9m 78 descri\u00e7\u00f5es dividas em 4 subclasses de qualidade de dor . Os descritores foram divididos em 20 subcategorias, cada uma contendo de 4 a 6 palavras. Os pacientes que tinham de escolher uma ou nenhuma das palavras em todas as subcategorias, e o \u00edndice num\u00e9rico foi calculado por meio da soma do n\u00famero de op\u00e7\u00f5es escolhidas, com valor m\u00e1ximo de 20. A vers\u00e3o brasileira do MPQ foi adaptada transculturalmente para o portugu\u00eas brasileiro e clinimetricamente testada em estudos anteriores.A vers\u00e3o brasileira2 de di\u00e2metro e uma taxa de aplica\u00e7\u00e3o de 0,5kgf/cm2/s.O LDP foi medido por meio de alg\u00f4metro com a ponta medindo 1cm Por\u00e9m, em nosso estudo, uma fisioterapeuta conduziu a avalia\u00e7\u00e3o preliminar de confiabilidade entre avaliadores. Essa professional avaliou 10 indiv\u00edduos para medir confiabilidade intrateste, com um intervalo de pelo menos 48 horas. A an\u00e1lise intrateste (ITT) indicou confiabilidade excelente para resultados da interven\u00e7\u00e3o .Os resultados da algometria de press\u00e3o tem mostrado boa validade. previamente marcados com l\u00e1pis dermatogr\u00e1fico: 5cm do processo espinhoso para esquerda e direita da vertebra L3 e L5. Al\u00e9m disso, outros dois pontos foram utilizados como controle e marcados 5cm lateral a tuberosidade da t\u00edbia \u00e0 esquerda e direita do m\u00fasculo tibial anterior. Ap\u00f3s marca\u00e7\u00e3o dos pontos, a ponta do alg\u00f4metro foi posicionada perperdincular a pele e pressionada gradualmente em taxa de 0,5kgf/cm2/s, iniciando com zero kgf/cm2/s. Os participantes foram instru\u00eddos para falar \u201cpare\u201d quando a press\u00e3o com a ponta emborrachada do alg\u00f4metro causasse dor. Ap\u00f3s, este valor foi registrado por pesquisadores e representava o LDP do momento. Ap\u00f3s 1 minuto, duas medi\u00e7\u00f5es adicionais foram realizadas em cada ponto, sendo a m\u00e9dia calculada para cada um dos pontos.O LDP foi medido bilateralmente nos pontosTodas as an\u00e1lises estat\u00edsticas foram realizadas utilizando o SPSS vers\u00e3o 25.0. Os dados foram apresentados como m\u00e9dias\u00b1desvio padr\u00e3o da m\u00e9dia e estavam sujeitos a an\u00e1lise de esfericidade e homocedasticidade da varia\u00e7\u00e3o por meio dos testes de Mauchly e Levene, respectivamente. O intervalo de confian\u00e7a foi estabelecido em 95% para todas as an\u00e1lises. A avalia\u00e7\u00e3o das diferen\u00e7as entre e dentre os grupos e an\u00e1lise de vari\u00e2ncia com medidas repetidas foram utilizadas para estimar os valores param\u00e9tricos. Para os dados n\u00e3o-param\u00e9tricos, o teste de rank Wilcoxon assinado foi adotado para o teste de diferen\u00e7a entre os grupos, e o teste de Kruskal-Walllis para diferen\u00e7as entre os grupos. O n\u00edvel de signific\u00e2ncia foi estabelecido em p<0,05. signific\u00e2ncia estat\u00edstica de 95% (1\u03b2 erro de probabilidade), erro \u03b1 n\u00edvel de probabilidade de 0,05 e amostra poss\u00edvel de perda de at\u00e9 5%. Consequentemente, 35 pacientes por grupo foram necess\u00e1rios.Empregou-se o programa GPower 3.0 para calcular o tamanho da amostra. A signific\u00e2ncia estat\u00edstica foi estabelecida em 0,95, tamanho de efeito moderado ,Um total de 105 pacientes foram avaliados entre agosto de 2015 e agosto de 2017. Nenhum participante desistiu de integrar o estudo. O diagrama dos padr\u00f5es de consolida\u00e7\u00e3o do relato de ensaio \u00e9 mostrado na A A A Burst em pacientes com DLC avaliados que utilizaram END, MPQ, e algometria de press\u00e3o.Este estudo avaliou os efeitos analg\u00e9sicos imediatos dos modos TENS Convencional e Pelo conhecimento dos autores, atualmente este \u00e9 o primeiro estudo da que comparou os efeitos analg\u00e9sicos imediatos de dois modos diferentes de TENS em DLC. O al\u00edvio imediato da dor \u00e9 necess\u00e1rio para permitir aos pacientes a realizar exerc\u00edcios de reabilita\u00e7\u00e3o, j\u00e1 que a dor pode ser um fator limitante ou intenso que pode ocorrer durante a sess\u00e3o de terapia.Burst foram igualmente efetivos em providenciar al\u00edvio imediato da dor. O TENS de alta e baixa frequ\u00eancia promove analgesia por meio da libera\u00e7\u00e3o de opioides end\u00f3genos no n\u00edvel espinhal e medula rostral ventral. As frequ\u00eancias de diferentes est\u00edmulos levaram a libera\u00e7\u00e3o de opiodes diferentes, ou seja, alta frequ\u00eancias (maior de 50Hz) est\u00edmulo de receptores \u03b4-opiodes e baixa frequ\u00eancias (menor de 10Hz) receptores de \u00b5-opioide ativo. Considerando que os caminhos analg\u00e9sicos n\u00e3o s\u00e3o os mesmos, esperou-se que as respostas tamb\u00e9m poderiam ser distintas. Contudo, tal distin\u00e7\u00e3o n\u00e3o foi observada. Esses dados corroboram com estudo conduzido por Cheing et al., em que TENS foi aplicado em indiv\u00edduos com dor cr\u00f4nica e redu\u00e7\u00e3o do padr\u00e3o da dor verificado por meio do escore EVA ap\u00f3s aplica\u00e7\u00e3o \u00fanica. Tousignant-Laflamme et al., comparou os resultados da 1\u00aa rodada de aplica\u00e7\u00e3o de TENS acupuntura em per\u00edodos com tempos diferentes (15 e 30 minutos) em 11 pacientes com DLC e n\u00e3o encontrou diferen\u00e7as no escore VAS entre os grupos. Por\u00e9m, n\u00e3o houve compara\u00e7\u00e3o com GP.Neste estudo, n\u00e3o identificou-se diferen\u00e7a significativa nos resultados da END entre os Grupos TENS. Portanto, os modos TENS Convencional e avaliou os efeitos analg\u00e1sicos de TENS e correntes din\u00e2micas no LDP no Grupo TENS. Em um ensaio conduzido por Giesbrecht et al., o LDP em pacientes com DL foi comparado ao de pacientes saud\u00e1veis. Os resultados revelaram que o grupo com DL teve menor LDL, que indica sensibilidade no local. De acordo com estudo, a sensibiliza\u00e7\u00e3o central foi melhorado por ativa\u00e7\u00e3o de nociceptores prim\u00e1rios de menor di\u00e2metro (fibras A\u03b2), permitindo baixa-intensidade de estimulo para ser percebido como dor e causa de mudan\u00e7as de curta dura\u00e7\u00e3o em neur\u00f4nios espinhais.Na avalia\u00e7\u00e3o por algometria por press\u00e3o, os resultados positivos foram observados, com um aumento no LPD variando de 8% para 15% em grupos tratrados com TENS comparado com uma redu\u00e7\u00e3o de 2% a 5% no GP. Ebaldi et al., avaliou hiperalgesia generalizada em indiv\u00edduos com DL com hernia\u00e7\u00e3o de disco intervertebral. Na avalia\u00e7\u00e3o de algometria os pacientes com DL apresentaram LDP menor do que o Grupo Controle.Um estudo de O\u2019Neil et al., indiv\u00edduos saud\u00e1veis foram avaliados antes, imediatamente ap\u00f3s, e de 20 a 30 minutos ap\u00f3s a aplica\u00e7\u00e3o de TENS. Os participantes foram divididos em grupos de quatro que receberam frequ\u00eancias diferentes de est\u00edmulo el\u00e9trico e um GP. A regi\u00e3o hipot\u00eanar foi o ponto utilizado por an\u00e1lise de algometria e cotovelo por estimula\u00e7\u00e3o el\u00e9trica. N\u00e3o observou-se diferen\u00e7as significativas em qualquer um dos pontos de avalia\u00e7\u00e3o.Al\u00e9m disso, no estudo de Gomes et al.,Um ensaio de 240 indiv\u00edduos saud\u00e1veis avaliou os efeitos de TENS com diferentes frequ\u00eancias (4-110Hz), intensidades , e local de aplica\u00e7\u00e3o . A dura\u00e7\u00e3o de pulso foi fixada em 200\u00b5s com uma aplica\u00e7\u00e3o de dura\u00e7\u00e3o de 30 minutos. O alg\u00f4metro foi aplicado no primeiro inter\u00f3sseo dorsal. Os pacientes foram avaliados imediatamente anterior a aplica\u00e7\u00e3o da interven\u00e7\u00e3o, durante a aplica\u00e7\u00e3o (cada 10 minutos), imediatamente ap\u00f3s a aplica\u00e7\u00e3o da interven\u00e7\u00e3o, e cada 10 minutos ao longo dos primeiros 60 minutos ap\u00f3s a interven\u00e7\u00e3o. Portanto, um aumento de LDP causado por efeitos analg\u00e9sicos de TENS pode ser possivelmente observado, indicando melhora da dor.Inicialmente, houve um aumento no LDP quando est\u00edmulo de baixa frequ\u00eancia foi aplicado com alta intensidade e mantida por cerca de 30 minutos ap\u00f3s o est\u00edmulo. Em posi\u00e7\u00e3o, a melhora imediata foi notada com alta frequ\u00eancia com estimulo de alta intensidade; por\u00e9m, a melhora n\u00e3o foi mantida.O principal ponto forte deste estudo foi a redu\u00e7\u00e3o imediata na intensidade do DL nas aplica\u00e7\u00f5es de TENS, que foi de maior relev\u00e2ncia na pr\u00e1tica cl\u00ednica de terapeutas f\u00edsicos. A randomiza\u00e7\u00e3o dos participantes e compara\u00e7\u00f5es com GP aumentou a qualidade do estudo e refor\u00e7ou sua aplicabilidade. Uma das limita\u00e7\u00f5es do nosso estudo \u00e9 a falta de avalia\u00e7\u00e3o 24 ou 48 horas ap\u00f3s a aplica\u00e7\u00e3o \u00fanica de corrente para verificar sua efetividade para redu\u00e7\u00e3o de dor e outra quest\u00e3o \u00e9 a n\u00e3o ado\u00e7\u00e3o da Escala de Percep\u00e7\u00e3o do Efeito Global que foi utilizada para avaliar a percep\u00e7\u00e3o global de melhoria dos pacientes e foi importante para avaliar as melhorias ap\u00f3s o programa de fisioterapia.burst foram efetivos para modular a dor e aumentar o limiar de dor \u00e0 press\u00e3o entre indiv\u00edduos com dor lombar cr\u00f4nica. Contudo, n\u00e3o se encontrou resultados significantes para indicar o melhor tratamento para dor lombar cr\u00f4nica.Os dois modos de aplica\u00e7\u00e3o de estimula\u00e7\u00e3o el\u00e9trica nervosa transcut\u00e2nea convencional ou"} +{"text": "Over 1250 data points from the Devonian Duvernay shale were utilized to create and validate the model. These datasets were obtained from three wells; the first was used to train the models, while the data sets from the other two wells were utilized to test and validate them. Support vector machine (SVM), random forest (RF), and decision tree (DT) were the ML approaches tested, and their predictions were contrasted with three empirical correlations. Various AI methods' parameters were tested to assure the best possible accuracy in terms of correlation coefficient (R) and average absolute percentage error (AAPE) between the actual and predicted TOC. The three ML methods yielded good matches; however, the RF-based model has the best performance. The RF model was able to predict the TOC for the different datasets with R values range between 0.93 and 0.99 and AAPE values less than 14%. In terms of average error, the ML-based models outperformed the other three empirical correlations. This study shows the capability and robustness of ML models to predict the total organic carbon from readily available logging data without the need for core analysis or additional well interventions.Unconventional resources have recently gained a lot of attention, and as a consequence, there has been an increase in research interest in predicting total organic carbon (TOC) as a crucial quality indicator. TOC is commonly measured experimentally; however, due to sampling restrictions, obtaining continuous data on TOC is difficult. Therefore, different empirical correlations for TOC have been presented. However, there are concerns about the generalization and accuracy of these correlations. In this paper, different machine learning (ML) techniques were utilized to develop models that predict TOC from well logs, including formation resistivity (FR), spontaneous potential (SP), sonic transit time (\u0394 Due to the continuous oil and gas exploitation, conventional hydrocarbon reserves are gradually depleted, and the production rates of the current reservoirs are significantly declining.Conventional hydrocarbon reserves are gradually depleting due to the continuous oil and gas exploitation and the production rates of the current reservoirs dramatically dropped , 2. SourTotal organic carbon (TOC), which has been widely considered as a quantification of the hydrocarbon generation potentials \u201310, is oSeveral authors developed empirical correlations to determine TOC, and these models were developed based on the experimental TOC measures on drilling cuttings or core samples and the corresponding well logs. Then, these developed correlations are proposed to be applied to determine the TOC for different wells \u201319. Thest), bulk density (RHOB), neutron porosity (CNP), gamma ray (GR), and spectrum logs of thorium (Th), potassium (K), and uranium (Ur).Artificial intelligence (AI) has been used in different industries . AI techTOC is a vital parameter to characterize the unconventional resources. Experimental analysis can be used to measure the TOC, but it is expensive, time-consuming, and does not give a continuous profile for the total organic carbon with depth. Empirical correlations can be used to estimate TOC. However, there are concerns about the generalization and accuracy of these correlations. In this paper, the application of different AI techniques in TOC prediction in Devonian shale formation from the well logs will be tested. These well logs include sonic transient time, resistivity, bulk gamma ray, bulk density, and spectral GR logs of Th, Ur, and K and neutron log porosity. The next sections in the paper describe the methodology that was used to build the ML models, followed by building the model. The models were then tested and validated with different data sets. A sensitivity analysis was conducted to investigate the importance of input logging parameters in the predicted TOC values.In this study, three machine learning tools were used to estimate the TOC as a function of eight well logs records. Experimental data for TOC from three different wells have been collected together with their corresponding well logs. 891, 291, and 82 data points from Well-A, Well-B, and Well-C were used to train, test, and validate the AI models all, respectively. All wells are in Devonian Duvernay shale, which is an organic liquid-rich source rock. The sedimentary basin is located in Alberta, Canada, with 61.7 billion barrels and 443 trillion cubic feet of oil and gas reserves, respectively [Rock-Eval 6 was employed to estimate the actual TOC values of drilling cuttings from different wells. Tests procedures are shown in Prior to the AI model's training, outliers, incomplete, or unrealistic data points were eliminated from the data used to construct the model. Data points that contain any value that is away from the mean of the data with three times the standard deviation were considered as an outlier. The statistical characteristics of Well-A's dataset are illustrated in The AI model was trained and optimized in this work using Well-A dataset that contains 891 data points with wide ranges of TOC and well logs values. The effect of various parameters inside the AI algorithms was tested to optimize the models, by running the AI tools inside a mutable for loops in MATLAB.In SVM models, different kernel functions, values for kernel options, epsilon, and regularization were tested while in RF, different sets of number of trees, maximum number of level in each tree, and maximum number of features were used. In DT, various values for maximum tree depth, minimum sample split, and maximum number of features were tested. The correlation coefficient between the known and predicted TOC and the average absolute percentage error (AAPE) were used as evaluation criteria.In addition, different sets of inputs were used to evaluate the significance of each well log parameter in TOC prediction. Seven sets were considered, the least one includes four parameters, and the most comprehensive consists of all eight parameters, as shown in The accuracies of the generated models were tested and validated using 291 and 82 data points from Well-B and Well-C, respectively. The two wells are in the same field as Well-A. The performance of AI models was also compared with that of currently existing correlations, such as the Schmoker model, the logR approach, and the Zhao et al. model.t, CNL, FR, GR, and spectral GR. The training dataset consisted of 891 data points from Well-A, while the testing dataset contains 291 data points from Well-C. This section presents the results obtained using each method.The AI models were trained for TOC estimation based on eight well log data of RHOB, \u0394Using data set from Well-A and Well-B, different trials have been applied using support SVM with changing some tuning parameters inside the algorithm, such as kernel function and regularization. The best results were achieved using the Gaussian kernel function. Good results have been achieved in the training dataset with a 7.1% average error; however, the accuracy in the testing datasets was relatively low with an average error of 19.7%. The correlation coefficients were 0.974 and 0.856 for training and testing, respectively. R value which equals 0.864 and which reduces the ability to generalize the model. In This technique resulted in perfect fitting in the training dataset with a 0.994 correlation coefficient as shown in R which equals 0.989. In comparison with DT, RF performed better in testing with a 0.931 correlation coefficient, and its performance with the training data set was very close with From the previous analysis, RF was chosen to perform the sensitivity on the inputs. Seven different sets of the well logs information are tested and reported in R, and Zhao et al. correlations.As shown in the previous results, good matching accuracies for Devonian shale's TOC have been achieved by the two methods in training and testing datasets. As an additional validation, eighty-two data points from Well-C were kept hidden from the AI tool during model construction utilized later to ensure the generalization of the new models. This well is located in the vicinity of the Well-I and Well-II that has been used in model building. The validation data points fall in 140\u2009ft depth range. The accuracy of the two AI techniques was compared with that of three of the available models for TOC estimation, namely, the Schmoker, \u0394logR of 0.94. DT and SVM models were less accurate than RF; however, in terms of AAPE, both were better than the other three correlations with AAPE values less than 16.4%. Zhao et al. [R predictions with AAPE values range between 20% and 25% and R values between 083 and 0.83 while the least favorable results in the validation dataset were from the Schmoker model with AAPE above 48%.The TOC values obtained from different empirical correlations and AI-based models against the actual values of Well-C dataset are presented in o et al. correlatt logs and level of maturity (LOM) (\u0394logR method) or bulk gamma ray, FR, and \u0394t or RHOB logs or a combination of FR and \u0394o et al. correlatt, FR, K, Th, and Ur logs, which helps to obtain a continuous profile for TOC with depth without the need for core analysis or additional well interventions. Similar to any developed model, we recommend employing the developed models using input parameters within the same model's inputs ranges to ensure reliable results. For future work, more data will be collected to validate the developed models and other ML techniques will be applied.The developed models were able to accurately predict the TOC from the convention well log including CNP, RHOB, GR, \u0394R values of 0.99 and 0.93, respectively, and AAPE values of 5.3% and 13.8% in the same order.In training and testing datasets, the three AI algorithms produced good matches; however, the RF-based model has the best accuracy. The RF model was able to predict the TOC for the training and testing datasets, with Data from a different well were hidden entirely from the AI tools and used to validate the built model. In this dataset, the RF model produced a 0.94 correlation coefficient and a 14% AAPE.The AI-based models' predictions were compared with three other empirical correlations. The AI models yielded more accurate results contrasted to the other models which resulted in AAPEs greater than 20%.This study established three models for TOC prediction in Devonian shale from conventional well logs and spectral GR logs using different machine learning techniques and approximately 1250 data points from three wells. The employed ML techniques were support vector machine (SVM), decision tree (DT), and random forest (RF). A summary of the findings reported in this paper is as follows :In train"} +{"text": "Tnfsf11 gene, which encodes RANKL, is complex and involves distant regulatory regions. Nevertheless, cell culture studies suggest that an enhancer region near the transcription start site is involved in the control of Tnfsf11 expression by hormones such as 1,25-(OH)2 vitamin D3 and parathyroid hormone, as well as the sympathetic nervous system. To address the significance of this region in vivo, we deleted the sequence between -510 to -1413 bp, relative to Tnfsf11 exon 1, from mice using CRISPR-based gene editing. MicroCT analysis of the femur and fourth lumbar vertebra of enhancer knockout mice showed no differences in bone mass compared to wild type littermates at 5 weeks and 6 months of age, suggesting no changes in osteoclast formation. RNA extracted from the tibia, fifth lumbar vertebra, thymus, and spleen at 6 months of age also showed no reduction in Tnfsf11 mRNA abundance between these groups. However, maximal stimulation of Tnfsf11 mRNA abundance in cultured stromal cells by PTH was reduced approximately 40% by enhancer deletion, while stimulation by 1,25-(OH)2 vitamin D3 was unaffected. The abundance of B and T lymphocytes in the bone marrow did not differ between genotypes. These results demonstrate that the region between -510 and -1413 does not contribute to Tnfsf11 expression, osteoclast support, or lymphocyte production in mice under normal physiological conditions but may be involved in situations of elevated parathyroid hormone.The cytokine RANKL is essential for osteoclast formation during physiological and pathological bone resorption. RANKL also contributes to lymphocyte production, development of lymph nodes and mammary glands, as well as other biological activities. Transcriptional control of the Tnfsf11 gene, which encodes RANKL, is expressed by a broad range of cell types in response to a variety of different conditions and signals stimulate osteoclast formation in part via stimulation of Tnfsf11 transcription [Tnfsf11 to PTH and 1,25(OH)2D3 in cells of the osteoblast lineage. Specifically, we used reporter constructs derived from bacterial artificial chromosomes (BACs), as well as chromatin immunoprecipitation coupled with next generation sequencing (ChIP-seq), to identify an enhancer located 75\u201376 kb upstream from the transcription start site (TSS) that mediates the response to both PTH and 1,25(OH)2D3 [Tnfsf11 expression, decreases osteoclast number, and increases bone mass under normal physiological conditions [Tnfsf11 expression by PTH, and deletion of this region from the mouse genome also decreases Tnfsf11 expression and bone resorption, and increases bone mass [Hormones such as parathyroid hormone (PTH) and 1,25-dihydroxyvitamin Dcription . In prev5(OH)2D3 . Importanditions . We alsoone mass .Tnfsf11 transcription is complex and involves multiple distant enhancers. Nonetheless, studies by others have implicated regions more proximal to the transcriptional start site (TSS) in the control of Tnfsf11 expression by PTH and 1,25(OH)2D3. For example, deletion of a sequence resembling a vitamin D response element (VDRE) from a position 922 bp upstream from the TSS blunts 1,25(OH)2D3-stimulation of a promoter-reporter construct in an osteoblastic cell line [Tnfsf11 expression by the sympathetic nervous system [Tnfsf11 expression by isoproterenol, a surrogate for activation of sympathetic nervous system signaling, also required the sequences at -922 and -788 [Together, these studies demonstrate that regulation of s system . More res system . Anotherand -788 . Howeverand -788 .Tnfsf11 TSS play a role in expression of this gene and thereby bone metabolism. We used CRISPR-mediated gene editing to delete the region between -510 to -1413 bp, relative to the TSS, from the murine genome. We found that mice lacking this region exhibited normal skeletal mass and architecture and that Tnfsf11 mRNA abundance in tissues was unchanged, albeit stimulation by PTH was moderately reduced in primary cell cultures.The goal of the current study was to determine whether the potential regulatory sequences located near the murine Tnfsf11 TSS were identified using the GPP sgRNA Designer tool [GTTTAGCCTCAGTTATCCCTCCAA-3\u2019; eKO Reverse 5\u2019-CCACGATCTCAAAGACAGGTCA-3\u2019; product size 261 bp for mice lacking the region between the cut sites. The sequence of the deleted allele was confirmed by TOPO TA-cloning of the 261 bp product and sequencing . For genotyping experimental mice, a third primer located within the deletion region was introduced to the PCR for identifying wild type alleles: WT Reverse 5\u2019-CCCACCCCATTCTTTTCCTA-3\u2019; product size 185 bp. Breeding of founder mice and all subsequent offspring was performed with C57BL/6J mice. All mice were fed lab diet 8640 (Envigo), provided water ad libitum, and were maintained on a half day (12 hour) light/dark cycle. The institutional Animal Care and Use Committee of the University of Arkansas for Medical Sciences approved all protocols involving eKO mice and their wild type (WT) littermates.Single guide RNAs (sgRNAs) targeting regions -510 and -1413 bp upstream of the ner tool . The sgRner tool . The sg-BMD was measured using the PIXImus densitometer . Software version 2.0 was used for acquisition and analysis of the images. Three different sites were measured. The femur measurement encompassed the right femur. The spine measurement was a rectangle that enclosed the 8 largest vertebrae (last 2 thoracic and 6 lumbar vertebrae). The total body measurement encompassed the whole body minus the calvarium, teeth, mandible, and majority of the tail . Isoflurane sedation was used to keep the animals motionless during the 4 minute scan. Animal respiration, heart rate, and the righting reflex were monitored during and after sedation.E\u2009 = \u200955\u2009kVp, I\u2009 = \u2009145\u2009\u03bcA, integration time\u2009 = \u2009200\u2009ms). Femurs were scanned beginning immediately distal to the third trochanter to the beginning of the distal growth plate. Cortical dimensions were determined at the diaphysis. Trabecular bone measurements at the distal femur were made on 151 slices beginning 8\u201310 slices away from the growth plate in order to avoid the primary spongiosa, and proceeded proximally. The fourth lumbar vertebra (L4) was scanned from the rostral growth plate to the caudal growth plate. Trabecular bone analyses were performed on contours of cross-sectional images, drawn to exclude cortical bone, as described for femoral trabecular bone. All trabecular measurements were made by drawing contours every 10\u201320 slices and using voxel counting for bone volume per tissue volume and sphere-filling distance transformation indices. Calibration and quality control were performed weekly using five density standards, and spatial resolution was verified monthly using a tungsten wire rod. Beam hardening correction was based on the calibration records. All nomenclature, symbols, and units adhered to guidelines in the literature [MicroCT scanning was used to measure cortical and trabecular architecture of the femur and the trabecular architecture of the fourth lumbar vertebra (L4). Bones were dissected, cleaned of soft tissues, fixed in 10% Millonig\u2019s formalin overnight, and transferred gradually from 70 to 100% ethanol. Dehydrated bones were scanned using a model \u03bcCT40 scanner to generate three-dimensional voxel images (1024\u2009\u00d7\u20091024 pixels) of bone samples. A Gaussian filter was used to reduce signal noise and a threshold of 220 was applied to all scans, at medium resolution (terature .6 cells per well of a 12 well plate (9 wells per genotype per time point). Cells were cultured until reaching 70\u201380% confluency. Three wells of each genotype were treated with vehicle (0.03% ethanol), 10 x 10\u22127 M bovine PTH (1\u201334), or 1x10-8 M 1,25(OH)2D3 for 1, 4, 12, or 24 hours. The experiment was performed twice; once using male mice and once using female mice.Left femurs and tibias were removed from 3 eKO or 3 WT mice under sterile conditions. Soft tissues were removed using sterile gauze and the distal and proximal epiphyses were cut off the shafts using a scalpel blade. Alpha Minimum Essential Medium (ThermoFisher) supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin-glutamine (ThermoFisher) was used to flush bone marrow stromal cells into a 12-well plate with marrow from similar genotyped mice pooled together. The cells were then reseeded at a density of 5x10ex vivo bone marrow cultures according to the manufacturer\u2019s instructions. RNA quantity and 260/280 ratio were determined using a Nanodrop instrument (ThermoFisher), and RNA integrity was verified by resolution on 0.8% agarose gels. Five hundred nanograms of RNA was used to synthesize cDNA using the High-Capacity cDNA Reverse Transcription Kit (ThermoFisher) according to the manufacturer\u2019s directions. Transcript abundance in the cDNA was measured by quantitative PCR using TaqMan Universal PCR Master Mix (ThermoFisher) and Taqman assays. PCR amplification and detection were carried out on an ABI StepOnePlus Real-Time PCR system (ThermoFisher) as follows: 2-min holding stage at 50C followed by a 10-min initial denaturation at 95C, 40 cycles of amplification including denaturation at 95C for 15 sec and annealing/extension at 60C for 1 min. The following TaqMan assays from Life Technologies were used: RANKL (Mm00441908_m1), OPG (Mm00435452_m1), and the housekeeping gene ribosomal protein S2 (Mm00475529_m1). Gene expression was calculated using the comparative threshold cycle (\u0394Ct) method [Total RNA was isolated from tissues by homogenization in Trizol Reagent (ThermoFisher) according to the manufacturer\u2019s instructions. The following tissues were used for RNA isolation: cortical bone from the tibia after flushing out the bone marrow and scraping the periosteum with a scalpel, whole L5 vertebrae, thymus, and spleen. The RNeasy Plus Mini kit was used to extract mRNA from KL Mm004408_m1, OPBone marrow cells were collected from left femurs and tibias of 5 female eKO and 5 female WT mice by removing both epiphyses and flushing out cells with PBS containing 3% FBS. The cells were then washed and incubated with anti-mouse CD16/CD32 for 5 minutes on ice. Then the samples were stained for 30 minutes on ice with the following antibodies: 5 \u03bcg/ml anti-CD3-FITC for T cells, 2 \u03bcg/ml anti-CD19-APC-Cy7 for B cells, 0.5 \u03bcg/ml anti-CD11b/Mac1-APC for monocyte progenitors, and 0.5 \u03bcg/ml anti-Ter119-PerCP/Cy5.5 for erythroid cells. Cells were washed to remove unbound antibodies and then fixed with 4% paraformaldehyde in phosphate-buffered saline for 20 minutes before analysis using a BD FACS Aria flow cytometer (BD Biosciences). Data analysis was conducted using FlowJo Software . Fluorescence Minus One (FMO) controls (BD Biosciences) guidance was followed for appropriate gating for the different cell populations.g for 20 minutes to separate serum from cells. Soluble RANKL in blood serum was measured using a mouse Quantikine kit according to the manual provided by the manufacturer. A five-parameter regression formula was used to calculate the sample concentrations from standard curves using GraphPad statistical software.Blood was collected by retro-orbital bleeding into a microcentrifuge tube and allowed to clot at room temperature for 2 hours. It was then centrifuged at 2000 \u00d7 Two-way ANOVA, Student\u2019s t test, or Repeated Measures Mixed Effects models for measurements over time were used to detect statistically significant genotype or treatment effects, after determining that the data were normally distributed and exhibited equivalent variances. In some cases, Wilcoxon rank-sum tests were used in place of Student\u2019s t-test and log, square root, or rank transformations were used to obtain normally distributed data. Pairwise comparisons and contrasts of genotypes were estimated in the ANOVA models. All t tests were two-sided. Tukey or Benjamini-Hochberg corrections were used for multiple comparison adjustments for each family of tests of pairwise comparisons and contrasts. Statistical analyses were performed using GraphPad Prism 8 or SAS version 9.4.Tnfsf11 TSS [Fig 1A). Moreover, our previous in vitro studies demonstrated that promoter-reporter constructs containing up to 2 kb of sequence upstream from the TSS were unresponsive to either PTH or 1,25(OH)2D3 [Tnfsf11 transcription. To directly address this question in vivo and in the context of the endogenous Tnfsf11 gene, we deleted a DNA fragment from -510 to -1413 bp, relative to the TSS of the Tnfsf11 gene, from the mouse genome . This region contains the putative VDRE, CRE, and NFATc1 binding sites identified by others [Figs S1).We have previously identified functional enhancers located 75\u201376 kb and 22 kb upstream from the murine sf11 TSS ,10. ConsFig 2A and 2B). While there was no difference in body weight in female mice, eKO males weighed slightly less than controls at 16 and 25 weeks . Consistent with the latter, male eKO mice exhibited slightly lower lean and fat mass but no change in femur length .We selected one founder mouse lacking the region between -510 and -1413, indicated by the arrows in Fig 3A\u20133D). Similarly, cancellous bone volume in the spine also did not differ between genotypes . Taken together, the BMD and micro-CT results demonstrate that deletion of the -510 and -1413 region did not alter bone mass or architecture in the cancellous or cortical bone compartments during growth or adulthood. These results also suggest that osteoclast formation and bone resorption were unaffected by the deletion.To determine if eKO mice exhibited differences in skeletal architecture and bone volume, we performed micro-computed tomography (micro-CT) analysis on right femurs and fourth lumbar vertebrae (L4) of mice of both sexes at 5 weeks and 6 months of age. Femoral cancellous bone volume did not differ between genotypes at either age, nor did femoral cortical thickness , tibial cortical bone, spleen, and thymus of WT and eKO mice at 6 months of age by quantitative PCR. We did not observe differences in Tnfsf11 mRNA levels between genotypes in any of these tissues , nor did we detect differences in circulating RANKL protein . Consistent with no change in Tnfsf11 mRNA in the spleen and thymus, the percentage of B and T lymphocytes in the bone marrow, as measured by CD19-positive and CD3-positive cells, respectively, was not altered by deletion of the putative enhancer .Since micro-CT analysis did not reveal any differences between WT and eKO mice at either age in either sex, we limited further analysis to female mice. While there were no differences in bone mass, deletion of the enhancer could still have a small impact on expression of Tnfsf11 expression, we examined the responsiveness of this gene to PTH or 1,25(OH)2D3 in bone marrow stromal cells isolated from eKO and WT littermates. PTH potently stimulated Tnfsf11 mRNA abundance in cells from both genotypes but the maximal stimulation was reduced by approximately 40% in cultures from the eKO mice . This finding was replicated in a second experiment performed in cells from mice of the opposite sex . In contrast, the potent stimulation by 1,25(OH)2D3 was not consistently changed by deletion of the enhancer region . To confirm that the bone marrow cultures from each genotype had reached similar stages of development, we measured expression of Tnfrsf11b and its response to the hormones. Suppression of Tnfrsf11b by either hormone was unaffected by deletion of the enhancer region .Lastly, as a stringent test for a possible role for the -510 and -1413 region in Tnfsf11 abundance leads to increased cancellous and cortical bone mass [Tnfsf11 mRNA in bones of mice lacking the -510 to -1413 region. Therefore, our results do not support a role for the -510 to -1413 region, and the putative transcription factor binding sites therein, in the control of Tnfsf11 expression in the bones of growing or adult mice under normal physiological conditions.Osteoclasts are essential during bone growth for removal of calcified cartilage beneath the growth plate, for modeling of bone shape, and for remodeling of bone matrix. Consequently, reductions in osteoclast formation or activity necessarily result in altered bone mass and structure. Specifically, reduced osteoclast number due to reduced one mass ,21. HoweTnfsf11 mRNA in thymus and spleen and again observed no changes after deletion of the potential enhancer. Consistent with this, we did not observe any changes in B or T lymphocyte abundance in the bone marrow. Although the -510 to -1413 region does not contribute to Tnfsf11 expression in bone or lymphocytes, it remains possible that this region is involved in Tnfsf11 expression in cell types important for M cell, lymph gland, or mammary tissue development, which were not examined in this study. It is also possible that this region contributes to Tnfsf11 expression in bone under conditions that were also not examined here. For example, the finding that the response to PTH in vitro was blunted suggests that this region may be involved in the elevated Tnfsf11 expression associated with states of PTH excess. However, it is important to note that PTH is a tonic stimulator of Tnfsf11 under normal physiological conditions [Tnfsf11 mRNA in bones of the eKO mice suggests that the enhancer is not required for response to physiologically normal levels of PTH, PTHrP, or 1,25(OH)2D3.As RANKL has functions in non-skeletal tissues ,4,22, wenditions . Thus, tTnfsf11 enhancer mouse model, with only slightly less genetic heterogeneity, clearly revealed differences in bone mass and gene expression in mice lacking a more distal enhancer [Tnfsf11 expression, the contribution is small. The genetic heterogeneity may also have contributed to the slight difference in body weight observed at some ages in the male eKO mice.One limitation of our study was that the mice were not in a homogeneous genetic background. The majority of the animals used in our study were in a mixed C57BL/6-BalbC background with a slightly higher contribution from C57BL/6 than BalbC. Nonetheless, our previous study of a different enhancer . This, cTnfsf11 by 1,25(OH)2D3, previous studies identified a putative VDRE within this region [Although we found no evidence that the -510 to -1413 region contributes to the control of s region . It is is region . Thus, rS1 FigA) PCR products from the genotyping PCR described in the Material and methods section were fractionated on an agarose gel, stained with ethidium bromide, and imaged. The size of the eKO product is 261 bp and that of the WT product is 185 bp. Representative results are shown for a homozygous eKO mouse, heterozygous eKO mice, and a WT mouse. (B) The sequence of the eKO allele was determined by TA-cloning of the 261 bp PCR product and sequencing. The relevant portion of the sequencing result is shown aligned to the sequence of the WT allele. 855 bp of the WT allele were ommitted from the figure to reduce the size of the sequence. The PAMs for each of the sgRNAs used to create the deletion are highlighted in green.((TIF)Click here for additional data file.S2 FigA) and female (B) wild type and homozygous eKO mice. n = 11 to 18 per group, p values determined using 2-way ANOVA. (C-D) Femur length of 5-week-old and 6-month-old mice using right femurs measured with calipers. n = 3 to 18 per group, p values determined using 2-way ANOVA. Values are means \u00b1 s.d..Serial analysis of lean and fat mass using Piximus-derived fat percentage and total tissue mass data beginning at 8 weeks of age until 25 weeks of age in male ((TIF)Click here for additional data file.S3 FigA-D) Tnfsf11 (RANKL) and Tnfrsf11b (OPG) mRNA measured by Taqman RT-PCR in RNA prepared from bone marrow stromal cell cultures from female mice of the indicated genotype treated with vehicle, 10\u22127 M PTH, or 10-8M 1,25(OH)2D3 for the indicated times. Values are the mean fold-change, \u00b1 s.d., of triplicate wells; p values determined by comparing eKO versus WT values at the same time point by Repeated Measures Mixed Effects models.((TIF)Click here for additional data file.S1 Raw images(TIF)Click here for additional data file. 22 Feb 2021PONE-D-21-02120Deletion of a putative promoter-proximal Tnfsf11 regulatory region in mice does not alter bone mass or Tnfsf11 expression in vivoPLOS ONEDear Dr. O'Brien,Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE\u2019s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.The reviewers noted a few important gaps in your MS. \u00a0Most important is to document success of the targeting strategy. \u00a0In addition, more details about growth and body composition are sought. \u00a0While thew overall decision is classified as \"major revision\" because of their importance to the story, making the requested changes should be relatively straightforward.plosone@plos.org. When you're ready to submit your revision, log on to https://www.editorialmanager.com/pone/ and select the 'Submissions Needing Revision' folder to locate your manuscript file.Please submit your revised manuscript by Mar 29 2021 11:59PM. 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The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data\u2014e.g. participant privacy or use of data from a third party\u2014those must be specified.The Reviewer #1:\u00a0YesReviewer #2:\u00a0Yes**********4. Is the manuscript presented in an intelligible fashion and written in standard English?PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.Reviewer #1:\u00a0YesReviewer #2:\u00a0Yes**********5. Review Comments to the AuthorPlease use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. Reviewer #1:\u00a0This work from MacLeod et al indicates that a region upstream of the RANKL (Tnfsf11) gene does not play an essential role in regulating bone structure, although it may be partially required for normal response of bone marrow-derived stromal cells to PTH stimulation of that gene. The data presented is clear, and limitations are clearly out lined in the discussion, but the rationale of the study and the success of the targeting strategy are not clear.Major concerns:1. No data is presented to show that the targeting strategy was effective. This is a major shortcoming as it calls all conclusions into question. Please show that recombination was successful.2. Figure 1 is not very clear and the statement of hypothesis is a bit muddy in some instances. For those who are interested in RANKL regulation, but are not conversant with all the regions, it becomes difficult to follow. It is really important to show in Figure 1 how the targeted region aligns with the previously described enhancers at 22kb, and 75-76kb. You that you are targeting a region lacking \"hormonal responsiveness\" (lines 207-211), but this is also the region that contains the CRE and VDRE? Please clarify.3. RANKL is also highly responsive to IL-6, and this group has published much on that issue. Where is the putative IL-6 response site (STAT binding element) compared to the region deleted?4. The in vitro studies showing downregulation of the PTH effect on RANKL is based on a single cell preparation. This is not adequate and should either be removed or repeated so it is clear that the result was reproducible.Minor concerns:1. Methods: please provide more detail on the regions measured by micro-CT.2. Please indicate in all figure legends whether error bars are SD or SEM.3. The title for the legend of Figure 4 is a bit misleading as this also contains in vitro data.4. Since PTHrP is required for normal osteoclast formation and acts through the same receptor as PTH, you should include PTHrP in your statement on line 316 that the enhancer is not required for physiological action.5. For full data transparency, you should show individual data points for Figure 2, and Figure 4C-G.Reviewer #2:\u00a0An enhancer near to the transcriptional start site has been reported for the Tnfsf11 gene, that codes for RANKL. In this study, the authors generate a transgenic mouse where in this enhancer has been eliminated completely. They evaluate the impact of the loss of this enhancer in vivo and in vitro and conclude that outside of high PTH conditions, this enhancer does not appear to play a role in bone. This is a beautifully written and clear paper. The experiments conducted are appropriate to address the role of this enhancer in Tnfsf11 and the results are presented in a clear fashion. I do have some recommendations and a couple of questions on issues that were not addressed in this paper.1. Figure 2. Please provide body length to help interpret the body weigh issue. Are the knockout mice leaner, lacking mean mass or just over all smaller? The data on lean and fat mass should already be available from the DXA files.2. Figure 3. Related to the comment above, please provide femur lengths. Given the nearly significant result for cortical thickness for the 5 week old females, it would be ideal to present cortical area and marrow area as well. Please place the P values for the within gender comparisons for this figure for transparency purposes.3. Figure 4E. Arguably you do have a response to Vit D3. It is hard to tell what error bar belongs to what point. Please provide the P value for the 24 hour time point. The statements in the discussion regarding Vit D and this enhancer need to be tempered and not be such an absolute. With three samples for this assay, what is your power of detection for a difference here? The error bars are certainly wider for the KO than for the WT.4. The issue of body weight is not mentioned in the discussion. Do you have any explanation for the reduction in body weight? As this mouse is not completely without a phenotype, the discussion needs to be expanded to reflect this.**********what does this mean?). If published, this will include your full peer review and any attached files.6. PLOS authors have the option to publish the peer review history of their article digital diagnostic tool,\u00a0 1 Apr 2021Response to ReviewersReviewer 1.Major concerns:1. No data is presented to show that the targeting strategy was effective. This is a major shortcoming as it calls all conclusions into question. Please show that recombination was successful.We regret that we did not more carefully explain how we confirmed the sequence of the deleted allele. We now state clearly that a PCR product containing the deletion junction was cloned and sequenced. An example of the PCR and an alignment of the sequence of the deleted allele with the WT allele is presented in new S1 Fig. 2. Figure 1 is not very clear and the statement of hypothesis is a bit muddy in some instances. For those who are interested in RANKL regulation, but are not conversant with all the regions, it becomes difficult to follow. It is really important to show in Figure 1 how the targeted region aligns with the previously described enhancers at 22kb, and 75-76kb. You that you are targeting a region lacking \"hormonal responsiveness\" (lines 207-211), but this is also the region that contains the CRE and VDRE? Please clarify.Our previous in vitro studies showed that the first 2000 bp of DNA upstream from the TSS were unable to confer responsiveness to either PTH or 1,25(OH)2D3 upon stable transfection of promoter-reporter constructs. In contrast, others have shown that similar constructs do display responsiveness to PTH and 1,25(OH)2D3, as well as isoproterenol. Thus, our previous studies suggest that this region is not important for Tnfsf11 responsiveness to these hormones while those of other groups suggest that it is important.In an attempt to resolve these conflicting results, we deleted a fragment containing the putative response elements identified by the other groups from the endogenous murine Tnfsf11 gene and examined the impact on bone mass and Tnfsf11 gene expression. We have edited the text of the Results section to include more of the information that we presented in the Introduction section in an effort to clarify the goal of the present study.Due to the large distance of the 22 kb and 75-76 kb enhancers from the TSS, their inclusion in the alignment in Fig 1 is not practical. Since no aspect of the current study deals with these distant enhancers, it may in fact be confusing to include them in the diagram in Fig 1, which deals exclusively with the proximal promoter region that we study in the current work.3. RANKL is also highly responsive to IL-6, and this group has published much on that issue. Where is the putative IL-6 response site (STAT binding element) compared to the region deleted?IL-6 is a member of the gp130 family of cytokines and we have shown that the enhancer mediating the bulk of the response to these cytokines is located 88 kb upstream from the TSS . However, there are no conflicting results regarding the location of the response elements to gp130 family cytokines. Therefore, we think that discussion of their location in the present work will likely be distracting.4. The in vitro studies showing downregulation of the PTH effect on RANKL is based on a single cell preparation. This is not adequate and should either be removed or repeated so it is clear that the result was reproducible.We thank the reviewer for making this point and have included a second experiment performed using cells from female mice. The first experiment used cells from male mice. The results of both experiments are very similar, showing a mild reduction in the response to PTH at 4 hr of treatment. The second experiment showed a mild increase in the response to 1,25(OH)2D3 in cells from the eKO mice at the 24 hr time-point. Since this did not occur in the first experiment, this latter result likely represents experimental variation. We have included the new results in S3 Fig.Minor concerns:1. Methods: please provide more detail on the regions measured by micro-CT.We have added details for both the femur and vertebral measurements.2. Please indicate in all figure legends whether error bars are SD or SEM.We apologize for this omission and this information is now included in the figure legends.3. The title for the legend of Figure 4 is a bit misleading as this also contains in vitro data.We agree and have altered the title for this figure legend.4. Since PTHrP is required for normal osteoclast formation and acts through the same receptor as PTH, you should include PTHrP in your statement on line 316 that the enhancer is not required for physiological action.We have included PTHrP in this conclusion.5. For full data transparency, you should show individual data points for Figure 2, and Figure 4C-G.We have included individual data points for Fig 4C. However, when we did so for the line graphs in Fig 2 and Fig 4, the points representing the means were obscured and the graphs become essentially unintelligible. Thus, for these later graphs, we think it is clearer not to include individual data points.Reviewer #21. Figure 2. Please provide body length to help interpret the body weigh issue. Are the knockout mice leaner, lacking mean mass or just over all smaller? The data on lean and fat mass should already be available from the DXA files.Unfortunately we did not collect body length measurements on these mice. Because the mice are not uniformly extended in the DXA scans, we could not use the scans for such measurements. As suggested by the reviewer, we have included the lean and fat mass measurements from the DXA scans in a new figure, S2 Fig. Overall, the male eKO mice are slightly leaner and have slightly less fat. 2. Figure 3. Related to the comment above, please provide femur lengths. Given the nearly significant result for cortical thickness for the 5 week old females, it would be ideal to present cortical area and marrow area as well. Please place the P values for the within gender comparisons for this figure for transparency purposes.The femur lengths did not differ by genotype and are now included in the new S2 Fig. As suggested, we now include the within gender P values in Fig 3.3. Figure 4E. Arguably you do have a response to Vit D3. It is hard to tell what error bar belongs to what point. Please provide the P value for the 24 hour time point. The statements in the discussion regarding Vit D and this enhancer need to be tempered and not be such an absolute. With three samples for this assay, what is your power of detection for a difference here? The error bars are certainly wider for the KO than for the WT.We now include the P value (p = 0.163) for the 24 hr time-point. Based on the lack of significance here, and our finding that in a second experiment, the response to 1,25(OH)2D3 was increased in cells from the eKO mice at 24 hr, we conclude that there is not a consistent change in the response to this hormone. 4. The issue of body weight is not mentioned in the discussion. Do you have any explanation for the reduction in body weight? As this mouse is not completely without a phenotype, the discussion needs to be expanded to reflect this.We think the mostly likely explanation for the slightly lower body weight of the male eKO mice is genetic heterogeneity. The female eKO mice appear to be slightly heavier than controls, but this does not reach statistical significance. The overall effect size is similar in both sexes, just in the opposite direction. Based on this, and the fact that there is no evidence in any previous Tnfsf11 loss-of-function study that this cytokine plays a role in the control of body weight, we think that the mostly likely explanation for these effects is that our animals are not in a pure genetic background. We have now mentioned this possibility in the Discussion section. 19 Apr 2021Deletion of a putative promoter-proximal Tnfsf11 regulatory region in mice does not alter bone mass or Tnfsf11 expression in vivoPONE-D-21-02120R1Dear Dr. O'Brien,We\u2019re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.Within one week, you\u2019ll receive an e-mail detailing the required amendments. When these have been addressed, you\u2019ll receive a formal acceptance letter and your manuscript will be scheduled for publication.http://www.editorialmanager.com/pone/, click the 'Update My Information' link at the top of the page, and double check that your user information is up-to-date. If you have any billing related questions, please contact our Author Billing department directly at authorbilling@plos.org.An invoice for payment will follow shortly after the formal acceptance. To ensure an efficient process, please log into Editorial Manager at onepress@plos.org.If your institution or institutions have a press office, please notify them about your upcoming paper to help maximize its impact. If they\u2019ll be preparing press materials, please inform our press team as soon as possible -- no later than 48 hours after receiving the formal acceptance. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information, please contact Kind regards,Robert Daniel Blank, MD, PhDAcademic EditorPLOS ONEAdditional Editor Comments :Reviewers' comments:Reviewer's Responses to QuestionsComments to the Author1. If the authors have adequately addressed your comments raised in a previous round of review and you feel that this manuscript is now acceptable for publication, you may indicate that here to bypass the \u201cComments to the Author\u201d section, enter your conflict of interest statement in the \u201cConfidential to Editor\u201d section, and submit your \"Accept\" recommendation.Reviewer #1:\u00a0All comments have been addressedReviewer #2:\u00a0All comments have been addressed**********2. Is the manuscript technically sound, and do the data support the conclusions?The manuscript must describe a technically sound piece of scientific research with data that supports the conclusions. Experiments must have been conducted rigorously, with appropriate controls, replication, and sample sizes. The conclusions must be drawn appropriately based on the data presented. Reviewer #1:\u00a0YesReviewer #2:\u00a0Yes**********3. Has the statistical analysis been performed appropriately and rigorously? Reviewer #1:\u00a0YesReviewer #2:\u00a0Yes**********4. Have the authors made all data underlying the findings in their manuscript fully available?PLOS Data policy requires authors to make all data underlying the findings described in their manuscript fully available without restriction, with rare exception (please refer to the Data Availability Statement in the manuscript PDF file). The data should be provided as part of the manuscript or its supporting information, or deposited to a public repository. For example, in addition to summary statistics, the data points behind means, medians and variance measures should be available. If there are restrictions on publicly sharing data\u2014e.g. participant privacy or use of data from a third party\u2014those must be specified.The Reviewer #1:\u00a0YesReviewer #2:\u00a0Yes**********5. Is the manuscript presented in an intelligible fashion and written in standard English?PLOS ONE does not copyedit accepted manuscripts, so the language in submitted articles must be clear, correct, and unambiguous. Any typographical or grammatical errors should be corrected at revision, so please note any specific errors here.Reviewer #1:\u00a0YesReviewer #2:\u00a0Yes**********6. Review Comments to the AuthorPlease use the space provided to explain your answers to the questions above. You may also include additional comments for the author, including concerns about dual publication, research ethics, or publication ethics. Reviewer #1:\u00a0(No Response)Reviewer #2:\u00a0I would like to thank the authors for their consideration of all comments made by both reviewers for this manuscript.**********what does this mean?). If published, this will include your full peer review and any attached files.7. PLOS authors have the option to publish the peer review history of their article (If you choose \u201cno\u201d, your identity will remain anonymous but your review may still be made public.Do you want your identity to be public for this peer review? For information about this choice, including consent withdrawal, please see our Privacy Policy.Reviewer #1:\u00a0NoReviewer #2:\u00a0No 30 Apr 2021PONE-D-21-02120R1 Tnfsf11 regulatory region in mice does not alter bone mass or Tnfsf11 expression in vivo Deletion of a putative promoter-proximal Dear Dr. O'Brien:I'm pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department. onepress@plos.org.If your institution or institutions have a press office, please let them know about your upcoming paper now to help maximize its impact. If they'll be preparing press materials, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact plosone@plos.org. If we can help with anything else, please email us at Thank you for submitting your work to PLOS ONE and supporting open access. Kind regards, PLOS ONE Editorial Office Staffon behalf ofProfessor Robert Daniel Blank Academic EditorPLOS ONE"} +{"text": "Home health monitoring has the potential to improve outpatient management of chronic cardiopulmonary diseases such as heart failure. However, it is often limited by the need for adherence to self-measurement, charging and self-application of wearables, or usage of apps. Here, we describe a non-contact, adherence-independent sensor, that when placed beneath the legs of a patient\u2019s home bed, longitudinally monitors total body weight, detailed respiratory signals, and ballistocardiograms for months, without requiring any active patient participation. Accompanying algorithms separate weight and respiratory signals when the bed is shared by a partner or a pet. Validation studies demonstrate quantitative equivalence to commercial sensors during overnight sleep studies. The feasibility of detecting obstructive and central apneas, cardiopulmonary coupling, and the hemodynamic consequences of non-sustained ventricular tachycardia is also established. Real-world durability is demonstrated by 3\u00a0months of in-home monitoring in an example patient with heart failure and ischemic cardiomyopathy as he recovers from coronary artery bypass grafting surgery. BedScales is the first sensor to measure adherence-independent total body weight as well as longitudinal cardiopulmonary physiology. As such, it has the potential to create a multidimensional picture of chronic disease, learn signatures of impending hospitalization, and enable optimization of care in the home. Hospitalizations for HF are often preceded by fluid accumulation with congestion and associated shortness of breath. If identified early, HF exacerbations can often be managed in the outpatient setting with temporary intensification of diuretic therapy3. Unfortunately, early detection currently relies on patient self-recognition and self-reporting of symptoms, which is unreliable, particularly during the COVID-19 pandemic, when patients are reluctant to go to hospitals4.Heart failure (HF) is among the most challenging chronic conditions to manage. It affects more than 6 million patients in the US and costs more than $30B per year, largely due to the high burden of inpatient care required to manage recurrent exacerbations5. In clinical studies, lack of adherence to self-measurement combined with variability of HF decompensation trajectories has prevented identification of a one-size-fits-all threshold of weight change that can trigger interventions to improve outcomes compared with usual clinic-based care10. Wearables are similarly limited by the need for patient engagement to charge and utilize sensors and apps12. Implantable pulmonary artery pressure sensors overcome some of these limitations and demonstrated reductions in HF hospitalization rates in the CHAMPION trial; however, the technology requires an invasive procedure, is costly, and requires ongoing adherence to patient-initiated data collection and transmission15. Patients with implanted cardiac rhythm management devices can be monitored for worsening HF using multiparameter monitoring of cardiopulmonary metrics; however, many HF patients (particularly those with preserved ejection fraction) do not require implantable pacemakers and defibrillators16. Given the limitations of existing remote monitoring technologies, we developed a non-invasive, multiparameter measurement technology that passively collects total body weight and dynamic cardiopulmonary physiology in the home bed without requiring any active patient participation.Patients are advised to self-measure daily weights and vital signs and to report sudden changes, but success of these strategies is limited by the need for patient engagement because there are no adherence-independent weight measurement devices for the outpatient setting21. For example, a modified stand-upon weigh scale was shown to classify HF patients based on 30-s BCGs, but the short duration of monitoring and the requirement for patient-initiated self-measurement leave it vulnerable to poor patient adherence24. Respirations and BCGs are often measured by piezoelectric or electromechanical film sensors placed above or below mattresses or bed frames, or via bedside radiofrequency transmit-receivers29; however, because these sensors do not span the entire body and are primarily sensitive to high frequency dynamic signals, they are unable to measure total body weight30. Since it is not uncommon for patients to present to the emergency room with 20 lbs of fluid overload, and because that amount of fluid does not accumulate in 1\u20132\u00a0days but instead does so gradually over days to weeks, a non-contact adherence-independent weight sensor has potential to enable early intervention for fluid overload in HF.Passive home monitoring has been achieved by embedding sensors into a wide range of everyday objects, facilitating measurement of respiratory signals and ballistocardiograms from healthy individuals and patients with chronic diseasesWe developed an under-the-bed mechanical sensing platform that achieves adherence-independent noncontact longitudinal physiological monitoring of total body weight, high fidelity respiratory signals, and BCG-derived heart rates. We validated the technology by comparing BedScales to ground truth data from commercial bathroom scales, chest respirometers, nasal flow sensors, and electrocardiograms used in overnight clinical sleep studies. We also demonstrated the feasibility of detecting respiratory pathologies including tachypneas, central and obstructive sleep apneas, and periodic breathing; ventricular arrhythmias and their hemodynamic consequences. Finally, we demonstrate the real-world durability of the platform by showing data from an example patient with HF and ischemic cardiomyopathy during a 3-month recovery from coronary artery bypass grafting surgery. By eliminating the need for patient participation, BedScales has the potential to improve care of patients with difficult-to-manage chronic cardiopulmonary diseases such as heart failure.Roughly one third of life is spent asleep in bed. This is a unique setting in which cardiopulmonary physiology can be longitudinally measured without requiring correction for level of activity and\u00a0without obscuration by musculoskeletal movements. To leverage this ideal diagnostic window, we designed, manufactured, and validated a non-contact fully adherence-independent home monitoring system called BedScales. It consists of low-profile force sensors beneath each leg of a conventional home bed, recliner, or couch, which measure and transmit 80\u00a0Hz sampled data to a cloud computing environment via WiFi, where physiological parameters and signals are quantified including total body weight, detailed respiratory waveforms, ballistocardiograms, and musculoskeletal movements, all without requiring any conscious patient participation, management of devices, engagement with apps, or self-application of wearables Fig.\u00a0a. Each nDigitized and amplified data from the individual force sensors is automatically transferred via micro-USB to a wall-powered central communications module, which in turn transmits it to a HIPAA-affiliate Amazon Web Services environment via WiFi. We selected a hardwired connection between the sensors and communications box to avoid the need for Bluetooth troubleshooting and to provide an indefinite power source that does not require battery changes. Once stored in a time-series database in the cloud, the data can be synchronously or asynchronously processed to create custom analytics, visualizations, and dashboards for permission-dependent sharing with patients, healthcare providers, or family and friends. In summary, the fully automated and adherence-independent install-once-use-indefinitely platform assumes nothing about patients\u2019 technical literacy and does not require an accompanying smartphone or laptop computer. It is therefore accessible to patients who are socioeconomically disadvantaged, geographically distanced, or physically or cognitively impaired. Furthermore, because the devices are scalably manufactured, they are suitable for appropriately powered clinical studies.Commercial weigh scales require that patients remember to self-initiate daily standing weight measurements, which limits their utility to engaged patients who can safely and stably stand on a home floor scale. Hospital beds measure patient weights when they are in bed using non-contact sensors, but they do so only at a single time point, leaving them vulnerable to unmeasured errors when blankets, pillows, books, and devices are added between the time of zeroing and measuring weight. In contrast, BedScales measures the weight of the bed and its contents continuously across time, which allows separate quantification of persons and objects based on the times that they are added or removed. For example, one can see the separate addition of a glass, increasing amounts of water, and recurrent placement and removal of a smartphone Fig. . Weights2\u2009=\u20090.99, n\u2009=\u2009162, spanning 100\u2013800 lbs) Fig.\u00a0. Figure\u00a0Individuals often share the bed with a partner or pet Fig.\u00a0a; howeveWhen a patient is asleep in bed, episodic musculoskeletal movements are separated by comparatively long movement-free intervals during which low variance physiological signals such as respirations and ballistocardiograms can be measured. This provides opportunities to perform adherence-independent longitudinal quantification of respiratory rate and detection of episodic tachypneas, apneas, and periodic breathing. BedScales respiratory signals arise from the dynamic redistributions of load that accompany chest wall movement during inspiration and expiration. To convert signals from multiple sensors into a single patient respiratory signal, we first performed bandpass frequency-dependent filtering of the individual sensor signals (cutoffs at 0.167\u00a0Hz and 1.5\u00a0Hz). We then used principal component analysis within a sliding window to calculate eigenvalues that, when multiplied by individual sensor signals and algebraically summed, create a single respiratory source signal for peak finding Fig.\u00a0a,b. The 24. To validate the measurements, we installed BedScales beneath the legs of a hospital bed during overnight sleep studies and compared the resulting signal to those obtained from the standard commercial chest belt respirometer Fig.\u00a0d-f. Thes32. Bandpass filtering of BedScales signals (5\u00a0Hz and 50\u00a0Hz cutoffs) revealed characteristic BCG morphologies from the individual scales with defined waves that follow each QRS complex of the electrocardiogram (ECG)les Fig.\u00a0a,b. A siles Fig.\u00a0c,d. Thisles Fig.\u00a0d, as a hles Fig.\u00a0e, and asles Fig.\u00a0f.Figure 34. We identified one sleep study patient with a high burden of ventricular ectopy, including premature ventricular contractions, ventricular couplets, triplets, and non-sustained ventricular tachycardia (NSVT) indicating respirophasic variation and cardiopulmonary coupling Fig.\u00a0a,b33,34.VT) Fig.\u00a0c,d. PremVT) Fig.\u00a0c-e. Toge37. During the\u2009~\u20098-h sleep study, we longitudinally measured respiratory signals from the BedScales along with the commercial chest respiratory belt as well as the nasal pressure airflow monitor. After aligning data, we quantified regions with low variance indicating a lack of respiratory waveforms. Regions greater than 10\u00a0s were defined as apneas and their distribution is shown in Fig.\u00a038. Taken together, these data demonstrate the feasibility of performing high fidelity BedScales monitoring of normal and pathologic respiratory dynamics and their hemodynamic consequences without the need for obtrusive adherence-dependent sensors.We next examined an overnight sleep study patient with sleep disordered breathing who had a high burden of central and obstructive sleep apneas (CSA and OSA) Fig.\u00a0a. OSA isTo demonstrate the durability of BedScales during longitudinal monitoring in a real-world environment of a patient\u2019s home, we show an example case of a\u2009~\u200960\u00a0year-old man who presented to the hospital with volume overload and newly discovered severely depressed ejection fraction due to ischemic cardiomyopathy. He was diuresed to euvolemia and discharged home with a plan to return several days later for coronary artery bypass grafting (CABG). At the time of discharge, BedScales were installed under his home recliner where he slept each night. We performed monitoring for several days prior to scheduled surgery and for 3\u00a0months during recovery. During that time, we quantified, every 30\u00a0s, (i) his in-bed and out-of-bed status as a binary quantity Fig.\u00a0a-c, (ii)39. We quantified his RR for 7-days \u201cpre-surgery\u201d, 7-days immediately \u201cpost-surgery\u201d, and 7-days at the end of \u201crecovery\u201d protocols IRB # 171480 and IRB # 180160. All experimental protocols were approved by UCSD HRPP IRB. Informed consent was obtained from all subjects.Custom housing was designed using Solidworks . Tooling was machined and injection molded parts were manufactured by S. Pawlicki. 50\u00a0kg strain gauges were purchased from Sparkfun or a comparable vendor. Custom circuit boards were designed in CircuitMaker (Altium) and included Hx711 integrated circuit technology (Avia Seminconductor) with a gain of 64 and a sampling frequency of\u2009~\u200980\u00a0Hz. The circuit board design\u00a0was based on an Hx711 breakout board schematic,\u00a0a load cell combinator\u00a0schematic, and their corresponding Eagle files, all from Sparkfun .\u00a0The circuit boards communicated via microUSB to a Raspberry Pi which received electrical power from the building wall outlet, which in turn powered each sensor. The transducers and custom circuit were snap fit into the housing which was secured with screws. Rubber tops were die-cut from rubber sheets and the scales were securely fixed to plastic plates using double-sided adhesive pads. The Raspberry Pi communicated data using the subject\u2019s home WiFi and securely transmitted data to Amazon Web Services S3\u00a0buckets. All analyses were performed in Python and Matlab .Bed sensor weight validation was performed by using five healthy volunteers and two static weights in various permutations to span a large range of loads. Each person was measured on a commercial bathroom scale before lying on a bed with the BedScales sensor under each of its 4 legs. The 4 scales were calibrated together by fitting coefficients that minimized the variance when the same load was applied in different places. The final weights were calculated by subtracting the total load measured after and before each permutation of individuals and weights was placed on the bed (3 measurements were made of each permutation and these were averaged together). Two-point calibration that minimized the measurement error was then used to convert from arbitrary units (AU) to pounds (lbs). Longitudinal weight comparisons were made by installing BedScales under a home bed and comparing to self-measurement on two separate commercial floor scales at the beginning of each night of sleep. The limits of sensitivity were tested by placing the sensors beneath a 4-leg couch and placing an empty glass on a flat cutting board. At 20\u00a0s intervals, 15\u00a0mL (0.033lbs) aliquots of water were added. The glass was removed and replaced with and without water, and a smartphone was repeatedly added and removed.BedScales respiratory signals were generated by frequency-dependent filtering with cutoffs of 0.167\u00a0Hz and 1.5\u00a0Hz. A single respiratory signal was derived by linearly combining the individual sensor respiratory signals weighted by PCA eigenvalues calculated for each 12.5\u00a0s window. A moving variance algorithm was used to isolate regions of steady physiology (regions\u2009<\u200910\u00a0s were rejected) and peak finding was performed on these regions. The respiratory rate was calculated using the median inter-peak interval during a 5-min moving window with a shift of 30\u00a0s. For validation, the BedScales respiratory signal and chest belt were subject to additional smoothing , windows were required to have no more than 45\u00a0s of unstable physiology, and regions with technical artifacts in the chest belt were excluded. Data was aligned and compared to a simultaneously recorded respiratory chest belt with respiratory rates quantified using the same method.BCG signals from each scale were derived by frequency-dependent filtering (Butterworth) with cutoffs of 5\u00a0Hz and 50\u00a0Hz\u00a0. These signals were then smoothed using a moving mean filter and moving variance filters. The resultant signals from each scale were then summed to create a composite signal, which was filtered using another frequency-dependent filter (Butterworth) with cutoffs of 1\u00a0Hz and 50\u00a0Hz. The resultant signal was a single peak measure of BCG. For in-home data, steady regions as defined by the respiratory signal were isolated and analyzed. For validation,\u00a0a moving variance algorithm was used to isolate regions of steady physiology (regions\u2009<\u20095\u00a0s were rejected) and peak finding was performed on these regions. The heart rate was calculated using the median inter-peak interval during a 5-min moving window with a shift of 30\u00a0s. For validation, windows were required to have no more than 45\u00a0s of unstable physiology and a region with technical artifacts in the ECG was excluded. Data was aligned and compared to a simultaneously recorded ECG signal\u00a0with heart\u00a0rates quantified using the same method.The weights of persons sharing a bed were determined by measuring the calibrated sum of all sensors across time and extracting the large differential weight changes. The weight changes were then classified into two groups termed person 1 and person 2. Simultaneity tests were performed by instructing 2 persons to get into and out of bed at specified temporal intervals in the following sequence\u2014Person 1 (IN), Person 2 (IN), Person 1 (OUT), Person 2 (OUT) and then repeated but exchanging Person 1 and 2. To explore the limits of simultaneity that would still permit decoupling of person weights, we systematically decreased the interval between Person 1 then 2 (or 2 then 1) getting into and out of bed and repeated the experiment for several time intervals , until the maneuver could not reach a steady position in the allotted interval.41. Given this estimate, the Kalman smoothing algorithm was used to extract the mechanical respiratory patterns of the two sources42. Validation was performed by simultaneously but independently measuring each respiratory signal using two respiratory belts . Interbreath intervals were compared by measuring the error between each demixed signal and each ground truth respiratory belt signal. For each individual, the absolute error between the putative demixed source signal and each respiratory belt signal was calculated and compared using a t test.Demixing of respiratory signals obtained from two simultaneous sleepers was performed using a hidden Markov model. Mechanical respiratory sources were interpreted as latent signals that evolve in a stochastically continuous manner, according to a linear additive Gaussian model, mixed through a linear operation with additive sensor noise to give rise to the signals at the four detectors. Interpreting the linear operation as unknown, we used the Expectation\u2013Maximization algorithm to obtain the maximum-likelihood estimateBedScales were installed beneath the legs of a conventional hospital bed in the Clinical and Translational Research Institute where overnight sleep studies were conducted. As part of another ongoing study, subjects underwent standard in-laboratory polysomnography (PSG) with electroencephalogram (EEG), electro-oculogram, submental and leg electromyogram for sleep staging; nasal pressure and thermistor for airflow measurement; thoracic and abdominal piezoelectric bands for respiratory effort; arterial oxygen saturation monitoring at the finger; and electrocardiogram monitoring for safety. Patients slept supine. Sleep state, arousals, and respiratory events were scored by a registered sleep technologist according to standard American Academy of Sleep Medicine 2012 Recommended Criteria. Signals from the thoracic piezoelectric band and the BedScales were aligned using custom python scripts.BedScales were delivered to and installed under each patient\u2019s bed (or recliner or couch). Signals were recorded locally and transmitted securely via the patient's WiFi to an Amazon Web Services S3 bucket which activated a cloud pipeline of analytics that extracted the physiologic parameters automatically. The daily average respiratory rate from each patient was derived by averaging the epochs (defined every 30\u00a0s) that had a physiologically reasonable bpm (bpm\u2009>\u20096 and bpm\u2009<\u200940). Values in the respiratory heatmap below 10\u00a0bpm were not displayed. The average heart rate from each patient was derived by averaging the epochs that had a bpm\u2009>\u200960 and\u2009<\u2009120. Values in the heart rate heatmap below 60\u00a0bpm were not displayed.P values less than 0.05 were considered significant and are indicated by asterisks as follows: *P\u2009<\u20090.05, **P\u2009<\u20090.01, ***P\u2009<\u20090.001, ****P\u2009<\u20090.0001.Statistical analysis was performed using custom python scripts or GraphPad Prism software. All data are represented as mean values\u2009\u00b1\u2009standard deviation unless indicated otherwise. For two-group comparisons, a two-tailed nonparametric Mann\u2013Whitney test was used unless otherwise specified. All analyses except respirophasic inspiratory versus expiratory BCG magnitudes were unpaired. Supplementary Figures.Supplementary Table S1."} +{"text": "Recent evidence suggests that the box H/ACA small nucleolar RNA (snoRNA)-ended long noncoding RNA (lncRNA), SLERT, plays a critical role in gene regulation. However, its role in cancer remains undetermined. Herein, we explored its implication in human endometrial cancer (EC).EC plasma and tissue samples were collected for the detection of SLERT expression using qRT-PCR method. The functional investigation was tested by CCK-8 and transwell assays. Luciferase reporter, RNA pull-down, and immunoprecipitation (RIP) assays were used to determine the regulatory network involved in SLERT. The in vivo effect of SLERT was tested by caudal vein lung metastasis model.6A levels of BDNF mRNA; then, the m6A sites were read by IGF2BP1, enhancing BDNF mRNA stability, followed by the activation of BDNF/TRKB signaling, an inducer of EMT. The animal model showed that overexpression of SLERT increased EC cell lung metastasis, and this effect was effectively blocked by BDNF silencing or treatment with TRKB inhibitor k252a. Clinically, EC patients have high levels of SLERT both in tissue or plasma, which might be used as a biomarker of diagnosis and prognosis.Stable knockdown of SLERT significantly inhibited EC cell (KLE and AN3CA) migration and invasion, while it did not affect cell viability. SLERT induced epithelial-mesenchymal transition (EMT) via elevating N-cadherin and Vimentin and downregulating E-cadherin. Further investigation showed that SLERT directly binds to METTL3, increasing the mOur findings, for the first time, uncover the metastasis-promoting effect of SLERT in EC via in vitro and in vivo evidence, providing a potential therapeutic target for metastatic EC treatment.The online version contains supplementary material available at 10.1186/s12957-022-02821-w. Endometrial cancer (EC) is a kind of epithelial malignant tumor occurring in the endometrium, most commonly occurring in perimenopausal and postmenopausal women . It is tLong non-coding RNA (lncRNA) is a class of endogenous RNA molecules that do not encode proteins and the transcript length is over 200nt, which can regulate gene expression on multiple levels . LncRNA Recently, a novel lncRNA, SLERT, has been reported to enhance pre-rRNA transcription via evicting DDX21 suppression on Pol I transcription, leading to decreased tumorigenesis , 12. How2.This study included 92 EC tissues and 28 normal tissues, which were collected from Huaihe Hospital of Henan University. The fresh tissues were stored in liquid nitrogen immediately to prevent RNA degradation. In addition, plasma samples were also collected from 40 EC patients and 18 healthy volunteers. Those who had received preoperative chemoradiotherapy were excluded. All procedures are approved by the Medical Institutional Review Board of Huaihe Hospital of Henan University . Two EC cells KLE and AN3CA were commercially purchased from China Center for Type Culture Collection (CATCC). DMEM medium was used to culture cells at 37\u00b0C with 5% COTotal RNA was extracted from EC cells using Trizol reagent . The Cytoplasmic & Nuclear RNA Purification Kit was used to isolate cytoplasmic and nuclear RNA fragments. Then, reverse transcription was conducted using SuperScript\u2122 reverse transcriptase (Invitrogen) and gene amplification and quantification were tested by PowerUp\u2122 SYBR\u2122 Green (Invitrogen) with the QuantStudio\u2122 7 Pro system (Invitrogen). GAPDH was used as the reference control for quantifying SLERT and BDNF expression.To visualize the location of SLERT in cells, the FISH Kit was purchased and the assay was conducted as per the manufacturer\u2019s instructions with minor modifications. In brief, the FAM-labeled probe against SLERT was synthesized and incubated with cells permeated with ethanol at 37\u00b0C for 4h. After washing for six times, cells were counterstained with DAPI and then observed with fluorescence microscope.Four shRNAs targeting SLERT were designed and inserted into YSH-LV001-shRNA lentiviral vector , followed by infection into KLE and AN3CA cells in the presence of polybrene. The stable cell lines were screened by 2ug/mL puromycin. The siRNAs targeting IGF2BP1, BDNF, and METTL3 were commercially purchased from Santa Cruz Biotechnology . To overexpress BDNF and SLERT, the full-length sequence was synthesized and inserted into pcDNA 3.0 vector (Invitrogen).Cell viability was detected by CCK-8 solution in 96-well plates, followed by assessment of absorbance at 450nm with the automatic enzyme-linked immunosorbent assay system. Cell migration was tested by wound healing assay, the migration distance in each well was recorded at 48h. Transwell chamber coated with matrigel was used to test cell invasion; the invaded cells were stained by crystal violet.KLE and AN3CA cells with stable SLERT knockdown were lysed by RIPA buffer supplemented with protease inhibitor cocktail . Protein quantification was carried out using Pierce\u2122 BCA Protein Quantitative Kit (Invitrogen). Total protein was separated by 10% SDS-PAGE gel and transferred to PVDF membrane. After blocking with 5% skimmed milk, the membrane was incubated with primary antibodies: anti-E-cadherin , anti-N-cadherin , anti-Vimentin , anti-Snail , anti-METTL3 , anti-BDNF , anti-TRKB , and anti-GAPDH . Lastly, the PVDF membrane was developed using SuperSignal West Pico PLUS (Invitrogen).The promoter of BDNF (2000bp) was synthesized and inserted into pGL3-basic vector , followed by transfection into HEK293T, KLE, and AN3CA cells using Lipofectamine 3000 (Invitrogen) according to the manufacturer\u2019s instructions. The promoter activity was tested by luciferase reporter system (Promega).6A antibody , the enrichment of BDNF mRNA was tested by qRT-PCR method.Total RNA was extracted from KLE and AN3CA cells using Trizol solution. Then, RNA was purified and fragmented with dynabeads mRNA Purification Kit (Invitrogen) and RNA fragmentation reagent (Invitrogen), respectively. After incubation with anti-mThe biotin-labeled probes against SLERT and its anti-sense were synthesized and incubated with cell lysate at 4\u00b0C overnight. After incubation with Magnetic Streptavidin Beads (Invitrogen) at 25\u00b0C for 2h, the enriched protein was washed and subjected to western blot assay. For RIP assay, the corresponding antibodies were added into cell lysate, and the enriched RNA was purified and tested by qRT-PCR assay.n=5 per group), labeled as OE-vector , OE-SLERT , OE-SLERT+ASO-BDNF , and OE-SLERT+k252a . After 4 weeks, the lung tissues were dissected and weighted, followed by H&E staining.A total of 20 NOD/SCID mice of similar weight were used in this study, which were approved by the Institutional Animal Care and Use Committee of Huaihe Hospital of Henan University. The mice were randomly divided into four groups or two-way (Bonferroni post hoc test) ANOVA, as appropriate. ROC curve and Kaplan-Meier plot were used to test the diagnostic and prognostic values of SLERT in EC, respectively. All results are the mean\u00b1SD of at least three independent experiments carried out in triplicate. P = 0.008), with an area under of ROC (AUC) value of 0.75 (95% CI: 0.6604~0.8476) (P < 0.001) , depth of invasion (P < 0.001), lymphovascular invasion (P = 0.006), and lymph node metastasis (P = 0.018), while it did not correlate with age and histology type (Table P = 0.006) , with an AUC value of 0.84 (95%CI: 0.7384~0.9394) Fig. A, wherea01) Fig. B and MMP01) Fig. C. Import02) Fig. D, hintin02) Fig. E. The re02) Fig. F\u2013H. Howe01) Fig. I, J, imp01) Fig. K, L.Fig.6A modification is critical for the stability of mRNA [6A. As expected, the m6A levels of BDNF were markedly decreased after SLERT knockdown (P < 0.01) Fig. A, B; oppner Fig. C, D. Fur01) Fig. E, F. Imp01) Fig. G; simila01) Fig. H, I, sugd by m6A . Silenci01) Fig. J, K. Mor01) Fig. L, M and 01) Fig. N, O.Fig.P < 0.05). However, the above effects were partially abolished by silencing of METTL3 or BDNF and lung/weight index were notably increased in SLERT-overexpressing group of cancer-related deaths are caused by primary tumor cells migrating far from their original site . Metasta6A methylation is an important epigenetic modification, which is a dynamic and reversible process regulated by methylase (writer), demethylase (eraser) and reader proteins that recognize m6A-modified RNA [6A are frequently dysregulated in human cancer, linking to malignant phenotype and dismal prognosis [6A writers, erasers, and readers in EC by comprehensive immunohistochemical analysis and found that METTL3, METTL14, FTO, HNRNPA2B1, and HNRNPC were significantly highly expressed, which were linked to shortened overall survival in EC [6A via METTL3, and this process was inhibited after SLERT knockdown, suggesting that SLERT is required for BDNF m6A methylation catalyzed by METTL3. Further RNA pull-down and RIP data revealed that SLERT directly bound to METTL3, facilitating the binding of METTL3 to BDNF mRNA. The fate of m6A-modified RNA varies according to different readers, including degradation, increased stability, or translation promotion or suppression. IGF2BP1 is a well-documented mRNA stability enhancer via m6A recognition [6A-dependent manner and promoted EC cell proliferation and invasion [6A and read by IGF2BP1, resulting in increased mRNA stability and maintaining breast cancer stem cell stemness [mfied RNA . It is ified RNA . The keyrognosis . A very al in EC . Herein,invasion . Myc prostemness . In this6A-dependent manner. Moreover, this study provides a potential diagnostic, prognostic marker as well as therapeutic target for patients with metastatic EC patients.In sum, our findings uncover that SLERT acts as a pro-metastasis lncRNA in EC by activating BDNF/TRKB signaling in a mAdditional file 1: Figure S1. Western blot analysis of Snail levels in KLE and AN3CA cells with SLERT knockdown.Additional file 2: Figure S2. The body weight of the mice in these four groups."} +{"text": "Ventilator-associated pneumonia (VAP) remains a common risk in mechanically ventilated patients. Different care bundles have been proposed to succeed VAP reduction. We aimed to identify the combined interventions that have been used to by ICUs worldwide from the implementation of \u201cInstitute for Healthcare Improvement Ventilator Bundle\u201d, i.e., from December 2004. A search was performed on the PubMed, Scopus and Science Direct databases. Finally, 38 studies met our inclusion criteria. The most common interventions monitored in the care bundles were sedation and weaning protocols, semi-recumbent positioning, oral and hand hygiene, peptic ulcer disease and deep venus thrombosis prophylaxis, subglottic suctioning, and cuff pressure control. Head-of-bed elevation was implemented by almost all studies, followed by oral hygiene, which was the second extensively used intervention. Four studies indicated a low VAP reduction, while 22 studies found an over 36% VAP decline, and in ten of them, the decrease was over 65%. Four of these studies indicated zero or nearly zero after intervention VAP rates. The studies with the highest VAP reduction adopted the \u201cIHI Ventilator Bundle\u201d combined with adequate endotracheal tube cuff pressure and subglottic suctioning. Multifaced techniques can lead to VAP reduction at a great extent. Multidisciplinary measures combined with long-lasting education programs and measurement of bundle\u2019s compliance should be the gold standard combination. Ventilator-associated pneumonia (VAP) is one of the main types of infection in critically ill mechanically ventilated patients, leading to increased mortality, morbidity, hospital stay, economic and psychological costs for patients and their families ,2,3,4. DIn 1983, the CDC published the guidelines for the prevention of nosocomial pneumonia, which were specialized for VAP in 2003 .Staphylococcus aureus (CA-MRSA) and hospital-associated MRSA (HA-MRSA). Apart from that, the communities have faced several viral outbreaks, such as SARS-1, SARS-2, and MERS. All the above severe respiratory syndromes and population aging have led to increased rates of ventilated patients [In the last two decades, a major problem has been the increasing rates of occurrence of community-associated methicillin resistant patients . As VAP A great number of studies examined the effectiveness of the different combinations of interventions for VAP prevention. Several reviews have summarized the findings of those studies ,12,13,14A systematic literature review was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines . A comprThree independent authors were responsible for removing the duplicates, screening the title and abstract, and analyzing the full content of the studies in accordance with the inclusion criteria. Two independent authors selected the included studies and another independent author resolved possible disagreements. To be included in our systematic review, studies had to (a) be published in the English language; (b) be pre\u2013post observational studies; (c) include adult critically ill patients, intubated at least 48 h, admitted to all kinds of adult ICUs; (d) evaluate the implementation of care bundles in VAP prevention by thoroughly presenting all the combined interventions and calculating the pre and post intervention VAP rate; (e) compare with the individual intervention\u2019s implementation for VAP prevention; and (f) be published after the implementation of \u201cIHI Ventilator Bundle\u201d. Additionally, we excluded protocols, conference papers, abstracts, posters, and letters to editors and editorials.For each study, the main characteristics were extracted: authors, country, data collection period, study setting, sample size, age of patients, measures, implementation of an educational program, and main findings. The appraisal of quality of the included studies was performed by using the Risk of Bias in Non-randomized Studies of Interventions (ROBINS-I) . The cheThe study selection process is demonstrated in Finally, 38 studies were included in our review. The main characteristics of the studies are presented in The risk of bias assessment of the reviewed studies is demonstrated in All of the studies used multifaceted strategies to prevent VAP, as it was one of the inclusion criteria. The number of the included ventilator bundle elements varied from four to thirteen and is shown in Patients\u2019 outcomes are shown in p-values of less than 0.05 were considered significant.In nine studies ,26,30,422O and subglottic suctioning. Moreover, the study of Gallagher et al. (2012) [The studies with the highest rate of reduction ,19,25 im. (2012) indicate. (2012) , where \u201cWe identified thirty-three studies that adopted an education program, as demonstrated in Our systematic review demonstrated the variety of interventions included in ventilator care bundles and their implementation in VAP decrease in adult ICUs. Thirty-eight studies met our inclusion criteria and we found that the combined measures can prevent VAP toa greater extent. Moreover, our review showed that the number of intervention strategies varied widely in the included studies. Most of the studies examined whether ventilator bundle\u2019s compliance could be increased through health workers\u2019 training.VAP is one of the main healthcare-associated infections in intubated critically ill patients. VAP is associated with an exceeded duration of mechanical ventilation. The most common microorganisms responsible for VAP are pseudomonas aeruginosa, escherichiacoli, klebsiella pneumoniae and the Acinetobacter species from Gram-negative microorganisms and staphylococcus aureus from Gram-positive microorganisms. VAP diagnostic criteria differ among ICUs but usually require factors such as fever, leukocytosis, progressive infiltrate on chest X-ray, positive cultures from respiratory secretions and reduction in gas exchange ,9.During the last decades, there has been a great scientific concern about finding the best strategies to prevent VAP and ventilator-associated events . WorldwiA multimodal approach for VAP prevention includes functional, mechanical, and pharmacological measures . The mosA semi-recumbent position was established with the introduction of \u201cIHI Ventilator Bundle\u201d, though it was proposed much earlier in studies conducted from 1992 to 1999 to test its contribution in aspiration prevention ,45,46,47A notable result of our study is that ten studies did not control the daily readiness for extubating, when the presence of the endotracheal tube is one of the major predisposing factors for VAP development, as pathogens enter the trachea either by micro aspiration around the ETT cuff or by the biofilm formed in the inner side of the ETT. Moreover, only nine of the included studies followed the \u201cIHI VAP preventive Bundle\u201d. All of them, achieved a VAP reduction at least 36%, five of them showed a reduction of over 65% and two of them managed a zero after intervention VAP rate. These results may indicate the important contribution of the IHI VAP preventive Bundle, alone or in combination with other measures, in VAP reduction. It is noteworthy that the revised studies, carried out the last six years, did not use the \u201cIHI Ventilator Bundle\u201d but adopted some of its interventions.Another thing that was remarkable was that in many studies, hand hygiene was not mentioned to be monitored as an element in the VAP care bundles. We assume that measures such as hand hygiene and aseptic suctioning technique are of the most basic techniques and that they were taken for granted, along with all the other interventions for VAP prevention in the included studies. Another interesting conclusion in some studies ,23,28,36Over the past two decades, humanity has faced several viral outbreaks, causing severe respiratory syndromes, such as SARS-1, SARS-2 and MERS, which may have increased the percentage of intubated patients at a local level. According to the inclusion criteria, all the included studies were conducted after December 2004. Taking into consideration the data collection periods of the revised studies, in two studies ,19 carriThe study of Alvarez-Lerma et al. (2018) was the Finally, most of the included studies, apart from the implementation of care bundles, adopted at the same time a multifaced educational program. Presentations, posters, videos, discussions, stimulating scenarios and feedback meetings were some of the ways that ICU staff was trained. Our systematic review underlined the wide range in VAP reduction. This wide fluctuation may have been affected, to some extent, by either the country\u2019s healthcare system or the extent variety of educational programs and different degree of personnel\u2019s adherence, but at the same time the studies\u2019 results highlighted that multidisciplinary health professionals\u2019 education and frequent monitoring of adherence can lead to better patients\u2019 outcomes.There are several limitations in this systematic review. At first, as we searched for relevant studies only in English language, other potential studies written in different languages were not included in our review. Also, we included three databases in our research, and thus additional studies could be found in other databases. In addition, application of vigorous inclusion and exclusion criteria may preclude some studies to be included in our review. Moreover, among the reviewed studies, there was a heterogeneity, to a greater extent, of the study settings. In particular, differences in study quality are indicative of great differences in studies design, since seven studies had a high level of quality, 20 had a moderate level, and 11 a low level. However, we should notice that heterogeneity among studies was not a limitation of our review but an unavoidable effect of the studies design. Therefore, we should consider our results with caution since it is difficult to establish solid conclusions.From our systematic review, it emerged that there is a considerable variation between the VAP care bundles and education programs in ICUs worldwide. This variation leads to discrepancies and does not allow comparability, to a greater extent, between the studies to find the gold standard VAP care bundle. The IHI VAP preventive Bundle seems to be a very useful tool in VAP reduction, combined with adequate ETT cuff pressure and subglottic suctioning, without forgetting the hand hygiene and aseptic suctioning technique. ICUs should adopt basic practices that prevent or decrease VAP rates, and as a result, mortality, duration of mechanical ventilation, length of stay, and healthcare costs. Moreover, the strategies should be multifaceted and supported by a long-term education program by ensuring compliance in the care bundle. These multidisciplinary strategies and education programs should be common in all ICUs. At least in the same country, national or cross-sectional randomized controlled studies need to be carried out in order to be able to compare the measures\u2019 efficacy and find the best combination of preventive interventions."} +{"text": "To compare the value of contrast-enhanced CT (CECT) and non-contrast-enhanced CT (NCECT) radiomics models in differentiating tuberculosis (TB) from non-tuberculous infectious lesions (NTIL) presenting as solid pulmonary nodules or masses, and develop a combine radiomics model (RM).This study was a retrospective analysis of 101 lesions in 95 patients, including 49 lesions (from 45 patients) in the TB group and 52 lesions (from 50 patients) in the NTIL group. Lesions were randomly divided into training and test sets in the ratio of 7:3. Conventional imaging features were used to construct a conventional imaging model (IM). Radiomics features screening and NCECT or CECT RM construction were carried out by correlation analysis and gradient boosting decision tree, and logistic regression. Finally, conventional IM, NCECT RM, and CECT RM were used for combine RM construction. Additionally, we recruited three radiologists for independent diagnosis. The differential diagnostic performance of each model was assessed using the areas under the receiver operating characteristic curve (AUCs).The CECT RM outperformed the conventional IM , the NCECT RM , and three radiologists. The diagnostic efficacy of the combine RM was best in the training and test sets.The diagnostic efficacy of the CECT RM was superior to that of the NCECT RM in identifying TB from NTIL presenting as solid pulmonary nodules or masses. The combine RM had the best performance and may outperform expert radiologists. In 2020, approximately 9.9 million people worldwide fell ill with tuberculosis (TB), equivalent to 127 cases per 100,000 population. The majority of these patients are located in the WHO regions of South-East Asia 43%), Africa 25%) and the Western Pacific 18%) .This study was approved and exempted the informed consent of patients by the ethics committees of the three hospitals.Inclusion criteria: (1) TB was confirmed by sputum testing or pathology combined with DNA testing, clinical symptoms, medical history, and tuberculin testing, and NTIL was confirmed by pathology or effective anti-inflammatory therapy; (2) lesions located in the peri-pulmonary field; (3) CECT images with layer thickness \u2264 2.5 mm on initial examination; (4) lesions presenting as solid nodules or masses without internal calcification; (5) patients with no history of lung surgery, radiotherapy or chemotherapy. According to the inclusion criteria, 73 patients from the First Affiliated Hospital of Dalian Medical University (Center 1) from January 2010 to February 2022, 25 patients from the Second Affiliated Hospital of Dalian Medical University (Center 2) from April 2019 to February 2022, and 15 patients from Dalian Public Health Clinical Center (Center 3) from December 2017 to July 2021 were collected retrospectively. The exclusion criteria were as follows: (1) Artifacts in CT images had an impact on evaluation. (2) Patients did not have NCECT images with slice thickness \u2264 2.5 mm. Eighteen patients were excluded according to the exclusion criteria.The patient selection process is shown in Two demographic characteristics (gender and age), two clinical characteristics , and five hematological examination indices were recorded at the time of patient admission.Chest CT scans were performed using multi-slice spiral CT (16 or more) from three companies . The patient was routinely scanned from the tip of the lung to the base under inspiration. Scanning parameters: tube voltage, 120\u2013140 kVp; tube current, 140\u2013630 mA or automatic adjustment; matrix, 512 \u00d7 512; reconstruction thickness, 1\u20132.5 mm; reconstruction interval, 1\u20132.5 mm. CECT scans were performed using a non-iodine ion contrast agent, with an injection rate of 2.5\u20133.0 mL/s, and scanned at 55\u201360 s after contrast injection.CT imaging analysis was done jointly by two thoracic diagnostic radiologists (A and B) without knowing the type of lesions, and different opinions were discussed and agreed upon. Each lesion was observed in the lung window , mediastinal window , axial, sagittal, and coronal views. The conventional imaging features of each lesion were analyzed, including (1) semantic features: lobulation, spiculation, cavity, pleural traction, and location; and (2) quantitative features: axial position maximum diameter. The apical and posterior segments of the upper lobe (S1 and S2), as well as the superior segment of the lower lobe (S6), are TB predilection sites . Therefohttp://slicer.org/) by radiologist A. The regions of interest (ROIs) were outlined layer by layer on the entire lesion from NCECT and CECT images respectively under a mediastinal window and the cavity areas were removed. Radiologist A blinded the pathological diagnosis of the lesions.The original images were imported into the open source software 3D slicer , gray-level dependence matrix (GLDM), gray-level run length matrix (GLRLM), gray-level size zone matrix (GLSZM), and neighborhood gray-tone difference matrix (NGTDM).Assessment of intra- and inter-group agreement was performed by randomly selecting 32 lesions from 101 lesions, and independent measurements were completed by radiologist A and radiologist B with unknown final pathological results. And independent measurements of data were completed again by radiologist A 1 month after the first measurement. Inter- and intra- class correlation coefficients (ICCs) were used to assess the inter- and intra- observer agreement of feature extraction, with ICC > 0.75 indicating good to excellent agreement .via a linear combination of selected features that were weighted by their respective coefficients.Firstly, the radiomics feature data of the training and test sets were normalized respectively. Secondly, the radiomics features were screened in the training set using correlation analysis and gradient boosting decision tree (GBDT). Differences in conventional imaging characteristics of TB and NTIL groups were compared by univariate analysis. Finally, logistic regression was used to establish four models, i.e., conventional imaging model (IM), NCECT RM, CECT RM, and combine RM. The combine RM was formed by the combination of conventional IM, NCECT RM, and CECT RM. The radscore was calculated for each patient The receiver operating characteristic (ROC) curves were performed to evaluate the discriminative performance of the models. The areas under the ROC curves (AUCs), sensitivities, specificities, and accuracies were calculated. The calibration curves were plotted, and the fits of the models were estimated by the Hosmer-Lemeshow (H-L) test. The decision curve analysis (DCA) was performed to estimate the clinical utility of the models.We invited a senior radiologist C (with 25 years of diagnostic chest imaging experience), a Mid-level radiologist D (with 12 years of diagnostic chest imaging experience), and a junior radiologist E (with 4 years of diagnostic chest imaging experience) to diagnose all lesions independently. Their diagnostic ability in identifying TB from NTIL presenting as solid pulmonary nodules or masses was assessed. The value of RMs was further clarified by comparison with radiologists.t-test was used; when they did not conform to the normal distribution, they were expressed as median and Mann-Whitney U test was used. A two-tailed p-value < 0.05 indicated statistical significance.All statistical analyses of the data were performed using R (version 4.1.1). Categorical variables were expressed as the \u201cnumber of cases (percentage)\u201d and the chi-square test or Fisher's exact test was used to evaluate differences between the TB and NTIL groups. When continuous variables conformed to the normal distribution, they were expressed as mean \u00b1 standard deviation and There were 45 patients (49 lesions) in the TB group , and 50 patients (52 lesions) in the NTIL group . The patients in the NTIL group included 7 cases of pulmonary fungal infection, 11 cases of organized pneumonia, 9 cases of inflammatory granuloma, and 23 cases of pulmonary inflammation.P > 0.05). In the test set, more patients in the TB group had a history of smoking than in the NTIL group (P = 0.025).The clinical data of patients in the training and test sets are shown in P = 0.001). In the test set, the differences in all features were not significant (P > 0.05).The comparison of conventional imaging features between two groups in the training and test sets is shown in The radiomics feature selection processes were the same for both the NCECT and CECT RMs. A total of 107 radiomics features were extracted. First, 17 features with ICC < 0.75 were excluded. The consistency assessment showed the ranges of the inter- and intra-class correlation coefficient values for the remaining 90 radiomics features were 0.765\u20130.994 and 0.853\u20131.000, respectively. This indicated good to excellent inter- and intra-observer agreement for the 90 radiomics features. Then, after filtering by correlation analysis, 28 NCECT radiomics features and 29 CECT radiomics features were retained. Finally, the GBDT algorithm selected 11 NCECT radiomics features and 10 CECT radiomics features from them as the best features.All conventional imaging features were used to establish the conventional IM. The modeling formulas of the NCECT and CECT RMs are shown in In the training set, the AUCs of NCECT RM and CECT RM were higher than that of the conventional IM . CompareIn this study, the best performance was achieved with the combine RM in the training and test sets, with AUCs of 0.922 and 0.833 , accuracies of 0.829 and 0.774 .P > 0.05, In the training set, the H-L test showed no significant difference between the four models and the ideal model .Our results showed that cavity was the only conventional imaging feature that differed significantly between the two groups in the training set, and it was more common in TB lesions, accounting for 32.35%. An analysis of the imaging presentation of pulmonary granulomatous lesions also mentioned that cavity was highly suggestive of Mycobacterium tuberculosis infection . The proGiven that there was only cavity with the discrepancy, we decided to use all conventional imaging features for modeling to avoid missing important information. However, even though conventional IM integrated all conventional imaging features, its diagnostic power is unsatisfactory .Essentially, CT images provide much more information than the macroscopic appearances we can see with the naked eye. Radiomics then provides us with a large number of quantitative features that reflect the microstructure of the lesion, mainly including intensity histogram, shape-based features, and texture features, and the extraction process of these features is automatic , 25. TheWang et al.'s RM based on NCECT to identify TB and community-acquired pneumonia in children was also quite effective, the AUC reached 0.837 . In addiCECT scan is a commonly used auxiliary examination method in clinical diagnosis and treatment. The results of several studies have shown that the radiomics characteristics of tumors are altered after the uptake of contrast agents, particularly first-order histogram features and textural features \u201332. SeveThe AUC and accuracy of the combine RM were the highest among all models, and all indicators were relatively balanced in the training and test sets. This study also found that radiologists were not able to distinguish well between TB and NTIL presenting as solid pulmonary nodules or masses, even for experienced radiologists . Moreover, there is a great deal of subjectivity and fortuity in the diagnosis of radiologists with different qualifications. Computer-aided diagnosis has become an irreversible trend in clinical work, and artificial intelligence has great prospects in the field of thoracic radiology . The comThere are some limitations to our study. Firstly, this is a retrospective study with unavoidable selection bias. Secondly, after strict inclusion and exclusion criteria, our sample size was small, so a larger sample is needed to validate this study in the future. Thirdly, the NTIL group in this study has a large variety of diseases, and the value of using radiomics to help differential diagnosis of one of these diseases with TB will be discussed in the future. Fourthly, only CECT images scanned 55\u201360 s after contrast agent injection were selected in this study, the influence of different delayed scanning times on CECT's differential diagnostic ability can be further studied in the future. Fifthly, the use of multiple CT scanners in this study affected the reproducibility of the radiomics data. Although we used resampling for correction, we still need to use batch correction to minimize acquisition-related radiomics variability and verify the generality of the final model by stress testing in future studies. Finally, we used the rather time-consuming manual method of outlining ROIs. Compared with it, semi-automatic measurements are not only convenient and fast but also may have a better inter-observer agreement .In conclusion, radiomics helped to differentiate TB from NTIL presenting as solid pulmonary nodules or masses, and CECT may be a better choice. Combine RM we built obtained the best diagnostic efficacy and may outperform expert radiologists.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.Approval was obtained from the Ethics Committee of the First Afliated Hospital of Dalian Medical University, the Ethics Committee of the Second Hospital of Dalian Medical University, and Dalian Public Health Clinical Center Ethics Committee. Written informed consent for participation was not required for this study in accordance with the national legislation and the institutional requirements.WZ: conceptualization, methodology, investigation, data curation, writing\u2014original draft, and writing\u2014review and editing. ZX: investigation, writing\u2014original draft, and writing\u2014review and editing. DT: validation, investigation, and data curation. KW, XL, and DQ: resources and data curation. MZ: software, visualization, and formal analysis. ZL: conceptualization, investigation, writing\u2014review and editing, supervision, and project administration. All authors contributed to the article and approved the submitted version.Author MZ was employed by GE Healthcare. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."} +{"text": "Leishmania genus. Benzylamines are a class of compounds selectively designed to inhibit the squalene synthase (SQS) that catalyzes the first committed reaction on the sterol biosynthesis pathway. Herein, we studied seven new benzylamines (SBC 37\u201343) against Leishmania amazonensis. After the first screening of cell viability, two inhibitors (SBC 39 and SBC 40) were selected. Against intracellular amastigotes, SBC 39 and SBC 40 presented selectivity indexes of 117.7 and 180, respectively, indicating high selectivity. Analysis of the sterol composition revealed a depletion of endogenous 24-alkylated sterols such as episterol and 5-dehydroepisterol, with a concomitant accumulation of fecosterol, implying a disturbance in cellular lipid content. This result suggests a blockade of de novo sterol synthesis at the level of SQS and C-5 desaturase. Furthermore, physiological analysis and electron microscopy revealed three main alterations: (1) in the mitochondrion; (2) the presence of lipid bodies and autophagosomes; and (3) the appearance of projections in the plasma membrane. In conclusion, our results support the notion that benzylamines have a potent effect against Leishmania amazonensis and should be an exciting novel pharmaceutical lead for developing new chemotherapeutic alternatives to treat leishmaniasis.Leishmaniasis is a neglected disease caused by protozoan parasites of the The leishmaniasis transmission was reported in 98 countries and 3 territories on 5 continents. The estimated world prevalence for all clinical manifestations of the disease is 12 million, with 58,000 visceral leishmaniasis (VL) cases and 220,000 cutaneous leishmaniasis (CL) cases per year2. Multiple species, including L. amazonensis and L. braziliensis, are the causative agents of cutaneous leishmaniasis (CL) in the New World. Furthermore, L. amazonensis can also cause mucocutaneous leishmaniasis (MCL), which results in progressive destruction of the naso-oropharyngeal mucosa, and diffuse cutaneous leishmaniasis (DCL), the most severe cutaneous form of the disease that is characterized by a diffuse infiltration in the skin of the patients and highly resistant to all kind of chemotherapy available4. Besides the severe clinical manifestations of leishmaniasis, few drugs are available for its treatment.Leishmaniasis is an endemic neglected disease caused by several species of 5. Second-line treatments are based on the use of amphotericin B formulations and pentamidine. In some countries in Asia, Africa, and Europe, miltefosine (Impavido) is currently the first-line treatment6. Among the chemotherapeutic agents available, miltefosine is the only orally available treatment; however, it is teratogenic and not indicated for women of fertile age8. None of the available drugs can be considered ideal due to their high toxicity, long duration of treatment, and severe side effects, which often lead to treatment abandonment. Furthermore, these treatments do not eliminate the parasite in the infected individuals10.Pentavalent antimonials have been the first treatment choice for more than 70\u00a0years in some countries worldwide, despite the parenteral route of administration, high toxicity, and cost17. Benzyl farnesyl amine mimetics are one class of selective inhibitors of the squalene synthase (SQS), an important enzyme that catalyzes the condensation of two molecules of farnesyl pyrophosphate to produce squalene. SQS catalyzes the first committed step in sterol biosynthesis, and its inhibition does not affect the synthesis of isoprenoids, which are also essential molecules for eukaryotic cells. For years, the research for specific squalene synthase inhibitors (SQSi) has involved intensive efforts of industrial and academic investigators because of the potential use of these inhibitors in the treatment of coronary heart disease and hypercholesterolemia20.Trypanosomatids and fungi have an endogenous requirement of ergosterol and other 24-alkylated sterols for growth and survival, which are absent in mammal cells. Thus, the enzymes involved in the sterol biosynthesis pathway are interesting targets for new treatments, and several works have shown the effect of different sterol biosynthesis inhibitors (SBIs) in trypanosomatids23, we early investigated the antiprotozoal potential of BPQ-OH (3-biphenyl-4-yl-3-hydroxyquinuclidine), an arylquinuclidine, with significant biological activity against T. cruzi and L. mexicana, resulting in growth inhibition, cell lysis, and complete depletion of endogenous squalene and 24-methyl sterols27. Against L. amazonensis, BPQ-OH presented a potent growth inhibition effect against both developmental stages of the parasite leading to several changes in the ultrastructure of promastigotes28. Furthermore, the other two quinuclidine derivatives, E-5700 and ER-119884, also showed potent antiproliferative effects against T. cruzi30 and L. amazonensis, also in combination with C14\u03b1-demethylase inhibitors [azoles]23.Among the selective inhibitors of the SQSLeishmania amazonensis. Several aspects of the anti-Leishmania activity of these compounds were investigated at different times of treatment; we studied the antiproliferative, ultrastructural, and biochemical effects induced by them. Moreover, we found one derivative that is among the most selective compounds for the parasite with low toxicity for the mammalian cells.This work evaluated the effect of another class of selective SQS inhibitors, the novel mimetics of benzyl farnesyl amine, against Leishmania amazonensis used in this study was gently provided by the Leishmania Collection of the Instituto Oswaldo Cruz (code IOCL 0071-FIOCRUZ). It has been maintained via inoculation into the base of BALB/c mouse tails. Amastigotes were obtained from the lesions of infected mice and transformed into promastigotes that were maintained in Warren\u2019s medium [brain heart infusion plus hemin and folic acid]24 supplemented with 10% fetal bovine serum (FBS) at 25\u00a0\u00b0C. Infective metacyclic promastigotes were used to obtain intracellular amastigotes in macrophage culture. Firstly, peritoneal macrophages from BALB/c mice were harvested by washing them with Hanks\u2019 solution and plated in 24-well tissue culture chamber slides, allowing them to adhere to the slides for 24\u00a0h in RPMI medium (Gibco) supplemented with 10% FBS at 37\u00a0\u00b0C in 5% CO2. After this, adherent macrophages were infected with metacyclic promastigotes at a macrophage-to-parasite ratio of 1:10 at 35\u00a0\u00b0C in 5% CO2 for 2\u00a0h. These cultures were maintained for 24\u00a0h in RPMI medium supplemented with 10% FBS for the assays with intracellular amastigotes.The MHOM/BR/75/Josefa strain of 25. Structures of those N-[4-[benzyloxy] benzyl]-benzene-methaneamine derivatives, SBCs 37\u201343, are displayed in Fig.\u00a0Trypanosoma cruzi25. Compounds were used as hydrochloride salts, purified by one recrystallization from analytical grade ethanol, and dried in a high vacuum at room temperature.The benzyl farnesyl amine mimetics, SBCs 37\u201343, were prepared by chemical synthesis at IQ-UNICAMP according to an experimental procedure previously described by C\u00e4mmerer and SouzaL. amazonensis promastigotes and peritoneal macrophages using the CellTiter 96\u00ae\u00a0Aqueous MTS Assay 29. For analysis in promastigotes, we started the culture at a cell density of 1\u2009\u00d7\u2009106 cells/ml in Warren\u2019s medium supplemented with 10% FBS. After 24\u00a0h, different concentrations of SBC 37\u201343 were added to the cultures. Cell viability and the cytotoxic effects were measured at 24, 48, and 72\u00a0h of treatment, when all groups, including untreated, were transferred to a transparent 96-well plate in triplicate. MTS/PMS assay reaction was quantified by optical density measurement at 490\u00a0nm in a microplate reader and SpectraMax M2/M2espectrofluorometer . As a negative control, parasites were fixed with 0.4% nascent formaldehyde for 10\u00a0min at room temperature before the incubation. Cytotoxicity effects of SBCs 37\u201343 in murine macrophages were also evaluated using the same MTS/PMS assay reaction described above. After washing the peritoneal cavity of the BALB/c mice with Hanks's solution, murine macrophages were obtained and cultivated in a transparent 96-well plate with RPMI medium supplemented with 10% FBS and maintained at 37\u00a0\u00b0C in 5% CO2. After 24\u00a0h of cultivation, SBCs 37\u201343 were added at different concentrations. Macrophage viability was measured at 24, 48, and 72\u00a0h of treatment. MTS/PMS assay reaction was also quantified by optical density measurement at 490\u00a0nm in a microplate reader and SpectraMax M2/M2espectrofluorometer . The cytotoxicity concentration to reduce 50% of viable macrophages (CC50) was determined.For primary screening of the antileishmanial effects of the SBCs 37\u201343, we evaluated the cell viability and cytotoxicity effects in 6 cells/ml. After 24\u00a0h of growth, SBCs 37\u201343 were added at different concentrations from concentrated stock solutions, and cell densities were evaluated daily over 96\u00a0h of growth using a Neubauer chamber. Based on the analysis of CC50 and IC50 in promastigotes, and the cytotoxic effects in murine macrophages, three benzyl farnesyl amine mimetics were chosen for evaluation against intracellular amastigotes. To evaluate the effects of compounds on L. amazonensis intracellular amastigotes, macrophages were infected as described previously and incubated with different concentrations of compounds after 24\u00a0h of infection. Fresh medium was added daily until 3\u00a0days . After this time, cultures were fixed in Bouin\u2019s solution17, washed with 70% ethanol, distilled water, and then stained with Giemsa solution for 1\u00a0h. The number of intracellular amastigotes was obtained after the count in light microscopy. Association indexes /total number of macrophages) were determined and used as a parameter to calculate the percentage of infection for each condition used in this study. The concentration that inhibited 50% of growth (IC50) and selective index (SI) was calculated. The CC50 and IC50 were calculated by fitting the values to a non-linear curve analysis. The regression analyses were performed with SigmaPlot 10 software.After evaluating the cell viability and cytotoxic effects by MTS assay in promastigote forms, we also analyzed the effects of the SBCs 37\u201343 in the growth of promastigotes. For this, promastigote cultures were initiated at a cell density of 1.0\u2009\u00d7\u200910m) of the untreated and treated promastigotes was analyzed using the JC-1 fluorochrome . This lipophilic and cationic mitochondrial vital dye accumulates in the mitochondria in response to \u0394\u03c8m since its fluorescence indicates an energized mitochondrial state15. JC-1 exists as a J-monomer that, in the absence of \u0394\u03c8m, accumulates in low concentration with emission wavelength at 530\u00a0nm (green fluorescence); however, in the presence of \u0394\u03c8m, JC-1 accumulates as J-aggregates with emission at 590\u00a0nm (red fluorescence). Parasites were prepared as previously described16. For each sample, 1\u2009\u00d7\u2009107 parasites were incubated with 10\u00a0\u03bcg/ml JC-1 for 25\u00a0min, with readings made every minute using a microplate reader and spectrofluorometer SpectraMax M2/M2e . As a positive control for the mitochondrial membrane depolarization, cells were incubated with 2\u00a0\u03bcM FCCP during the initial 20\u00a0min of reading. After 20\u00a0min, 2\u00a0\u03bcM FCCP was added to all samples to abolish the \u0394\u03c8m. The relative \u0394\u03c8m values were obtained, calculating the ratio between the reading at 590\u00a0nm and 530\u00a0nm (590:530\u00a0nm). Experiments were independently repeated at least three times in triplicate, and the graphic shows the mean and standard deviation of one representative experiment.Mitochondrial transmembrane electric potential , and incubated with 10\u00a0\u00b5g/ml H2DCFDA in PBS for 1\u00a0h at 25\u00a0\u00b0C. After 1\u00a0h, cells were washed and resuspended in PBS, added to a black 96-well plate, and then analyzed in a microplate reader and spectrofluorometer SpectraMax M2/M2e , using the pair of 507\u00a0nm and 530\u00a0nm wavelengths as emission and excitation wavelengths, respectively.As described previously, intracellular ROS levels were evaluated in control and compound-treated promastigotes7 cells were harvested, washed in PBS (pH 7.2), and incubated with 10\u00a0\u00b5g/ml Nile Red for 20\u00a0min. After this step, cells were washed twice in PBS, resuspended in 200\u00a0\u00b5l of PBS, and then added to a black 96-well plate. Readings were taken in a microplate reader and spectrofluorometer SpectraMax M2/M2e , using the wavelengths 485 and 538\u00a0nm for excitation and emission, respectively.For analysis of lipid bodies accumulation, 1.0\u2009\u00d7\u2009104, 1.25% potassium ferrocyanide, and 5\u00a0mM CaCl2 in 0.1\u00a0M cacodylate buffer (pH 7.2) for 30\u00a0min. For scanning electron microscopy, promastigotes were dehydrated in ethanol and critical point-dried in CO2. After that, samples were sputtered with a thin gold layer and observed under a FEI Quanta 250 scanning electron microscope. For transmission electron microscopy, cells were dehydrated in acetone and embedded in epoxy resin. After that, ultrathin sections were stained with uranyl acetate and lead citrate and observed under a Zeiss 900 electron microscope.First, control and treated promastigotes and intracellular amastigotes were fixed in 2.5% glutaraldehyde in 0.1\u00a0M cacodylate buffer (pH 7.2) for 1\u00a0h at room temperature. Second, the samples were postfixed in a solution containing 1% OsO27. Furthermore, tomogram generation by R-weighted back-projection was performed using ETOMO, and virtual slices were manually segmented using 3DMOD, which was also used to produce 3D models.For electron tomography, ribbons of 200\u00a0nm thick serial sections were obtained by ultrathin sectioning in an ultramicrotome. These ribbons were collected in formvar-coated copper slot grids. After that, colloidal gold particles (10\u00a0nm) were deposited on both surfaces of the sections, being used as fiducial markers for the alignment of the tilted views. Single-axis tilt series (\u00b1\u200960\u00b0 with 1\u00b0 increment) were produced from samples using Xplore3D software and a Tecnai-G2 electron microscope operating at 200\u00a0kV. 3D reconstruction was performed using the IMOD software packageLeishmania amazonensis promastigotes to analyze the effects of SBC 39 and SBC 40 on the free sterol composition of the promastigotes, as described previously28. Neutral lipids were analyzed by MS, and mass spectra were obtained by electron ionization (EI) at 70\u00a0eV according to the protocol published previously28. The assignment of structures was based on relative chromatographic behaviors, as well as the characteristic fragmentation patterns in MS, and by comparison of the mass spectra with those available in the National Institute of Standards and Technology (NIST) Research Library located at the NIST Mass Spectrometry Data Center.Total lipids were extracted from control and drug-treated Five calibration standards were prepared from the pure standard of cholesterol,\u00a0ergosterol,\u00a0\u03b2-sitosterol, stigmasterol, and 5\u03b1-ergosta-8(14)-en-3\u03b2-ol purchased from Sigma-Aldrich Co. Different calibration solutions were prepared using ethyl acetate as solvent. To quantify cholesterol and ergosterol, standards were used at different concentrations of 0.08, 0.10, 0.25, 0.50, and 1.0\u00a0mM to plot the standard curve. From each calibration solution, 1 \u03bcl was injected (run in triplicate) into the GC\u2013MS system to achieve the regression plot of various concentrations versus their peak area. To estimate the level of endogenous sterol/cell in control and drug-treated cultures, the total areas of the corresponding chromatographic peaks were divided by the cell densities of the cultures.P was\u2009<\u20090.05(*),\u2009<\u20090.01(**), and\u2009<\u20090.001(***).All the graphics in the figures were created using the means of three independent experiments, and the bars represent the standard deviations of the means. The statistical significance of differences among the groups was assessed using the one-way or two-way analysis of variance (ANOVA) test, followed by Bonferroni\u2019s multiple-comparison test in the GraphPad Prism 5 software. Results were considered statistically significant when Leishmania parasites were approved by the Ethics Committee for Animal Experimentation of the Health Sciences Centre, Federal University of Rio de Janeiro (Protocols n. IBCCF 096/097/106), according to the Brazilian federal law . Furthermore, all animals received humane care in compliance with the \u201cPrinciples of Laboratory Animal Care\u201d formulated by the National Society for Medical Research and the \u201cGuide for the Care and Use of Laboratory Animals\u201d prepared by the National Academy of Sciences, USA. The experiment involving animals follows the recommendations described in the ARRIVE guidelines.The experiments using BALB/c mice to isolate macrophages and to maintain Leishmania amazonensis promastigotes. Six of them had CC50 lower than 5\u00a0\u00b5M , indicating an excellent effect against Leishmania amazonensis. After that, these six inhibitors were analyzed against Leishmania amazonensis promastigotes to evaluate their potential antiproliferative effects. Figure\u00a050 values in the nanomolar range after 72\u00a0h of treatment .Figure\u00a050 values of 33.94\u00a0\u00b5M, 40.53\u00a0\u00b5M, 40.65\u00a0\u00b5M, and 39.15\u00a0\u00b5M, respectively. Based on these results obtained for macrophages and promastigotes, we decided to evaluate the effects of three of them against intracellular amastigotes. Figure\u00a050 values found to them were in the nanomolar range after 48\u00a0h of treatment or SBC 40 (30\u00a0nM\u2013300\u00a0nM) for 48\u00a0h induced reduction of the relative level of the main endogenous sterol 5-dehydroepisterol in about 50%. However, promastigotes in the presence of SBC 39 displayed a significant accumulation of ergosta-8, 24(24\u2032)-dien-3\u03b2-ol (fecosterol) to 40%, while in the presence of the highest concentration of SBC 40, fecosterol reached only 16% increased as the concentration of SBC 40 increased, clearly showing the effect on the synthesis of sterols . SQS is responsible for the reaction that catalyzes the first committed step in the SB pathway, thus, not interfering with isoprenoid production and its metabolites36. Several squalene synthase (SQS) inhibitors, such as quinuclidine derivatives, have been studied as potent SB inhibitors. The first quinuclidine derivatives tested against Leishmania sp. were BPQ-OH, ER-119884, and E5700, inducing cell death associated with the depletion of the parasite\u2019s endogenous sterols31. Furthermore, another quinuclidine derivative, WSP 1267, showed a potent effect against Candida albicans, C. parapsilosis, and C. tropicalis, with a MIC50 of 2\u00a0\u00b5g/ml37. Some quinuclidine derivatives were also able to inhibit the recombinant L. major SQS at submicromolar concentrations, exhibiting selectivity action for the parasite enzyme21. Benzyl farnesyl amine mimics were also reported to be selective inhibitors of human SQS. They have been explored for their potential use in developing cholesterol-lowering treatment options for hypercholesterolemia in men38.Sterol biosynthesis (SB) is an essential metabolic pathway in Leishmania amazonensis. The compounds SBC 39 and SBC 40 showed the most pronounced effects on the growth of L. amazonensis intracellular amastigotes, associated with low cytotoxicity in mammal cells and a higher selectivity index of 117.7 and 180, respectively. These selectivity indexes are higher than those found after treatment with posaconazole and itraconazole, two potent inhibitors of the growth of L. amazonensis16. Herein, the antiproliferative activities of SBC 39 and SBC 40 were in the nanomolar range against both extracellular promastigotes and intracellular amastigotes. Furthermore, the biological activity against Leishmania amazonensis was lower than those found for E5700 and ER-119884, two SQS inhibitors from Eisai Pharmaceutical Company, previously studied against Leishmania26. Nevertheless, it is noteworthy to mention that further development of E5700 and ER 119884 has been stopped because it caused testicle atrophy in a small animal experiment . Beyond this, quinuclidine derivatives often bear a certain risk of neurological side effects . Therefore, medicinal chemistry is increasingly interested in avoiding the quinuclidine moiety in early phase drug development. Finally, benzyl farnesyl amine mimetics have a much lower production cost, which is especially highly attractive for anti-infectious drug design in tropical emerging countries, where cost restrictions sometimes limit pharmaceutical development. Thus, these results should be regarded as a precious contribution to SB inhibitors research in tropical parasites with a high potential for further drug development.In this study, we report the activity of several benzyl farnesyl amine mimetics against 28. Transmission electron microscopy images indicated that mitochondrion ultrastructure was dramatically altered after treatment with SBC 39 and SBC 40 -dien-3\u03b2-ol (III) (fecosterol) in cells treated with both SBC 39 and SBC 40 indicated that these inhibitors not only impair the activity of the \u22068\u2009\u2192\u2009\u22067 double-bond isomerization (C-8 sterol isomerase/ERG2), but also the formation of a double bond between C-5 and C-6 in the B ring of sterols (C-5 sterol desaturase/ERG3).Using GC\u2013MS, we documented the relative composition (%) of eleven sterols in the membranes of es Table . In our 23. For instance, the ratio of cholesterol content to endogenous sterols in promastigotes control was 1:43. In contrast, in promastigotes treated with SBC 40 (at 300\u00a0nM), the cholesterol to endogenous sterols ratio changed dramatically to 1:7. The result is consistent with a blockade of de novo sterol synthesis at the level of SQS.Furthermore, the results Tables , 2 indicLeishmania amazonensis, and remarkably, they both have a dual mode of action (Fig.\u00a0Thus, the results suggest that both SBC 39 and SBC 40 have potent antiproliferative and selective growth inhibition effects on ion Fig.\u00a0. The firLeishmania sp. since they presented a very high specificity for the parasite. Furthermore, our findings justify future studies to better understand the mode of action and use in combination therapy with other SB inhibitors as a new therapeutic strategy that could reduce toxicity but increase the efficacy of treatment.In conclusion, our results support the notion that SBC 39 and SBC 40 are promising new chemotherapeutic agents against Supplementary Information 1.Supplementary Video 1.Supplementary Video 2.Supplementary Video 3."} +{"text": "Nitroepoxides were introduced as efficient substrates for the one-pot three-component synthesis of 2-iminothiazoles under catalyst-free conditions. Reaction of amines, isothiocyanates, and nitroepoxides in THF at 10\u201315\u00a0\u00b0C afforded corresponding 2-iminothiazoles in high to excellent yields. The reaction proceeds via the in situ formation of thiourea from an amine and an isothiocyanate, followed by nitroepoxide ring opening with the sulfur of thiourea, cyclization reaction, and dehydration cascade. The structures of products were confirmed by IR, NMR, HRMS analyses and X-ray crystallography. Particularly, the 2-imino-1,3-thiazolines are available in many biologically active compounds with antimicrobial, anticancer, anti-inflammatory, antihistaminic, antihypertensive, hypnotic, and anticonvulsant activities13. In addition, they were applied for the identification of human cells with positive myeloperoxidase reactivity14. For example 4-methyl-3-H-thiazoline derivative, PS-028, is a potent and selective GPIIb/IIIa receptor antagonist ethanolamine26, amidines27, diamines28, S-alkyl dithiocarbamates23 and etc.31 have been utilized in the reaction with nitroepoxides for the synthesis of valuable heterocyclic compounds including imidazoles, 1,3-thiazoles, benzodiazepines, thiazoline-2-thiones, and pyrazines.Nitroepoxides, which can be simply prepared from nitroalkenes by epoxidation reaction with H38. For this purpose, multicomponent reaction of isothiocyanates with amines in the presence of an electrophile is an efficient strategy for the synthesis of 2-imino-1,3-thiazolines. In this context, Yavari et al. developed a synthetic route for the synthesis of functionalized 2-imino-1,3-thiazoles using tetramethylguanidine, isothiocyanates, and 2-chloro-1,3-dicarbonyls 3 aziridines were also developed for the synthesis of 2-imino-1,3-thiazolines47.Multicomponent reactions (MCRs) provide simple access to complex molecules in a one-pot. MCRs have several advantages such as generation of several bonds in a single operation, avoiding the need for isolation and purification of reaction intermediates, saving in time, materials, solvents and energy1H and 13C NMR spectra of products were recorded on a Bruker AMX 300\u00a0MHz spectrometers referenced to internal Me4Si at 0.00\u00a0ppm. Reaction monitoring was carried out by thin-layer chromatography using TLC silica gel 60 F254 plates. HRMS (High Resolution Mass Spectra) was measured on a THERMO SCIENTIFIC Advantage and a THERMO SCIENTIFIC Exactive instrument equipped with an APCI source in the positive-ion mode. Mass analysis was performed using Agilent Technology (HP); 5973 Network Mass Selective Detector equipped with EI mode and ionization energy of 70\u00a0eV. The temperatures of the electronic-impact ion source and MS quadrupole were 230\u00a0\u00b0C. IR spectra were recorded on a Perkin-Elmer Spectrum RXI FT-IR Spectrometer. Nitroepoxides were prepared according to the literature procedures48.All chemicals and solvents were obtained from commercial sources and used as received. The n-hexane/EtOAc (7/3) . The structures of products were confirmed by IR, 1H NMR, 13C NMR, Mass/HRMS analyses and X-ray crystallography. Detailed information is available in the \u201cSupporting Information\u201d file.In a test tube equipped with magnetic stirrer bar, an isothiocyanate , an amine (1.2 equiv) and THF (4\u00a0mL) were mixed for 1\u00a0h at room temperature. Progress of the reaction was monitored by TLC. After complete consumption of the isothiocyanate, the reaction temperature was decreased to 10\u201315\u00a0\u00b0C and a nitroepoxide (1 equiv) was added and the mixture was stirred at 10\u201315\u00a0\u00b0C for 6\u00a0h. The solvent was removed under reduced pressure to afford yellow viscous oil. Purification was carried out by recrystallization in a minimum amount of MeOH or by column chromatography using silica gel and N,S-heterocycles and the chemistry of nitroepoxides52, herein we report a novel catalyst-free one-pot three-component route for the synthesis of 2-imino-1,3-thiazolines via nitroepoxide ring opening reaction and further stirring for 6\u00a0h at room temperature afforded a trace amount of corresponding product 4a . In addition, the nature and position of the substituents on the phenyl ring of nitroepoxides did not have significant impact on the reaction yield. High to excellent yields were obtained with both electron-donating and -withdrawing substituents on the phenyl ring. The structures of products were confirmed using IR, 1H and 13C NMR, and HRMS analyses. Characteristic signal in 1H NMR spectra is related to the methyl group which appeared around 2.2\u00a0ppm in all products. In addition, a signal at 158\u00a0ppm in 13C NMR was assigned for the carbon of the imine moiety in the products. In principle, two regioisomers may be generated in the reaction of in situ prepared thiourea with nitroepoxide, which is difficult to differentiate by 1H and 13C NMR analyses. For this purpose, to confirm the proposed structure for the products, the structure of 4p was elucidated using single-crystal X-ray analysis. ORTEP representations of 4p is shown in Fig.\u00a0Z, in which the substituent on the nitrogen of imine is in the same side with sulfur atom in the ring.With optimized reaction conditions in hand, the generality and scope of this protocol was examined using various amines, isothiocyanates, and nitroepoxides and the results are summarized in Table 2 with isobutyl amine was performed in THF for 1\u00a0h and the corresponding thiourea 5 was separated in 95% yield and characterized by 1H NMR analysis (See supporting information for 1H NMR spectra) (Scheme 5 with nitroepoxide was carried out and the corresponding 2-iminothiazole 4e was obtained in 90% isolated yield (Scheme 41, a proposed mechanism for this reaction is depicted in Scheme N,N\u2019-disubstituted thiourea A, which undergoes nucleophilic addition to nitroepoxide by sulfur atom to produce intermediate B. Elimination of nitrous acid from B affords the intermediate C. Then, intramolecular cyclization via the addition of aliphatic nitrogen to the carbonyl group in intermediate C provides the intermediate D, which undergoes dehydration reaction to afford the final structure 4. The nitrogen of aliphatic amine is superior to the nitrogen of aromatic amine for cyclization reaction due to the higher electron density and nucleophilic character.To propose a reasonable mechanism, two control experiments were carried out. Reaction of phenylisothiocyanate In conclusion, we have developed an efficient catalyst-free one-pot three-component procedure for the synthesis of 2-imino-1,3-thiazoles in high to excellent yields. The reaction proceeds via the ring-opening of nitroepoxides with in situ prepared thiourea derivatives. By this protocol, it is possible to prepare diversities of 2-imino-1,3-thiazolines with various substitution pattern. The main advantages of this protocol include catalyst-free conditions, simple reaction conditions, high to excellent yields and convenient operations without extra purification.Supplementary Information."} +{"text": "Despite the media are often described as critical for the success of the well-being agenda, there is wide dissatisfaction with their current level of interest. However, the media coverage of well-being metrics has been unresearched and, even when studies have been conducted, these employed unrobust methodologies, were limited to newspapers and to restricted samples of metrics. This paper fills such gap, providing also for the first time an analysis of radio and TV coverage of well-being metrics. The research was undertaken using Factiva (for newspapers) and TVEyes (for radio and TV) for the years of 2017\u20132021 and 2018\u20132021, respectively. The countries analysed are Scotland and Italy, both pioneers in the measurement of well-being. Findings reveal that media coverage of well-being metrics has been extremely low overall and that this was impacted negatively by the COVID-19 pandemic, which instead impacted positively on the reporting of GDP and related queries, showing that the main concern during the pandemic was the impact that this was going to have in terms of output, rather than in terms of well-being. Most composite indices, whose creation is often thought to help obtain greater media coverage, were almost if not even fully ignored by journalists, whereas metrics that lack an overall composite index but that are overseen by independent institutions and have been institutionalised were among the ones that were reported the most. According to the European Commission , p. 2, i., The success of these metrics was claimed in many studies to depend on their level of media coverage. For POINT (Policy Influence of Indicators) workshop participants, for instance, success for well-being metrics meant primarily to be \u2018quoted, mentioned and reported on an ongoing basis\u2019 , its inclusion clearly distorted his findings. Second, Morse (Human Development Report) or in fact any other alternative names (e.g. Index of Human Development). Therefore, his figures largely underestimate the number of articles published that cited the indices in question.However, the methodology employed by Morse was inconsistent, not transparent and incomplete. First, in two of his studies , 2013 he the UK\u2019 , p. 23. d, Morse , p. 1682The Guardian, for instance, is covered from July 14, 1984, whereas the Italian Corriere della Sera from January 27, 2009, although both were founded much earlier. This means that during the period he analysed , a similar study was undertaken to investigate the media coverage of 19 metrics in several English and French newspapers from 1990 to 2010. Although the overall number of mentions and that for each metric were not shown, the study found the Ecological Footprint to be the most mentioned one, followed by the HDI and to restricted samples of metrics. A rigorous analysis of media coverage of well-being metrics is therefore lacking. By using Factiva for the analysis of newspaper coverage and TVEyes for that of radio and TV, I was able to fill this research gap. Factiva enables us to search over 45,000 sources . TVEyes is a media monitoring tool that uses speech recognition technology to provide real-time transcripts of whatever is broadcast on radio and TV.The list of metrics that I searched for can be found below Table . All metThe WHR is the metric that was found in the highest number of articles (300), followed by the HDI 187) and the WWF LPI (116) with the BCI, the EURSPI, the GIP and the ISEW at the end of the scale , for a total of 1,015 articles . For instance, below are articles sorted by month for the WHR and the Bank of Scotland QoLS, which 40% of all articles refer to Fig.\u00a0. The majThe Daily Telegraph, The Times, Financial Times, The Guardian and The Independent of all articles were published in just five newspapers, namely ent Fig.\u00a0. No artiThe number of articles mentioning all 23 well-being metrics combined is 1.6% compared to the number of articles that mentioned at least once GDP over the same period, 2.7% compared to economic growth and 2% compared to recession Fig.\u00a0. CompareThe most mentioned metric over the 38-month period analysed was the WHR with 85 references, followed by the WWF LPI (48) and the Bank of Scotland QoLS (45), with 12 metrics at the end of the scale such as the BLI and the ISEW , for a total of 281 mentions Fig.\u00a0.Fig. 6NuOverall, the number of mentions decreased by 16% in 2020 compared to 2019 Fig.\u00a0. All metThe majority of mentions (54%) were made in just four months: November 2018, January and March 2019, and September 2020. These correspond to the months in which the 2018 GGCI, the 2019 Bank of Scotland QoLS, the 2019 WHR and the 2020 WWF LPI were issued, respectively. In fact, well-being metrics were mentioned again mostly, if not even only, in the months in which they got released. This trend can be seen clearly if we break down data by month for the top two metrics, which 47% of all mentions refer to , followed by the BES (713) and the ItaliaOggi QoLR (418), with the BCI, the IWI and the PIQ at the end of the scale , for a total of 2,685 articles Fig.\u00a0.Fig. 11NThe overall number of articles increased in 2018, decreased slightly the year after and more significantly by 28% in 2020 Fig.\u00a0. Since oSole 24 ORE issued a Youth Quality of Life sub-index. The issuing of metrics and their reporting is so interlinked that in December 2020, when the 2020 BES was supposed to come out before being postponed to 2021, only a handful of articles mentioned it. In other words, not only did the BES get reported upon its release, but it also did not when no BES Report was issued. Note how the institutionalisation of the BES, and the consequent publication of yearly MEF BES Reports and Appendices, has granted it frequent coverage throughout the year compared to other metrics. In fact, of all metrics studied both in Italy and Scotland, the BES is the only one that was reported on a regular basis, so much so that between 2017 and 2021 it was mentioned in at least one article in every but one month, not to mention that in 2021 the MEF BES Report received greater coverage than ISTAT\u2019s.Like in Scotland, the majority of articles were published in the months in which metrics were released and, within those months, almost exclusively on the day of their release. For instance, below is the number of articles sorted by month for the top two metrics, which 62% of all articles refer to, for the years of 2020 and 2021 Fig.\u00a0. The majSole 24 ORE, ItaliaOggi and Avvenire (responsible for their homonymous rankings) almost never talked about their competitors\u2019 metrics. Yet, they are among the top five newspapers in terms of references to the BES, despite this being a competitor. Interestingly, of all articles published by the first seven newspapers , only 6% are about the WHR and 3% about the GNH. The most read newspapers in the country thus did not seem interested in metrics that explicitly emphasise subjective indicators.Half of all articles appeared in seven newspapers Fig.\u00a0. Unlike The number of articles mentioning all 21 well-being metrics combined is 2.8% compared to the number of articles that mentioned at least once GDP over the same period, 14% compared to economic growth and 11% to recession Fig.\u00a0. CompareThe most mentioned metric over the 38-month period analysed was the Sole 24 ORE QoLR (with 762 references), followed by the BES (645) and the ItaliaOggi QoLR (375), with 9 metrics such LPI and the PIQ at the end of the scale , for a total of 2,256 references Fig.\u00a0.Fig. 16NOverall references increased particularly in November and December 2018, March and December 2019, December 2020, and March and December 2021 Fig.\u00a0. Such peThe majority of references 68%) were made in the months in which metrics were issued (except for those that are not updated regularly). Above, for instance, are references to the top four metrics sorted by month Fig.\u00a0. All the% were maThe majority of references within the above months were made on the days these metrics were issued Fig.\u00a0. Above, RTL 102.5, Radio Capital, RMC, Radio 105, R101, RDS and Radio Deejay). As for TV, the trend is similar, in the sense that the top five sources, which account for 65% of all TV mentions, are all-news channels. Breaking down data further by type of outlet, metric and source (not shown here) reveals again an interesting competition between Class CNBC and Radio 24 , with both almost completely ignoring each other\u2019s ranking. This shows the difference between commercial, newspaper rankings and official, institutionalised metrics such as the BES. In fact, despite the above and the BES being again a competitor, both were among the sources on which the BES was mentioned the most. Another finding worth noting is that metrics that more explicitly emphasise subjective indicators such as the GNH and the WHR were not mentioned frequently on \u201cnews and talk\u201d or less commercial sources. Indeed, of all mentions made on the first five radio stations and TV channels , less than 5% were about the WHR and only 1% about the GNH. This suggests that these metrics are not being taken seriously by the journalists working for these sources or by the policymakers interviewed by them.Slightly more than half of all references (54%) were made on TV Fig.\u00a0. As for As for the time of the day in which mentions were made, similarly to Scotland there were three peaks at 7 A.M., 12 P.M. and 5 P.M. Fig.\u00a0. Also siFinally, above is the number of references to all 21 metrics combined compared to that of GDP, economic growth and recession Fig.\u00a0. Data sodo get mentioned regularly and with a striking frequency.Findings revealed almost the exact same reporting patterns in both countries. Well-being metrics were mentioned occasionally, particularly during weekends in Scotland and on Mondays in Italy and mainly upon their publication in line with Morse , 2013. Swith that of just one. This also confirms claims made in the literature about the overexposure of GDP , the periods in which these got covered , but above all their overall level of coverage. In the case of newspapers, 1,015 articles were published on Scottish and UK-wide sources whereas 2,685 in Italy . In the case of radio and TV, 281 mentions were made on Scottish and UK-wide sources, whereas 2,256 in Italy, despite the Italian sample including eight fewer sources. Italy\u2019s much higher coverage, combined with the higher number of newspapers that showed interest in well-being metrics and Italy\u2019s longer and more established history of well-being measurement show, from a comparative perspective, that Italy is more advanced than Scotland as far as the promotion of the well-being agenda is concerned. Conversely, Scotland\u2019s coverage levels show that Heins and Pautz , p. 100 Sole 24 ORE and Radio 24, and ItaliaOggi and Class CNBC despite these not talking about each other\u2019s ranking. This seems to apply globally, too: it does not seem a coincidence that the WHR and the HDI, both United Nations outputs, have gained much more traction than the SPI, the HPI, the LPI and so on.Surprisingly, most composite indices were almost if not even fully ignored by journalists, whereas those metrics that lack an overall composite index such as the BES were among the ones that were picked up the most. It is often believed . Yet despite the higher number of years and the lower number of governments, the continued lack of attention towards the promotion of the NPF from the SNP-led government, whom the framework effectively belongs to, has determined its continued absence from the media landscape.11as a whole will have been in a way useless, as almost all the indicators included in the NPF already existed before its creation and the point of having them together is exactly to highlight the interconnectedness of different policy areas. The Scottish First Minister herself noted that, pointing out that \u2018the\u00a0National Performance Framework\u00a0is intended to be a cross-cutting framework and it is important not to see anything that we capture in isolation\u2019 . Regarding the first issue, this cannot be solved unless significant funding is available to cover the large amount of time needed to go through each article. Regarding the second issue, if articles about GDP were largely critical then during the COVID-19 pandemic and the economic crisis that followed those mentioning well-being metrics should have increased and those mentioning GDP and related terms decreased, whereas the opposite happened. In a way, then, the COVID-19 pandemic This article provided an in-depth investigation of media coverage of well-being metrics in Scotland and Italy on which no data were previously available. In fact, to the best of my knowledge no such data have ever been made available for any other country despite the key role that the media are believed to play for the success of the well-being agenda. It is true that studies were conducted about newspaper coverage (Bassi, et al., that is the problem. Second, expectations around composite indices should be reconsidered and more effort should be put into developing official, national and independent frameworks. Due indeed to their limited national relevance, global metrics generate mediocre to no coverage at all and in any case only upon their release, showing the ability of their promoters to craft good press releases rather than the capacity to spark a debate throughout the year which only the institutionalisation of an official, national, independent framework like the BES has shown to be capable of. Third, press releases should be sent considering the day and time journalists are more receptive and when the news cycle is more propitious. Data from both countries show that well-being metrics were mentioned particularly in the early morning, at lunchtime and in the afternoon, but not much during prime time. They also show that, as far as radio and TV are concerned, they get reported especially during weekends in Scotland and on Mondays in Italy, due to there being a sort of news vacuum in these days. Until well-being metrics gain currency and saliency, exploiting these days and times is going to be the most impactful strategy.What, then, could increase the media coverage of well-being metrics? First, more effort should be put into promoting the well-being agenda when the economy is expanding as opposed to periods in which growth is lacking. This is because in such cases the attention will predominantly be on how to put the economy back on a growth track because"} +{"text": "Micro electro-mechanical systems (MEMS) are used to record training and match play of intermittent team sport athletes. Paired with estimates of internal responses or adaptations to exercise, practitioners gain insight into players\u2019 dose\u2013response relationship which facilitates the prescription of the training stimuli to optimize performance, prevent injuries, and to guide rehabilitation processes. A systematic review on the relationship between external, wearable-based, and internal parameters in team sport athletes, compliant with the PRISMA guidelines, was conducted. The literature research was performed from earliest record to 1 September 2020 using the databases PubMed, Web of Science, CINAHL, and SportDISCUS. A total of 66 full-text articles were reviewed encompassing 1541 athletes. About 109 different relationships between variables have been reviewed. The most investigated relationship across sports was found between (session) rating of perceived exertion ((session-)RPE) and PlayerLoad\u2122 (PL) with, predominantly, moderate to strong associations (r = 0.49\u20130.84). Relationships between internal parameters and highly dynamic, anaerobic movements were heterogenous. Relationships between average heart rate (HR), Edward\u2019s and Banister\u2019s training impulse (TRIMP) seem to be reflected in parameters of overall activity such as PL and TD for running-intensive team sports. PL may further be suitable to estimate the overall subjective perception. To identify high fine-structured loading\u2014relative to a certain type of sport\u2014more specific measures and devices are needed. Individualization of parameters could be helpful to enhance practicality. Player monitoring in sports aims at optimizing training adaptations to improve performance and reduce injury risk . AdaptatInternal loading due to sports activities primarily results from movement-related force application demands. Forces need to be applied to the environment to cover running distances, perform changes in movement direction or accelerate or decelerate the body, e.g., during acceleration, stopping, or jumping tasks. Applying forces to the environment results in reaction forces acting on athletes\u2019 bodies, determining the external load stimulus applied to the biological system. Physical external loads applied over time result in different types of internal loads , which determine the body\u2019s adaptations. Knowledge of the internal response and adaptation to a given dose of external load is crucial for optimal, injury-free training progress. The internal load is influenced by individual factors such as age, gender, training experience, health status, and nutrition . The lin2max) tests using ergometry). Fourth, the assessment of the current health and fitness status, which, among other parameters such as genetics, age, and gender, make up the individual characteristics. Parameters of each category are included in this systematic review if a relationship to an external load parameter, measured during training or match play using a MEMS device, was assessed.In this framework, it can be distinguished between four different categories of internal parameter assessments: First, the internal load estimates collected during exercise, primarily made up of HR-based indices and RPE or session-RPE. Second, the exercise-induced responses measured post-exercise due to the delayed response of specific systems to activity, such as creatine kinase (CK), an indicator of muscle damage. Third, the body\u2019s adaptations may be assessed over time using force platforms or the inverse dynamics-based calculation of external joint moments), field settings offer greater ecological validity and the potential to reach larger numbers of athletes.In the field setting, external load variables can be measured using lightweight, body-worn sensors. With the introduction of global navigation satellite systems (GNSS) devices into the player monitoring market, the research around workload quantification and load monitoring has increased exponentially in the last 15\u201320 years ,15. NextHowever, keeping track of loading in team sports is a complex task: Running-based team sports are intermittent sports, consisting of hundreds of brief and very intense actions, such as jumps, tackles, changes of directions, accelerations, and decelerations . These mConsequently, sports scientists and tracking device manufacturers have created several parameters such as \u201cPlayerLoad\u2122\u201d (PL), \u201cimpact load\u201d, or \u201cleg stiffness\u201d, intending to capture load characteristics and their changes with, e.g., fatigue or training status. One of the main challenges in developing load parameters is to capture the demands of accelerating and decelerating, as well as turns and tackles. \u201cMetabolic power\u201d, for example, is one more recently developed approach that attempts to capture the demands of accelerating based on the assumption that this is comparable to the metabolic demands of running uphill . NeverthNew possibilities have been created using MEMS to quantify loading in team sports athletes. Nevertheless, a consensus on quantifying the \u201cinternal\u201d load of team sport athletes by \u201cexternal\u201d locomotor measurements is still missing ,21,22,23Consequently, two main challenges regarding load monitoring in team sport athletes have been identified: First, the complex task of quantifying the complex loading situation of intermittent team sports, and second, the difficulty of knowing the relationship between the given MEMS-based external load and the athlete\u2019s individual internal loading and consequently exercise-responses and adaptations within different domains. Identifying these relationships offers great potential to improve the understanding of individual load-response profiles.Therefore, this systematic review addresses MEMS-based external load parameters and their relationship to various internal parameters, encompassing biochemical, neuromuscular, subjective, cardiovascular, and further domains. This work could aid practitioners in choosing and interpreting appropriate parameters to monitor load in a time- and cost-effective manner to provide the appropriate stimuli to induce adaptations to improve sports performance and decrease the risk of injury.A systematic literature review was conducted based on the preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines . The folAfter the final selection was made, the quality of the selected studies was assessed using a 16-item checklist developed by Law et al. and modiData extraction was done using a custom-made sheet pilot tested on five randomly chosen articles. The sheet was redefined, and its final version was used by one reviewer (JH) who performed the data extraction. In case of unclear or missing data, corresponding authors were contacted. The following data were extracted from the studies: (1.) The type of team sport; (2.) the study sample ; (3.) the external parameters recorded or calculated; (4.) the internal parameters measured or calculated and/or the fitness assessment; (5.) the relationship between the external and internal parameters as indicated by statistical association or predictive measures.Data were categorized into groups consisting of the different team sports. Then, subgroups according to the parameters analyzed were created. The subgroups are based on A descriptive synthesis was undertaken with the data structured in a table containing the team sport, the studies included, the load parameters collected, including their frequency of use per sport, and the statistical relational measures between the external and internal parameters. The overall frequency of use of each external and each internal parameter was visualized using pie charts.n = 2178) that did not meet the criteria, 66 articles remained for the final analysis (n = 56) qualified as excellent regarding methodological quality. The most common item to lose quality points on was item 5: justification of the study sample size.The initial search returned 3573 articles. A total of 2234 records remained after removing duplicates; these articles were screened by title and abstracts against the eligibility criteria. After further exclusion of studies , Australian football (n = 11), basketball (n = 4), field hockey (n = 1), rugby union and rugby league (n = 8), soccer (n = 35), and tag football (n = 1). The participants were professional (n = 606), elite (n = 413), college/university (n = 402), and semi-professional (n = 120) athletes. n = 62 studies included male participants, totaling 1479 male athletes. Four studies studied female participants, accounting for n = 62 female athletes. The three most commonly external parameters recorded were distances in speed zones (n = 55), total distance (n = 46), and PL (n = 34), as depicted in n = 29), HR-based indices (n = 19), and well-being questionnaires (n = 17), as depicted in The articles included in this systematic review ranged from 2011 to 2019. The sports analyzed were: American football (n = 55) which was investigated in 82% of studies included in this review, followed by total distance (n = 46), analyzed in 67% of the research articles in this review, and PL (n = 34), occurring in 51%. (session-)RPE (n = 29) was most often investigated amongst the internal parameters, followed by HR-based indices (n = 19) and well-being questionnaires (n = 17), occurring in 45, 28, and 27% of research articles included in this systematic review, respectively. From the 66 articles included, 109 different relationships between external and internal parameters have been extracted. The most frequently analyzed relationship was between (session-)RPE and PL with predominantly moderate to strong associations (r = 0.49\u20130.84). The second most frequently analyzed relationship was between (session-)RPE and distances in speed zones with heterogeneous results. All results for the 109 relationships can be found in About 34 external and 32 internal parameters and parameter groups were included across all studies. Different HR-based and various (session-)RPE parameters were grouped and displayed in This systematic review aimed to enhance the knowledge around relationships between external, wearable-based load parameters and internal load, exercise-induced responses, adaptation parameters, and parameters of individual characteristics in running-based team sports. Knowledge about these relationships may reduce time- and possibly cost-intensive testing outside regular training. Acute fitness and fatigue states may be drawn only based on external load parameters. Additionally, the amount of data to be collected and analyzed could be reduced by collecting fewer internal parameters.Our systematic review is the first to include a myriad of external and internal parameters, focusing on external parameters collected from wearables only. This is crucial to enhance practicality and usability of parameters collected on-field. As the amount of data from wearable sensors and their use increase, it is inevitable to enhance the knowledge around these parameters and understand the dose\u2013response relationship of team sport athletes. The findings are discussed in the following sections.As some relationships have been examined by a minimal number of studies, results are discussed only when a systematic synthesis of results is feasible. In the following, results are discussed in categories of internal load, exercise-induced response, adaptation parameters, and individual characteristics . Figure Internal load parameters predominantly encompassed subjective ratings of exertion and HR-based indices.(session-)RPE had moderate to strong associations with total and relative distance in Australian football ,43 and sThe weak relationship between (session-)RPE and PL in American football may be due to different match demands compared to other team sports analyzed. American football players generally cover lower overall distances in games than soHeterogenous results between (session-)RPE and distances in speed zones for soccer players may be due to the larger volume of studies compared to other team sports included and due to the different methods used: partial correlations , within-Generally, indicators of total volume seem to result in higher associations than those expressed per minute or as a percentage of total volume. Rago et al. (2019) found a tendency of increasing correlations when speed thresholds were individualized rather than identical for all players.Heterogenous results between (session-)RPE and parameters describing accelerations and decelerations in soccer may be due to varying methods. Using partial and within-individual correlations , small tCorrelations between (session-)RPE and acceleration and deceleration parameters may be higher in basketball due to the high frequency of accelerations of 29.6 \u00b1 3.9\u201332.7 \u00b1 11.0 per minute in professional male players comparedFor practitioners, this means that estimation of (session-)RPE may be done most adequately with indicators of total volume such as total distance or PL in Australian football, soccer, and basketball players. In indoor team sports such as basketball, where total distance is not available from wearable sensors due to a lack of GNSS signal, parameters describing acceleration and deceleration may be used instead of total distance. Omitting (session-)RPE scales would save practitioners and players the time to analyze and fill out the scales.max [max [max [The relationships of HR-based parameters of internal load to external load parameters were analyzed in soccer players only. HR was divided in zones ,80, predmax ,84, and max ,60,72,79max ,101 thatmax . Banistemax , here Edmax ,60. Corrmax [max ,105. Premax [max ,106. ForOverall, among the internal load parameters, (session-)RPE, time spent in low- and medium-intensity HR zones, and TRIMP are best estimated using parameters of total volume such as PL and total distance. Time spent in high-intensity HR zones are not represented adequately by the external load parameters examined and may be recorded separately if of interest. Noteworthy is that HR-based indices do not adequately represent anaerobic training ,106. TheExercise-induced response are short-term changes in parameters. To detect changes, parameters are collected at two or more time points several hours apart. The first time point serves as a baseline measure, usually in a non-fatigued state. The next time point(s) occur(s) following exercise when athletes may be fatigued. Exercise-induced responses extracted from the studies included consist of well-being indicators, CK concentrations, HR-based indices, neuromuscular functioning, and biomarkers, among others.The relationships between well-being parameters and external load indicators were heterogeneous, possibly due to studies using different questionnaires analyzing either overall wellness ,50,68,76Results in Exercise-induced HR-based indices were found only in studies observing soccer players. Here, mostly negligible to small associations between several external load indicators and heart rate variability (HRV) were found ,83,84. FThe relationship between change in countermovement jump (CMJ) parameters as an indicator of neuromuscular fatigue and high-speed running parameters varied from negligible and nonsignificant to large and was assessed in soccer and tag football players. Three studies found negligible to small correlations ,84,86, wCMJ parameters\u2019 relationship to acceleration and deceleration parameters exhibited varying results. One study assessed relationships to parameters of overall volume, such as total distance, duration, and PL, only finding trivial or unclear effects in soccer players . For praIn American football, the number of impacts and peak head accelerations may indicate S100beta levels but not tau concentrations . Here, dConcentrations of secretory immunoglobulin A (sIgA), a marker of immune function, have been linked to high-intensity distance, total distance, and acceleration and deceleration parameters in soccer ,91. SmalOverall, numerous different exercise-induced responses were analyzed, as previously depicted in Adaptation parameters are assessed at a minimum of two time points several days, weeks, or months apart and they are commonly carried out in a non-fatigued state. Changes between measurements can be analyzed and viewed as adaptation. Adaptation parameters extracted from the studies included were related to changes in body mass, HR-based indices, intermittent and aerobic endurance capacities, sleep efficiency, and strength parameters.max was positively related to 10-week sprinting distance [Body mass seems to change in relation to 10-week sprinting distance, and total distance, but not session duration or average speed ,82. Chandistance . ACWR isdistance . Change distance . Monitor2max, improvements in aerobic endurance were strongly correlated to 10-week accumulated PL and acceleration [2max [2max increased despite decreasing duration. In this case, other factors, such as high mechanical loading, seem to represent improvements in aerobic capacity better than training duration.As indicated by a positive change in VOleration . Sessionon [2max . The durChanges in intermittent fitness, as indicated by the 30-15 intermittent fitness test, were observed by one study only, which found unclear and large relationships to high-intensity running, total distance, and PL, respectively . More reOverall, findings suggest that intensity seems particularly important to improve certain physiological capacities related to intermittent team sports. Volume and intensity need to be well-balanced in training programs to cause optimal adaptations.2max regarding total distance and intensity parameters in soccer players [Individual characteristics analyzed in relation to external parameters collected using wearables include intermittent and aerobic endurance capacity, neuromuscular performance parameters, and muscle architecture. As indicated by the Yo-Yo intermittent recovery test (YYIR), players with larger intermittent endurance capacities covered greater total distances in soccer and tag football ,94. Grea players . For praParameters of neuromuscular performance and muscle architecture were analyzed each by one study only; thus, a systematic synthesis is not feasible, and more research is needed to draw conclusions regarding those parameter relationships.Generally, based on the intensity and volume of external load experienced during training and match play, internal bodily reactions take place, which, in the long term, lead to adaptations and influence individual characteristics. Those characteristics determine how well a player can handle the external load. However, no consensus exists on the parameters encompassing internal load yet. Some researchers have a broad understanding of internal load parameters, including biochemical, neuromuscular, and hormonal responses ,2. OtherSome limitations to this systematic review are acknowledged in the following. These include the non-feasibility of synthesizing results for some parameter relationships. Given the wide variety of parameters, some relationships were analyzed by fewer studies to synthesize results systematically. Further, thresholds for speed zones differ across studies such that results may vary due to varying absolute or individualized thresholds used. Different hard- and software was implemented in the studies analyzed, which may cause a discrepancy in results. Manufacturers apply filters to the data during post-processing such that the same parameter could supposedly differ when obtained from another product. Even a software update could result in inconsistent results.2max. This finding, however, likely does not mean shorter training durations cause an increase in aerobic endurance, but rather other circumstances were in place, such as high running volume and repeated high-intensity events, that may elicit improved aerobic endurance.Correlations found do not mean causality; parameters might correlate because of other circumstances. Clemente et al. (2019) found a large negative correlation between training duration and VOn = 4) included in this systematic review studied female athletes [Few studies RPE, and in collision-based team sports, additionally markers of muscle damage may be omitted and replaced by external, on-field measurements, facilitating the work of practitioners. Relationships between external load and the other three internal parameter categories, exercise-induced responses, adaptations, and individual characteristics, are mostly ambiguous and need further verification. Until then, a holistic picture of an athlete may best be obtained by collecting external and internal parameters for those parameter groups. Due to the ever-increasing amount of data collected in both areas, external and internal, a sound understanding of the data and their sport-specific context becomes increasingly important. Good communication is crucial for all stakeholders to attain a common understanding of the data. Studies including female athletes have been noticeably little in number and should be increased in the future. Future work will need to validate novel methods of collecting internal parameters and their relationships to external parameters in order to understand the individual dose-response patterns."} +{"text": "Biological imaging is one of the primary tools by which we understand living systems across scales from atoms to organisms. Rapid advances in imaging technology have increased both the spatial and temporal resolutions at which we examine those systems, as well as enabling visualisation of larger tissue volumes. These advances have huge potential but also generate ever increasing amounts of imaging data that must be stored and analysed. Public image repositories provide a critical scientific service through open data provision, supporting reproducibility of scientific results, access to reference imaging datasets and reuse of data for new scientific discovery and acceleration of image analysis methods development. The scale and scope of imaging data provides both challenges and opportunities for open sharing of image data. In this article, we provide a perspective influenced by decades of provision of open data resources for biological information, suggesting areas to focus on and a path towards global interoperability. Biological imaging has a long history as a key approach for understanding biological systems. From early microscopy with hand-ground lenses through to modern, highly automated microscopes, imaging technology has developed hand in hand with our understanding of life itself. Since the dawn of computational microscopy, improved sensor technology, automation and scaling have enabled ever increasing volumes of image data from many different modalities to be collected.Not only are data volumes increasing rapidly, but deriving scientific results from imaging data increasingly requires complex analysis pipelines that act on raw images to eventually produce information principles and the BioImage Archive together providing free public archival of imaging data from any modality, at any scale.The European Molecular Biology Laboratory\u2019s European Bioinformatics Institute (EMBL-EBI) provides public data resources for many domains of biological research. This includes biological imaging, with the Electron Microscopy Public Image Archive addresses. They are used across the globe, with access in 2021 recorded from every United Nations (UN) member state country. Many resources are leading members of international consortia such as the International Nucleotide Sequencing Consortium, wwPDB or the ProteomeXchange Consortium. Together, they provide immense value to scientists worldwide.Providing access to imaging data at this scale reveals many challenges, some of which are unique to the imaging world, and others that are more general.Biological imaging is a broad term, covering a very wide range of imaging techniques, including the many variants of light-based microscopy as well as electron microscopy, scanning probe microscopy and others Fig.\u00a0. Each ofThese differences in capture methods, and their implementation in different instruments, have given rise to many different file formats in which image data can be represented. For those wishing to view, access and reuse open imaging data, this morass of file formats is a significant barrier.In addition, public image data resources must organise these different types of images in ways that ensure they remain findable. Organising data requires good-quality metadata . EMPIAR\u2019s largest individual datasets are over 70\u00a0terabytes in size, which can potentially take many weeks to transfer, particularly since network bandwidth is not scaling at the rate of data growth. Automated checks that run on data files often have to read the entire image, which scales linearly with image size. Scaling storage to cope with this data volume also requires significant financial and infrastructural commitment, although there are economies of scale that help large institutional data providers.Very large individual images are also challenging to work with. Such large individual images arise from different capture methods \u2013 vEM techniques can produce very large images, as can techniques that stitch 2D electron microscopy images or because of sectioning techniques where each section image is stored individually. These are difficult for traditional computer file systems to deal with; directories with millions of individual files cause significant performance issues for example. Such datasets also require carefully chosen ways in which to store metadata since supporting searches across millions of files can be challenging.Finally, the number of individual datasets that must be managed is a challenge. Human curation of datasets is often required to ensure appropriate standards have been followed. Time required for this curation scales linearly with the number of datasets that must be reviewed. Additionally, as datasets become more complex, time required for curation also increases, such as for the correlative or multimodal data described below for which curation requires expertise in multiple different biological data domains.Part of the increased complexity in biological imaging comes from multimodality. Imaging across scales often requires correlative approaches, for example, combining fluorescence microscopy to identify regions of interest followed by detailed exploration using electron microscopy . Nevertheless, data access patterns vary, with some datasets accessed much more frequently than others; hence, it is possible to tune storage solutions to access patterns. In particular, we can usefully combine faster storage for more recent and more frequently accessed datasets, with cheaper, more capacious storage for others. For rarely accessed data, images can be kept on \u201ccold storage\u201d such as magnetic tape. This \u201ctiered\u201d storage approach smooths the path towards long-term sustainability.Key early work on image metadata developed standards and models for image files is a set of guidelines developed by representatives of different subgroups within the wider biological imaging community , a \u201ctrain-the-trainer\u201d approach enables the widening of the reach of good practice. Furthermore, their feedback and guidance are critical to those attempting to develop user-friendly systems for submission, curation and reuse of image data.The bioimage analysis community is a critical piece of the image data puzzle . The challenges of sharing sensitive data compound the difficulties described above with the legal, ethical and regulatory demands on managing infrastructure for controlled access. However, there are clearly huge potential benefits to being able to share sensitive human imaging data, including the development of new clinical methods and diagnostic tools.The European Genome\u2013Phenome Archive (Brazma et al. Scientific communities and projects have limited resources when it comes to supporting data exchange and publishing. Standards implementations that are initially complex (Spellman et al. The main principles of building a successful scientific data archive are: capture experimental data and supporting metadata, i.e., scientific record; provide a simple data submission process; facilitate standards development, and adopt existing standards; and do not perform data value judgements or complex data integration. A value added resource may enrich, combine and curate datasets; reprocess data; run meta-analysis; etc. The two sets of functions are quite distinct and require different approaches to resource governance, processes and technical infrastructure.As data resources have developed, the need for large-scale collaborations to share the burden of management has grown. The International Nucleotide Sequence Database Collaboration (Cochrane et al. The results of individual experiments do not exist in a vacuum. The systems, organisms, processes, genes, compounds, tissues and cells studied by imaging are examined by multiple routes of study. As discussed above, imaging data is often at its most informative when it is combined with other kinds of data. Ensuring that image metadata are recorded in ways that allow linking to these other data supports such data integration. An example is the use of consistent identifiers for proteins in a localisation study so that they can be linked to a protein sequence database such as UniProt (The UniProt Consortium Beyond immediate reuse cases, providing well-organised biological data can bring unexpected benefits. The AlphaFold Protein Structure Database (Varadi et al. Considering the possibilities enabled by cloud-ready imaging formats, the development of metadata models and the guidance from other domains of biological data, we can suggest next steps to support large-scale open sharing of image data and metadata.As the community converges on a small set of imaging formats that can meet the needs of data generators, analysts and depositors, it becomes possible to build a rich ecosystem of tools and resources around them. In addition to reducing the need for data conversion, these formats provide a standardised protocol for online data access, mitigating the need for time-consuming downloads, and enabling rapid visualisation.Controlled vocabularies and full ontologies are key components of enabling data integration and comparison. Agreement on which of these to use across the community of those managing imaging data would be a good first step towards enabling interoperability.Although the needs of different data management systems differ, agreeing on a shared core set of metadata based on the REMBI principles would provide a basis for interoperability. An initial core set could be agreed across existing data providers and extended over time.Once standardised data formats, shared vocabularies and metadata schemas are aligned, collaboration across data resources that manage imaging data becomes much easier. Data can flow between pre-publication data management systems and public repositories, and federation across large-scale archives becomes possible.Open data is fundamental to good science. It supports reproducibility and auditability, without which science cannot prosper. It helps with method development by providing reference example data for image analysis tools and techniques. Open data is particularly critical for biological imaging because the field moves so fast, with new developments in technology allowing access to higher spatial and temporal resolutions, larger tissue volumes and more multiplexing.A recent case study in image data reuse highlights the benefits of open provision of well-annotated image data. The Nucleome Browser (Zhu et al. If these approaches can be enabled for the huge volumes of imaging data now captured, there are rich opportunities for examining the systems of life at higher detail and improving our understanding of biology at unprecedented scales. Taking lessons from the wider world of biological data and building a roadmap from data and metadata standards will enable realisation of this vision."} +{"text": "Atherosclerosis (AS) is a chronic inflammatory disease that might induce severe cardiovascular events, such as myocardial infarction and cerebral infarction. These risk factors in the pathogenesis of AS remain uncertain and further research is needed. This study aims to explore the potential molecular mechanisms of AS by bioinformatics analyses.GSE100927 gene expression profiles, including 69 AS samples and 35 healthy controls, were downloaded from Gene Expression Omnibus database and indenfied for key genes and pathways in AS.A total of 443 differentially expressed genes (DEGs) between control and AS were identified, including 323 down-regulated genes and 120 up-regulated genes. The Gene ontology terms enriched by the up-regulated DEGs were associated with the regulation of leukocyte activation, endocytic vesicle, and cytokine binding, while the down-regulated DEGs were associated with negative regulation of cell growth, extracellular matrix, and G protein-coupled receptor binding. KEGG pathway analysis showed that the up-regulated DEGs were enriched in Osteoclast differentiation and Phagosome, while the down-regulated DEGs were enriched in vascular smooth muscle contraction and cGMP-PKG signaling pathway. Using the modular analysis of Cytoscape, we identified 3 modules mainly involved in Leishmaniasis and Osteoclast differentiation. The GSEA analysis showed the up-regulated gene sets were enriched in the ribosome, ascorbated metabolism, and propanoate metabolism. The LASSO Cox regression analysis showed the top 3 genes were TNF, CX3CR1, and COL1R1. Finally, we found these immune cells were conferred significantly higher infiltrating density in the AS group.Our data showed the pathway of Osteoclast differentiation and Leishmaniasis was involved in the AS process and we developed a three-gene model base on the prognosis of AS. These findings clarified the gene regulatory network of AS and may provide a novel target for AS therapy.The online version contains supplementary material available at 10.1186/s12920-023-01533-8. Atherosclerosis is one of the most common diseases in the cardiovascular system, which threatens human health. The vascular disease causes many vascular complications affecting the cardiovascular system , 2. AS mThe pathological manifestations of AS exhibit the formation of fatty-fibrous plaque, including the adhesion of activated leukocytes, the proliferation of vascular smooth muscle cells (VSMC), and the degradation of the extracellular matrix, angiogenesis, and calcification. Advanced atherosclerotic plaques developed a morphologically fibrous cap composed of VSMCs proliferating and migrating from the vessel wall into the plaque. Our previous studies have reported that pathophysiological factors were involved in the development of AS, including the phenotypic switch of VMSC, the source of oxidative stress, and mitochondrial injury , 4.Atherosclerosis is primarily a chronic inflammatory disease of the arterial wall response to injury, which is mediated by the oxidized low-density lipoprotein (LDL) . In acutMore research will be needed in the public health and medical field and might reduce the incidence and mortality. Given that microarray analysis can provide an enormous amount of data about gene expression, these analysis methods have been used to predict novel prognostic markers and therapeutic targets . In the We used the \u2018GEOquery\u2019 package of R software (4.0.2 version) to download the GSE100927 microarray data from the GEO database. The GSE100927 data set was based on the GPL17077 platform and contained 104 samples, including 69 atherosclerotic arteries and 35 healthy arteries.The raw data of GSE100927 was read by the affy package in R (version 3.6.1) and the rate monotonic algorithm was used for the background correction and data normalization. Hierarchical clustering analysis was used to group into 2 similar expression patterns of AS arteries and control. The principal component analysis (PCA) was performed to define probe quality control by the \u201cstats\u201d R package. DEGs were screened by the \u201climma\u201d package. The cut-off criterion for statistically significant DEGs was determined as |log2FC|>\u20091 and P value\u2009<\u20090.05. The significant DEGs volcano map was drawn using the \u201cggplot2\u201d package. The 15 top up-regulated and down-regulated genes were drawn using the \u201cheatmap\u201d package.https://david.ncifcrf.gov/) [Candidate genes enriched functions and pathways were identified by using the online DAVID tool (rf.gov/) , which wrf.gov/) \u201313. Bothhttp://string-db.org) to evaluate the relationships of DEGs through protein\u2013protein interaction (PPI) information [These significant DEGs were conducted in the STRING database logistic regression to perform feature selection to screen diagnostic markers for AS arteries . The expWe analyzed the fractions of immune cells in each sample by CIBERSORT . We perfData were expressed as means\u2009\u00b1\u2009SD. Student\u2019s paired t-test analyses were performed in two groups. The significant differences were considered when P\u2009<\u20090.05. R version 4.0.2 was applied to perform the analysis.DEGs were separately identified to analyze the microarray data of GSE100927 from each chip. Figure\u00a0GO and KEGG pathway enrichment analysis of DEGs were performed by using the DAVID online databases. DEGs were classified into biological processes (BP), cellular components (CC), and molecular functions (MF). The top 7 GO enrichment analysis results of DEGs were listed in Fig.\u00a0Figure\u00a0We employed the STRING online database to construct a protein\u2013protein interaction (PPI) network of DEGs. DEGs were imported into the STRING database and a total of 421 nodes and 3529 edges were obtained with a scoring value\u2009>\u20090.4 contained 34 nodes and 361 edges with a score of 21.87. The top 7 genes were ITGAM, TLR1, CCR5, ITGAX, TNF, CD68, and CCR2 software. GSEA analysis showed that 122/186 gene sets were up-regulated, while 64/186 gene sets were down-regulated between AS group and the control group. Table To obtain the diagnostic markers genes, Least Absolute Shrinkage and Selection Operator (LASSO) Cox regression analysis was performed based on the gene expression of DEGs. We used the LASSO logistic regression algorithm to screen the diagnostic markers from 443 DEGs. We randomly selected 70% of samples in GSE10927 as the training set construction of the models and the remaining 30% of samples as the test set . A coefficient profile plot was produced against the log (lambda) sequence. Selection of tuning parameter (lambda) in the LASSO model used tenfold cross-validation via minimum criteria. Figure\u00a0The KEGG pathway enrichment analyses were related to infection, such as Leishmaniasis, and phagosome. To assess the potential associations, we analyzed the different immune cell types between AS group and the control group. The heatmap presented the different immune cell infiltration patterns between the two groups, where the colors displayed the infiltration density Fig.\u00a0A. In AddAtherosclerosis (AS), as a metabolic syndrome, posed a serious threat to human health and caused a series of complications affecting the cardiovascular system, including acute myocardial infarction and cerebral infarction . It is wBP analyses demonstrated that the up-regulated DEGs were mainly enriched in muscle contraction and negative regulation of the BMP signaling pathway, while the down-regulated DEGs were enriched in immune response and inflammatory response . These fWe identified the top 10 hub genes based on degree value by using Cytoscape software, including PTPRC, TYROBP, TNF, ITGAM, SPI1, LCP2, CSF1R, CCR5, and ITGAX. Nie reported that PTPRC was considered to be an immune marker gene in atherosclerosis development . Liu repThe functions of the genes in three important modules from PPI were selected to further perform the KEGG pathway enrichment analysis. The common three KEGG enrichment pathways were osteoclast differentiation, phagosome, and Leishmaniasis. These findings are in line with other research which reported that AS development is a result of imbalanced lipid metabolism and the AS progression was mediated by the phagosome of macrophages and smooth muscle cells . The finhigh monocytes and TNF-\u03b1 as potential therapeutic targets for preventing infection-induced cognitive dysfunction [To evaluate the prognostic genes based on DEGs of GSE100927, we applied the LASSO Cox regression model to construct a prognostic gene signature. We identified a gene signature involved in AS. The AUC value of the three-gene signature suggested the prediction ability and the prognostic values of these gene signatures. TNF-\u03b1 could induce the up-expression of CX3CR1 in endothelial cells and BV-2 microglial cells , 41. Morfunction . The novAS is a chronic inflammatory disease involving multiple types of immune cells. Chistiakov reported that Tregs mediated the immune response and the secretion of anti-inflammatory cytokines IL-10 and TNF-beta . CD8\u2009+\u2009TThe main pathological process of AS involved vascular endothelial cell damage, lipid deposition, and plaque formation . Our resThere are also some limitations in our study. Firstly, the hub genes and the gene signature were analyzed using gene expression profiles of human samples from online public databases; further studies using more human samples will be required to validate these marker genes by WB experiments. Secondly, more experiments need to examine these potential mechanisms related to the phagosome, Leishmaniasis, and osteoclast differentiation in tissue samples from patients with AS.In summary, our study displays a new comprehensive bioinformatics analysis of DEGs that might be involved in the AS progress. We identified 10 hub genes and three modules that were highly associated with AS. The KEGG pathways were mainly enriched in phagosome, Leishmaniasis, and Osteoclast differentiation. We developed and validated a three-gene signature to predict the prognosis of AS. These novel molecular targets are principal candidate genes for further investigations into biomarkers and molecular mechanisms.Additional file 1. Table S1. The KEGG pathway of Module 1, Related to Figure 4A. Table S2. The KEGG pathway of Module 2, Related to Figure 4B. Table S3. The KEGG pathway of Module 3, Related to Figure 4C."} +{"text": "The breast imaging-reporting and data system (BI-RADS) grading has a great advantage in diagnosing breast diseases, but with some limitations.The study analyzed the value of ultrasound-guided core needle biopsy (CNB) in diagnosing BI-RADS grades 3, 4, and 5 breast cancer.Breast cancer patients at BI-RADS grades 3 to 5 received breast ultrasonography, ultrasound-guided CNB and immunohistochemical examination. Receiver operating characteristic (ROC) curve was made to test diagnostic efficiency of regression model.Calcification was positively correlated with expression of estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor (HER)-2. The areas of 4 ROC curves were 0.752, 0.805, 0.758, and 0.847, and the 95%CI was 0.660 to 0.844, 0.723 to 0.887, 0.667 to 0.849, and 0.776 to 0.918, respectively. BI-RADS grades 3 to 5 were positively correlated with expression of ER, PR and human epidermal growth factor receptor-2 (HER-2). Statistical significance existed between grade 5 and expression of ER, PR and HER-2, and between grade 4 and expression of HER-2.The study demonstrates that BI-RADS can be used as an effective evaluation method in the diagnosis of breast diseases before invasive operation, and it has higher diagnostic accuracy if combined with pathological examinations. In recent years, the incidence of breast cancer has been increasing year by year, ranking the first among all female malignant tumors in China. With the improvement of breast cancer treatments, the survival of breast cancer patients has been further increased. However, some patients at advanced stage or with poor molecular typing have poor prognosis, leading to deaths of the patients. With the continuous improvement of diagnosis and treatment, the screening of breast tumors has been significantly improved, and greatly improved the survival of breast cancer patients. The examination of breast tumors is mainly through combining clinical palpation with imaging examination. Ultrasonography is the most widely used method in clinical application. It usually judges benign and malignant tumors according to the shape, boundary, hyperechoic halo around lesions, echo type and blood supply.,4 However, ultrasound examination still has its own limitations. In the examination of small breast cancer, the ultrasonographic signs are not obvious, leading to low detection rate and even missed diagnosis. The American College of Radiology has updated the new grading standard suitable for the Breast Imaging-Reporting and Data System (BI-RADS), providing a basis for the standardization and normalization of ultrasound diagnosis for breast lesions, which has enhanced the objectivity of ultrasound diagnosis of the disease. However, the application of BI-RADS grading in the diagnosis of atypical lesion or microlesions still has some limitations. Especially at grades 3 to 5, there are suspicious malignancies and misdiagnosis. For cases with atypical ultrasonographic signs, clinical manifestations should be considered, and puncture biopsy is needed. Core needle biopsy (CNB) is minimally invasive, simple and rapid for the sampling and diagnosis of suspicious lesions. It can avoid incision and enhance the cosmetic effect. Previous studies are mainly focused on how to improve the accuracy of lesions above grade 3 in BI-RADS classification using ultrasound technology.\u201310 When necessary, hormone-dependent tumor markers such as estrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor (HER)-2 are also considered. In addition, histological structure and morphological characteristics also determine the ultrasonographic manifestations of tumors, which are closely related to the occurrence, development, metastasis and recurrence of breast cancer.Breast cancer is a common malignant tumor in women.The aim of the present study is to retrospectively analyze the value of ultrasound-guided CNB in diagnosing BI-RADS grades 3, 4, and 5 breast cancer, and to study the correlation between ultrasonographic signs and molecular biological markers, in order to provide evidence for early diagnosis, treatment and prognosis evaluation of breast cancer.A total of 120 patients who were diagnosed with breast cancer by ultrasound at our hospital between May 2016 and May 2019 were included in the study. The age range of the patients was 25 to 65 yeas, and the median age was 48 years. The diameter of nodules ranged between 5\u2009mm and 25\u2009mm. The inclusion criteria were: diagnosis of BI-RADS grades 3, 4, or 5 by high-frequency ultrasound and color Doppler ultrasound, as well as comparison with postoperative pathological results. All procedures performed in the current study were approved by the Ethics Committee of China Medical University. Written informed consent was obtained from all patients or their families. were listed in Table The diagnostic criteriaCNB was performed under the guidance by Color Ultrasound Diagnostic Instrument . The frequency of the probe was 10 MHz. Magnum Disposable Core Tissue Biopsy Needle had a needle groove length of 20 to 22\u2009mm. During color Doppler ultrasonography of the breast, the details of the lesions were observed and recorded, and the distance between the mass and the nipple was measured. BI-RADS grading was performed according to the shape, margin, direction, internal echo, posterior echo, changes of surrounding tissues, calcification, blood supply and lymphatic drainage. Breast masses of BI-RADS grade 3 to 5 were selected for the study. The selection of puncture points should be maintained within the surgical reach. The distance between puncture point and pleura should be greater than the range of puncture needle. The insertion depth of biopsy needle should be determined according to the depth, size and correlation of the lesions. General operations retained 3 to 6 samples. After operation, the biopsy needle was pulled out, the size of the tissue strip was recorded and the tissues were placed in fixative fluid for subsequent pathological examination was determined by immunohistochemistry according to the manufacturer manual .Positive results of ER and PR were judged by the following 2 scoring systems. First, ten high-power visual fields of uniformly colored tumors were selected, the number of positive cells and the total number of cells were counted, and the percentage of positive cells was calculated. If no positive cells were observed, 0 score was recorded. If the percentage of positive cells was within 1% to 10%, 1 score was recorded. If the percentage of positive cells was within 11% to 50%, 2 scores were recorded. If the percentage of positive cells was within 51% to 80%, 3 scores were recorded. If the percentage of positive cells was\u2005>80%, 4 scores were recorded. The second scoring system depended on coloring depth. If no color was observed, 0 score was recorded. If light yellow was observed, 1 score was recorded. If light brown was observed, 2 scores were recorded. If dark brown was observed, 3 scores were recorded. The scores in the 2 systems were added up. If the total score was 0 or 1, negative result (\u2212) was achieved. If the total score was 2, weak positive result (+) was achieved. If the total score was 3 to 4, positive result (++) was achieved. If the total score was 5 to 6, strong positive result (+++) was achieved curve was drawn to test the diagnostic efficiency of regression model.The results were analyzed using SPSS 20.0 statistical software . Chi-square test was used to compare the counting data between groups. Spearman grade correlation analysis was used to analyze the correlation between indicators. P\u2005<\u2005.05) . Microcalcification was positively correlated with positive expression of ER, PR and HER-2 (P\u2005<\u2005.05) in ultrasound signs were positively correlated with biological indicators. Boundary burr, mass shape, mass size, lymph node enlargement and CDFI expression were not significantly different between groups for ER, PR and HER-2 P\u2005>\u2005.0. MicrocaROC curves for multiparameter regression models of microcalcification, ER, PR and HER-2 were drawn. The area under the curve for the 4 ROC curves were 0.752, 0.805, 0.758, and 0.847, respectively. Their 95% CI were 0.660 to 0.844, 0.723 to 0.887, 0.667 to 0.849, and 0.776 to 0.918, respectively . In BI-RADS grade 5, there was statistical significance for the expression of ER, PR and HER-2 (P\u2005<\u2005.5). In BI-RADS grade 4, there was no statistical significance for the expression of ER and PR (P\u2005>\u2005.5) Table . Early detection, accurate diagnosis and timely treatment of breast cancer directly affect the treatment effect and prognosis.,16 Ultrasound-guided puncture is convenient, minimally invasive and easy to be widely used in clinic. It has become the preferred examination method for breast diseases. In the 2017 edition of Guidelines and Specifications for the Diagnosis and Treatment of Breast Cancer, the Chinese Anti-Cancer Association recommends performing preoperative image-guided biopsy in qualified hospitals. The pathological and histological morphology of tumors is the basis of the ultrasonographic manifestations of breast cancer. Understanding this is essential for analyzing the ultrasonographic features of breast cancer. Mammary gland is a hormone-dependent organ. The expression of hormone receptors is often altered in the occurrence of breast cancer. Therefore, breast cancer is also considered as a hormone-dependent tumor. At present, there are more and more mature studies in breast cancer, and the molecular biological indicators widely used in clinic include ER, PR, and HER-2, which are closely related to the occurrence, development, diagnosis, treatment and prognosis of breast cancer. They are the most common biomarkers indicating the biological behavior and prognosis of breast cancer.In recent years, the incidence of breast cancer has shown an increase trend, and the age of onset is decreasing. Because the BI-RADS grading system is obtained through the relevant indicators of ultrasound images, and through the creation of risk assessment model with biomarkers, the risk assessment model has a certain correlation with the tumor progression before the clinical manifestation of the disease, which is particularly valuable in clinical application.The diagnostic method introduced here is based on the analysis of biological factors related to histological samples obtained from ultrasound images and biopsy. The higher the relevant grading index of BI-RADS grading system is, the worse the prognosis is. They are complementary to biological functions. Different grading and biological indexes can evaluate the development and progress of the tumor. The main reasons are the number of puncture samples, the accuracy of sampling and the pathological types of lesions. Puncture biopsy is mainly towards cystic tissues, and the solid part is usually little, so it is easy to misdiagnose highly differentiated and low-grade malignant breast cancer as benign nodules. Therefore, it is suggested that an accuracy of 99% can be achieved by taking at least 4 to 5 samples during CNB, while other researchers believe that multi-point puncture has no significant effect on diagnosis.\u201310 It is also believed that false negative rate is different among different pathological types. For example, ultrasound findings of in situ and highly differentiated carcinomas are atypical and classified as BI-RADS grade 3, resulting in missed diagnosis. In the meantime, some special types of benign lesions are classified as BI-RADS grade 5 because of malignant signs. This requires that we strictly follow the BI-RADS classification criteria, and observe the dynamic state of each section layer by layer.For patients with BI-RADS grades 3 to 5, the specificity in the diagnosis of breast lesions was 100%, and the sensitivities were 70.0%, 89.2%, and 80.9%, respectively. There was a positive correlation among the groups, with statistical significance. The false negative rate of CNB for patients at BI-RADS grade 3 was 30%, which was significantly higher than those of patients at BI-RADS grade 4 or 5, being acceptable according to a previous report.,20 The amount and size of calcification are closely related to the malignant degree of breast cancer and can be used as an important characteristic index for the diagnosis of breast cancer. In the present study, the expression of ER, PR, and HER-2 in calcification group was significantly higher than that in non-calcification group, being consistent with the above conclusions. A study shows that the diagnostic efficiency of lymph node biopsy for breast cancer is high, and the area under ROC is higher than 0.9.. The diagnostic efficiency of BI-RADS itself in breast masses with calcification, solid, cystic and cystic solid lesions is relatively lower than that of puncture biopsy, and the misdiagnosis rate is high due to the nature of the lesions. In the process of puncture, if the breast mass is cancerous with cystic lesions, the measured biomarker indexes are low or false negative. The combination of the 2 is able to diagnose breast lesions and help clinicians understand the correlation between ultrasound findings and pathology, so as to make a diagnostic model for clinicians before follow-up treatment.Ultrasound signs are closely related to pathological basis. The shape, boundary and size of tumors are very similar to those of pathological specimens. The growth patterns of breast cancer in tissue structure are different. In this study, the shape, size, marginal burr, calcification, lymph node enlargement and CDFI observed in ultrasound examination were positively correlated with the biological manifestations. Significant differences were only found between calcification and the positive expression of ER, PR and HER-2. Up to now, the mechanism of calcification in breast cancer remains unclear. However, it is reported that bone matrix proteins are found in breast cancer tissues, and calcification in calcium and phosphorus deposition environment formed by bone matrix protein metabolism is associated with positive expression of ER, PR, and HER-2.,22 In the present study, the expression levels of ER, PR and HER-2 were positively correlated with BI-RADS grades 3, 4 and 5. However, no statistical significance existed between grade 3 and the expression of ER, PR and HER-2. Statistical significance existed between grade 5 and the expression of ER, PR and HER-2. In addition, no statistical significance existed between grade 4 and the expression of ER and PR, while statistical significance existed between grade 4 and the expression of HER-2. When breast cancer occurs, cells containing ER, PR and HER-2 proliferate malignantly, causing positive expression of ER, PR and HER-2. This is obviously related to the changes among grades 3 to 5 in BI-RADS classification. For BI-RADS grade 4, the expression of ER, PR and HER-2 were not consistent, possibly because, with the increase of BI-RADS grades, HER-2 amplification and expression can inhibit cell apoptosis, regulate cell differentiation, promote cell proliferation and neovascularization, leading to breast cancer.In the present study, the results of pathological examination after breast puncture biopsy and pathological diagnosis of the specimens after operation were taken as the gold standard. The areas under the 4 ROC curves was 0.752, 0.805, 0.758, and 0.847, respectively, and the 95% CIs were 0.660 to 0.844, 0.723 to 0.887, 0.667 to 0.849, and 0.776 to 0.918, respectively. Therefore, the area under ROC curve has certain clinical significance in the diagnosis of malignant breast tumors. Different areas under ROC between lesion biopsy and lymph node biopsy might be due to their different ultrasound shapes. Lymph node enlargement or metastasis is usually discovered at stages higher than grade 5 in BI-RADS classification. Therefore, effective judgment of BI-RADS classification, combined with biological manifestations, can effectively improve the diagnostic efficiency. At present, there are few studies on the biological expression of breast cancer at BI-RADS grades 3 to 5, and many studies have focused on a single ultrasound indicator. In particular, predictive biomarkers for treatment stratification, diagnostic biomarkers for early detection and prognostic biomarkers for estimating the clinical outcome of patients are essential for the prevention of tumors. The pathogenesis of breast cancer should be studied from gene mutation, abnormal glycosylation level of protein, metabolism and imaging manifestations.The present study demonstrates that ultrasound-guided CNB is easy to operate and can cause weaker trauma, and thus has high diagnostic value for breast cancer in combination with BI-RADS classification. BI-RADS classification is accurate, but there is still a certain rate of missed diagnosis. The combination of breast ultrasound signs with BI-RADS classification can predict the biological indicators of breast cancer, such as ER, PR and HER-2. The presented achievements may improve the overall BC management by providing evidence for early diagnosis, treatment and prognosis evaluation of breast cancer. Although many tumors have been effectively treated, the exact mechanisms of tumor occurrence and development still remain unclear. Cancer is a complex systemic disease, involving various abnormalities of genes, proteins, metabolites and medical images.The authors wish to thank their department and research team for their help and dedication.Conceptualization: Tiejun He.Data curation: Tiemei Shi, Yabo Ju.Formal analysis: Tiejun He, Wendong Luo, Yabo Ju.Funding acquisition: Tiejun He.Investigation: Tiemei Shi, Wendong Luo, Yabo Ju.Methodology: Tiemei Shi.Project administration: Tiejun He, Wendong Luo, Ran Li.Resources: Tiemei Shi, Ran Li.Software: Tiejun He, Yabo Ju.Supervision: Tiemei Shi, Yabo Ju, Ran Li.Validation: Tiejun He, Wendong Luo, Ran Li.Visualization: Tiemei Shi, Ran Li.Writing \u2013 review & editing: Tiejun He.Writing \u2013 original draft: Tiemei Shi." \ No newline at end of file