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+{"text": "Angus M. GunnWestport, CT: Greenwood Press, 2004. 143 pp. ISBN: 0-313-31999-5, $55 cloth.The Impact of Geology on the United States: A Reference Guide to Benefits and Hazards . In this second volume, Angus M. Gunn provides an overview of human-made environmental disasters. He shows that although technology has given humankind enormous control over the environment, it has also proven to be a threat to our survival. Gunn categorizes these human-made disasters into a number of subtypes\u2014for example, mining disasters, dam failures, government actions, industrial explosions, oil spills, nuclear energy catastrophes, and terrorism. For each of these types of disaster, the book contains 26 case examples describing the events that led up to the disaster, the technical details of the event itself, the cleanup it necessitated, and its consequences. Some of the examples described in the book are famous\u2014for example, the Minimata mercury poisoning in Japan, the Buffalo Creek dam collapse in West Virginia, and the near accident at the Three Mile Island nuclear plant in Pennsylvania. Others have been almost forgotten, such as the deliberately induced great famine in Ukraine in 1932, which resulted from the massive collectivization of farms ordered by Stalin.This book was written as a sequel to an earlier volume by the same author on the impact of natural disasters, Toxic Turmoil: Psychological and Societal Consequences of Ecological Disasters. New York:Kluwer Academic, 2002). Another shortcoming of the book is that some of the medical information cited in the book appears to be incorrect, as presented in the case of the Chernobyl accident. Without due reference the author states that this accident caused a steady rise in miscarriages and birth defects in Belarus and will eventually have generated a death toll of over 5 million people. Claims such as these are entirely unfounded, as noted by Bard et al. .The book is well written and successfully combines factual information with good journalism. Gunn professes to stick to the tried-and-true methods of the hard physical sciences. The consequence of this choice is that the book makes no reference to the societal and psychological impacts of disasters. Interestingly, it is exactly some of the case examples given in the book, such as the Three Mile Island incident, that have led to the recognition of the importance of these secondary effects. This omission is especially obvious in the case of terrorist events, which are precisely intended to cause fear and social discord as much as physical damage. Readers who are interested in the full picture of the impact of human-made disasters, including their underlying psychological and societal dynamics, should therefore turn to other volumes (e.g., Havenaar JM, Cwikel JG, Bromet EJ, eds. Unnatural Disasters is a well-written book containing a wealth of historical details about some classical environmental disasters, but it is not suitable as a reference for public health purposes.In summary,"}
+{"text": "Msh2 in chromosome stability has been investigated in a targeted mouse model for HNPCC Msh2\u03947N. Chromosome aberration frequencies were similar in bone marrow of Msh2+/+Msh2+/\u2013and Msh2\u2013/\u2013mice and no differential effects of in vivo X-irradiation were noted. By contrast, the induction of sister-chromatid exchanges (SCEs) by methyl nitrosourea (MNU) was reduced in Msh2\u2013/\u2013and Msh2+/\u2013cells to ~20% and ~45% wild-type levels respectively indicating a phenotypic effect of haploinsufficiency of the mouse Msh2 gene. \u00a9 2000 CancerResearch CampaignThe role of"}
+{"text": "Studies on the regulation of phage \u00d829 gene expression revealed a new mechanism to accomplish simultaneous activation and repression of transcription leading to orderly gene expression. Two phage-encoded early proteins, p4 and p6, bind synergistically to DNA, modifying the topology of the sequences encompassing early promoters A2c and A2b and late promoter A3 in a hairpin that allows the switch from early to late transcription. Protein p6 is a nucleoid-like protein that binds DNA in a non-sequence specific manner. Protein p4 is a sequence-specific DNA binding protein with multifaceted sequence-readout properties. The protein recognizes the chemical signature of only one DNA base on the inverted repeat of its target sequence through a direct-readout mechanism. In addition, p4 specific binding depends on the recognition of three A-tracts by indirect-readout mechanisms. The biological importance of those three A-tracts resides in their individual properties rather than in the global curvature that they may induce. Gene expression is regulated primarily at the level of transcription initiation, mainly by \u03c3 factors and by transcription factors that facilitate or prevent interactions of the RNA polymerase (RNAP) with the promoter. To initiate transcription from specific promoters, the bacterial RNAP core must associate with the initiation factor \u03c3, which contains determinants that allow sequence-specific interactions with promoter sequences . A famil\u03bb can function as a repressor or activator, exerting the transition from one program of \u03bb gene expression to another upon the formation of a higher-order protein-DNA complex [Escherichia coli is a dimer capable of self-association to hexamers. TyrR dimers activate transcription, but TyrR hexamers repress transcription binding to targets that overlap the promoter sequence [Transcription factors are mostly regulatory proteins that bind to DNA sequences generally at or nearby the promoter sequence. These sequence-specific protein interactions are usually responsible for regulating transcription initiation . However complex . Similarsequence . Furthersequence \u201326. Bindsequence \u201333.2.Bacillus subtilis \u03c3A-RNAP [B. subtilis has a subunit composition of \u03b2, \u03b2\u2019, \u03b12 and \u03c9, homologous to the E. coli enzyme. The B. subtilis \u03c3A subunit is homologous to E. coli \u03c370; both recognize the same consensus sequences at the \u221235 and \u221210 hexamers [Phage \u00d829 gene expression is directed by the  \u03c3A-RNAP ,35. The hexamers \u201343.B. subtilis, only those genes involved in DNA replication and transcription regulation are expressed at early times . However, protein-DNA complex formation frequently requires additional interactions whereby bases not contacted by the protein and apparent unspecific interactions provide specificity by an \u201cindirect-readout\u201d mechanism \u201359. The Protein p4 is a DNA binding protein \u201366 that 10-helix. This is a novel fold, and searching for structurally related proteins [N-terminus where the polypeptide chain from Pro2 to Gln5 runs anti-parallel to the stretch from Arg6 to Asp11. This structural element, named \u201cthe N-hook,\u201d is a key feature for DNA recognition. In the structure of the p4-DNA complex, two p4 protomers are bound to the same face of the DNA helix. The DNA presents a continuous curved B conformation towards the bound protein that correlates with minor groove narrowing at the concave face and minor groove widening at the convex face, while the major groove is quite regular. The hooks, located at the tips of the p4 dimer, intrude into the DNA major groove making the only amino acid-base contact of the complex. The guanidinium group of Arg6 bonds with G at positions + and \u221213 (\u00b113). In addition, three positively charged patches in the p4 dimer interact with DNA backbone phosphates at three separated A-tracts  simulations of DNA and p4-DNA complexes to investigate the basis for the p4-DNA complex specificity . In the 6 . TherefoC-termini of \u03b11 helices contain a cluster of 12 positively charged amino acids, located after a kink that maintain the helix almost parallel to the DNA axis  the asymmetry of the DNA target is functionally required for p4-DNA interaction; (ii) p4 curves the DNA, and (iii) the distance from G \u2212 13 to G + 13 is about 90 \u00c5 while the 75 \u00c5 distance from the Argonomer A .5.B. subtilis, the host RNAP starts transcription from early promoters A2b and A2c. RNAP recognizes promoters A2c and A2b through interaction of the \u03b1 and \u03c3 subunits at the promoter elements [Upon \u00d829 infection of elements . Efficieelements . Syntheselements . This haelements . The nucelements ,80.6.The switch from early to late transcription of the \u00d829 genome is tightly regulated to ensure the appropriate sequence of gene expression. Repression of early promoters A2c and A2b and activation of the late promoter A3 are simultaneously regulated in a sophisticated manner by proteins p4 and p6, where protein p4 has a leading role in the process.54 would narrow it providing optimal complementarity between one p4 surface and its target. Therefore, p4 creates specificity in the protein-DNA complex using the intrinsic properties of minor groove A-tracts.The study of protein p4 revealed novel protein-DNA interaction paradigms: (i) The p4 structure adds a new DNA binding motif to the catalogue of DNA binding protein motifs, the N-hook; (ii) p4 recognizes its targets through direct-readout of the boundary guanines in its target sites and by two additional indirect-readout mechanisms. Both indirect-readout mechanisms are based on the malleability of A-tracts. First, remodeling the topology of the external A-tracks, p4 provides a better adjustment of the N-hook to the DNA. Second, the over-winding of the central minor groove by the insertion of ArgProtein p4 bound to its targets remodel 120 base pairs of DNA to the structure of a nucleoprotein-hairpin that is stabilized by the incorporation of p6. The nucleoprotein-hairpin is the key factor that coordinates gene expression since the switch from early to late transcription is the interplay between the RNAP and the p4-p6 complex for binding to the sequence containing promoters A2c, A2b and A3. Efficient promoter complex formation and transcription initiation requires the appropriate positioning of the RNAP at the promoter. The stability of the hairpin structure, which depends on the availability of proteins p4 and p6 in the cell, might be critical. The hairpin impairs the correct interaction of the RNAP at early promoters A2c and A2b and simultaneously activates late promoter A3 stabilising the primary transcriptional complexes."}
+{"text": "Mental illness currently ranks among the top ten causes of burden of disease in low-income countries . In the A substantive portion of the burden of mental disorders in low-income countries is thought to be attributable to many of the failures of human development as targeted through the Millennium Development Goals (MDGs), including poverty, HIV, and gender inequality. The evidence on depressive disorders and depressed mood is most well developed in this respect .l health .deliver these interventions in different contexts. Given the present lack of adequate mental health care systems financing and lack of adequate human resources for mental health in low-income countries, a scaled-up response will likely involve integration of treatment for mental disorders into primary health care settings [Even were the evidence base on mental health spillovers to be strengthened overnight, additional questions would need to be answered in order to determine how best to settings . Screenisettings . Screenisettings ,54, but settings , and sepsettings -8,56.Lancet\u2019s Global Mental Health series in 2007 [PLoS Medicine Packages of Care series in 2009 [Significant strides have been made in ensuring a greater prominence for mental health on the global agenda, reflected in the  in 2007  and 2011 in 2007 , the PLo in 2009 , and the in 2009 . As of y in 2009 . In low- in 2009 , and the in 2009 . A comprIn the years leading up to 2015, we hope that mental health advocacy will be intensified to ensure that programming and funding for prevention and treatment of mental disorders are not sidelined in future initiatives as they have been to date with regards to the MDGs  and non-"}
+{"text": "Escherichia coli datasets. We demonstrate that the ensemble-based network aggregation approach can be used to effectively integrate GRNs constructed from different studies \u2013 producing more accurate networks. We also apply this approach to building a network from epithelial mesenchymal transition (EMT) signature microarray data and identify hub genes that might be potential drug targets. The R code used to perform all of the analyses is available in an R package entitled \u201cENA\u201d, accessible on CRAN (http://cran.r-project.org/web/packages/ENA/).Reverse engineering approaches to constructing gene regulatory networks (GRNs) based on genome-wide mRNA expression data have led to significant biological findings, such as the discovery of novel drug targets. However, the reliability of the reconstructed GRNs needs to be improved. Here, we propose an ensemble-based network aggregation approach to improving the accuracy of network topologies constructed from mRNA expression data. To evaluate the performances of different approaches, we created dozens of simulated networks from combinations of gene-set sizes and sample sizes and also tested our methods on three  The rank-product method, introduced by Breitling et al An important application of network reconstruction is to identify hub genes in a network that might be biologically and pharmaceutically interesting. When we applied ENA to microarray data that was previously used to delineate an epithelial-mesenchymal transition (EMT) signature Reconstructed gene networks are often returned as a weighted undirected graph Specifically, suppose This algorithm can be efficiently applied to large networks with many reconstructed networks in The initial application was to leverage the rank-product method to \u201cbootstrap\u201d samples. Each time, we constructed the gene network using a randomly selected subset of the available samples. By repeating this process B times, we created a set Of course, this bootstrapping procedure inflates the computational complexity of GRN reconstruction by several orders of magnitude, as GRNs must be reconstructed B times rather than just once. Because each graph in The second application of the rank-product network merging method was to reconstruct an aggregated GRN, based on the output of multiple different reconstruction techniques. We have observed that reconstruction techniques perform differently based on different simulation settings 1, MD2\u2026. MDD as follows:The final application evaluated in this study was in the merging of networks constructed from different datasets. Historically, gene expression datasets have been collected from various sites on different microarray platforms with different procedures for tissue collection, which creates incompatibilities and difficulties when performing analyses on data from different datasets simultaneously. Because the rank-product method makes no assumptions about the distribution of the data at any point, we employ it to combine GRNs produced from different datasets, yielding a single aggregated GRN which aims to capture the consistencies in network topology from the GRNs produced on different datasets. We thus derive the aggregated network from datasets MDhttp://cran.r-project.org/web/packages/ENA/index.html), from which the compiled binaries, as well as all original source code, are also available for download.The code used to bootstrap samples and aggregate the resultant networks was written in the R programming language. We created an R Package entitled \u201cENA\u201d and made it available on CRAN  to control not only the R code developed for the ENA package, but also all code, reports, and data used in the aforementioned simulations and reconstruction techniques; all of this code is freely available at https://github.com/QBRC/ENA-Research. All the data analysis code used to generate the results in this study was compiled into a single report and can be reproduced easily using the knitr R package Additionally, we leveraged the Git revision control system via GitHub  of 0.25, 1, and 2. We used the methods described above to reconstruct three networks (one from each dataset) and then aggregated those networks. For comparison, we also combined all three datasets into a single dataset containing these 600 samples and then reconstructed a single network from this larger dataset.The performance of methods in this setting can be represented by a Receiver Operating Characteristic (ROC) Curve, which plots the True Positive Rate against the False Positive Rate, demonstrating the performance of the method at all relevant edge confidence score thresholds. The performance of a method can be quantified by calculating the Area Under the ROC Curve (AUC). The greater the AUC, the better the performance of the method represented. A perfect reconstruction would have an AUC of 1, while a random guess would obtain an AUC of 0.5. An alternative approach to evaluating gene regulatory network reconstruction is the Area Under the Precision Recall curve (AUPR). In a precision recall curve, recall  is plotted against precision .We found that bootstrapping samples can increase the accuracy of network inference. In our study, we randomly selected 70% of all samples and rebuilt networks and repeated the abovementioned process more than 100 times for each dataset to get the bootstrapping results. For example, the networks reconstructed from the dataset on the 231-gene network with a noise value of 0.25 can be compared to demonstrate variations in performance  and 2.Aside from optimizing individual reconstruction techniques, we found that combining different network reconstruction techniques that were executed on the same dataset also has the power to significantly improve the accuracy of the reconstructed networks. Using the dataset from the 83-gene network with 200 samples and a noise value of 0.25, we evaluated the comparative performance of each reconstruction technique, as well as that of the aggregated network. e.g., some methods perform well on larger networks, while others excel in datasets containing few samples. Thus, to have an aggregation technique that consistently outperforms or matches the best performing individual method eliminates the need to choose a single reconstruction technique based on the context.We also observed this trend to hold true across most of the datasets  datasets: 1) the Many Microbe Microarrays Database (\u201cM3D\u201d) E. coli Genome arrays; 2) the second dataset (\u201cStr\u201d) of expression data from laboratory evolution of E. coli on lactate or glycerol (GSE33147) E. coli to different perturbations and stresses, such as drug treatments, UV treatments and heat shock. The RegulonDB database E. coli, was thus used as a \u201cgold standard\u201d to evaluate the accuracy of the variously constructed networks.We then tested the ENA approach on three E. coli data (We were able to obtain similarly positive results by employing these approaches on the oli data . BootstrCDH1), vimentin (VIM), N-cadherin (CDH2) and/or fibronectin 1 (FN1), and followed a bimodal distribution pattern across the cell lines Network reconstruction of gene expression data helps identify hub genes that might be novel drug targets because of their role in engaging multiple molecules, a process that has been used to identify gene sets predictive of benefit for adjuvant chemotherapy in non-small-cell lung cancer ZEB1, which had the highest degree in the resulting ENA network, is a well-known EMT activator and tumor promoter that represses stemness-inhibiting microRNAs MARVELD3 is known as a tight junction molecule and has been shown to be downregulated during Snail-induced EMT EPHA1, the first member of the erythropoietin-producing hepatocellular (Eph) family of receptor tyrosine kinases, was recently shown to potentially play a role in carcinogenesis and the progression of several cancer types EPHA1 is also frequently mutated in NSCLC patients, along with other known \u201cdriver\u201d mutations Overall, we attempted to identify hub genes clinically interesting for NSCLC treatment. We thus employed multiple methods to build GRN networks and combined them via ENA. As shown in The ability to aggregate networks using the rank-product merging approach has proven to be a valuable contribution in reconstructing gene regulatory networks \u2013 and likely in other fields, as well. By bootstrapping a single dataset using a single approach such as SPACE, we were able to significantly improve the performance of the algorithm. By aggregating the networks produced by different reconstruction techniques on a single dataset, we were able to consistently match or outperform the best-performing technique for that dataset, regardless of fluctuations in the performance of any one algorithm. By aggregating networks constructed independently on different datasets capturing similar biological environments, we were able to reconstruct the network more accurately than would be possible using any one dataset alone. So far, the study of integration of gene regulatory networks has been continuously advancing. Both Marbach D. et al. 2012 It is likely that SPACE was the only method to show consistent and significant improvement from bootstrapping because the SPACE algorithm models gene regulation using linear regression; as a result, the network construction problem is converted to a straightforward variable selection problem. In SPACE, the variable selection problem is solved by sparse regression techniques with a symmetric constraint. By solving all the regression models simultaneously, SPACE attempts to accrue the globally optimized results. However, due to the instability in variable selection http://www.ingenuity.com/products/ipa) is a pathway and network database based on curated literatures. When we used IPA to analyze our data, ZEB1 was identified as a hub gene, which confirmed our discovery using the ENA approach. Additionally, predicted interactions such as the CDH1\u2013CDH3 interaction and the CLDN4-GRHL2 interaction were also confirmed .(DOCX)Click here for additional data file."}
+{"text": "Inflammation could be triggered by activation of transient receptor potential vanilloid 1 (TRPV1) channels in the cochlea and possibly other TRP channels. Targeting TRPV1 for knockdown has also been shown to be a useful strategy for ensuring otoprotection. Cisplatin entry into cochlear hair cells is mediated by various transporters, inhibitors of which have been shown to be effective for treating hearing loss. Finally, cisplatin-induced DNA damage and activation of the apoptotic process could be targeted for cisplatin-induced hearing loss. This review focuses on recent development in our understanding of the mechanisms underlying cisplatin-induced hearing loss and provides examples of how drug therapies have been formulated based on these mechanisms.Evidence of significant hearing loss during the early days of use of cisplatin as a chemotherapeutic agent in cancer patients has stimulated research into the causes and treatment of this side effect. It has generally been accepted that hearing loss is produced by excessive generation of reactive oxygen species (ROS) in cell of the cochlea, which led to the development of various antioxidants as otoprotective agents. Later studies show that ROS could stimulate cochlear inflammation, suggesting the use of anti-inflammatory agents for treatment of hearing loss. In this respect, G-protein coupled receptors, such as adenosine A Cisplatin is a widely used and effective drug for the treatment of solid tumors ranging from ovarian, lung, head, and neck to testicular cancer. Dose limiting side effects, such as ototoxicity, neurotoxicity, and nephrotoxicity, are generally encountered with a majority of patients treated with cisplatin-based chemotherapy. Methods to increase diuresis, such as hydration, have been shown to reduce nephrotoxicity. However, to date no effective FDA approved treatment for ototoxicity is available. Cisplatin-induced hearing loss is primarily in the high frequency range. It is bilateral and permanent and severely affects the quality of life for cancer patients. The incidence of cisplatin-induced hearing loss in children ranges from 22 to 77% . This raSeveral approaches have been undertaken over the past two decades to treat cisplatin ototoxicity. These include the use of localized or systemic administration of antioxidants or drugs which activate endogenous antioxidant systems. Cisplatin induces apoptosis of hair cells through activation of mitochondrial pathway which can be targeted to inhibit cell death. Another approach involves using anti-inflammatory agents which target the pro-inflammatory mechanisms associated with cisplatin treatment in the cochlea. Recent insights into the entry processes of cisplatin in hair cells and other cells in the cochlea should stimulate the development of drugs which specifically block entry of drugs into these cells without affecting cisplatin entry into cancer cells. This report reviews the currently accepted mechanisms underlying cisplatin-induced damage or death to cochlear cells and highlights how these mechanisms could guide the future development of effective otoprotective agents.2) to form superoxide . Environmental stimuli which increase the metabolic activity of the cochlea, such as loud noise, are expected to increase oxidative stress in the cochlea. The metabolic demand on the cochlea renders it very sensitive to hypoxic events and ischemia-reperfusion injuries , GSH.Px, and catalase (CAT). SOD catalyzes the conversion of  to H2O2 and O2 while CAT converts H2O2 to O2 and H2O. GSH.Px reduces H2O2 and possibly other peroxides. The enzyme GSH.Px also catalyzes the conversion of reduced GSH to its oxidized form (GSSG), in the process of detoxifying H2O2. In addition, glutathione reductase (GR) is important for the defense against ROS by aiding in the regeneration of GSH from GSSG . Two forms of SOD are expressed in the cochlea. A Cu/Zn isoform of SOD is localized in the cytosol, while a Mn-regulated isoform (Mn-SOD) is localized to the mitochondria , heat shock proteins, kidney injury molecule-1, anti-apoptotic proteins, transcription factors such as nuclear factor erythroid 2-related factor 2 (Nrf2) and signal transducer and activator of transcription (STAT) proteins, and a number of hormone and G-protein-coupled receptors . TherefoN-acetyl cysteine (NAC) protected against cisplatin-ototoxicity in rats  in the stria vascularis, spiral ganglion cells, and organ of Corti . In vitro studies performed in HEI-OC1 cells demonstrate that cannabinoid 2 receptor (CB2) agonists reduce cisplatin-induced cell killing  RNA protected against cisplatin-induced hearing loss and damage to the outer hair cells . Protection was likely mediated by reducing the expression of a downstream target of TRPV1, such as NOX3, activation of which promotes ROS generation and STAT1 activation, as indicated above. STAT1 can promote both the inflammatory and pro-apoptotic actions of cisplatin in the cochlea  channels in cisplatin-mediated ototoxicity . In a rair cells  . These data indicate that cisplatin induces apoptosis through activation of mitochondrial pathway and that inhibiting elements of this pathway could alleviate hearing loss.Several studies have implicated the mitochondrial pathways in the apoptosis of auditory cells after cisplatin treatment. Mongolian gerbils administered cisplatin showed deterioration in the responses of both distortion product otoacoustic emissions (DPOAE) and the endocochlear potential as compared with age-matched controls . The cisf Bcl-xL . Treatmed Bcl-xL  and protStress signals from the mitochondria also regulate the cleavage of pro-caspase-9 into its active form, caspase-9 . ActivatSeveral mechanisms underlying cisplatin-induced caspase activation has been proposed. For example, cisplatin-induced apoptosis and caspase-3 activation in HEI/OC1 cells was reduced after treatment with NF-\u03baB inhibitors, Bay 11-7085 and SN-50 . This fiThe general mechanism of action of cisplatin in tumor cells is that it forms crosslinks with the purine bases of the DNA, causing DNA damage. The overwhelming DNA damage caused by formation of DNA adducts interferes with the DNA replication and repair mechanisms, which subsequently induces apoptosis . The apop53, caspase-1, and caspase-3  and group B (Csb-/-) mice were hypersensitive to cisplatin. In contrast, Xpc-/- mice which were deficient in global genome repair enzymes showed normal sensitivity to cisplatin  genes and xeroderma pigmentosum complementary group C and A (XPC and XPA) genes were assessed in patients with osteosarcoma who were treated with cisplatin. These studies showed that polymorphisms in DNA repair gene, XPC, was associated with increased cisplatin-induced ototoxicity in cancer patients. Increased ototoxicity was associated with the CC genotype of XPC Lys939Gln . Ctr1 is highly expressed in the cochlea where it is localized to outer hair cells, inner hair cells, stria vascularis, and spiral ganglion neurons and contributes to drug entry and cell apoptosis. Decreasing cisplatin entry by intra-tympanic administration of copper sulfate, a substrate of Ctr1, protects against hearing loss induced by cisplatin . Cisplatchildren . Severalchildren . In factchildren . Cisplatchildren . Recent children . These ichildren . Prelimichildren . These schildren . These cchildren . The potmegalin gene polymorphisms in susceptibility to cisplatin ototoxicity . These investigators show that the accumulation of cisplatin in the renal proximal tubules is mediated by binding of cisplatin-metallothionein complex to megalin . These studies need to be extended in human clinical trials for final validation. The routes of drug administration would be a major issue, as systemic administration of these drugs could compromise cisplatin chemotherapeutic efficacy. In this regard, the use of a number of cisplatin transporters (namely Ctr1 and OCT2) and antioxidants would have the greatest likelihood of interfering with cisplatin antitumor efficacy (as discussed above). Localized delivery of these drugs into the cochlea would eliminate potential systemic toxicities but would require an additional minor surgical procedure to accomplish this goal. This would allow the use of a majority of the agents listed in this table for protection against hearing loss. Several of these drugs are currently in clinical use or in clinical trials for other indications. These include TNF-\u03b1 antagonists which are used for the treatment of chronic inflammatory diseases (Studies described above have identified a number of different mechanisms which mediated cisplatin ototoxicity and provide the basis for rational drug design to treat this debilitating condition. While most of the drugs which target these mechanisms have shown promise, efficacy studies are still in the experimental animal stage (see diseases  and EGCGdiseases . These adiseases . ContinuSS and VR conceived and together outlined this review. SS and VR wrote the manuscript. DM and LR critiqued and revised the manuscript.The 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."}
+{"text": "Cisplatin is a widely used chemotherapeutic agent for treatment of various cancers. However, treatment with cisplatin is associated with drug resistance and several adverse side effects such as nephrotoxicity, reduced immunity towards infections and hearing loss. A Combination of cisplatin with other drugs is an approach to overcome drug resistance and reduce toxicity. The combination therapy also results in increased sensitivity of cisplatin towards cancer cells. The mitogen activated protein kinase (MAPK) pathway in the cell, consisting of extracellular signal regulated kinase, c-Jun N-terminal kinase, p38 kinases, and downstream mediator p90 ribosomal s6 kinase (RSK); is responsible for the regulation of various cellular events including cell survival, cell proliferation, cell cycle progression, cell migration and protein translation. This review article demonstrates the role of MAPK pathway in cisplatin based therapy, illustrates different combination therapy involving cisplatin and also shows the importance of targeting MAPK family, particularly RSK, to achieve increased anticancer effect and overcome drug resistance when combined with cisplatin. Cisplatin has been widely used since its approval in 1978 against a wide spectrum of tumors including lung, ovarian, testicular, bladder, colorectal and head and neck cancers . AlthougCisplatin is the first platinum drug approved globally and has been used as a drug for cancer chemotherapy for more than 30\u00a0years . CytostaUpon cisplatin entry to the cell, it becomes activated by hydrolysis. Chloride ions are displaced by water molecules resulting in a strong positive electrophile that covalently binds to any nucleophile such as nitrogen in purine residues. The most reported reaction is the 1,2-intrastrand cross-linkage of the activated cisplatin with the purine residues at nitrogen (N7) which was demonstrated in vitro in DNA of cisplatin-treated salmon sperm cells. These results were further approved in another in vivo experiment that analyzed white blood cells of cancer patients , 16. CisAlthough cisplatin is widely used anticancer drug, significant challenges still remain in regards to cisplatin resistance and cisplatin induced toxicity. There are different mechanisms attributed towards tumor resistance to cisplatin.Cisplatin is introduced into cells either through passive diffusion or via transporters  . PreviouTreatment with cisplatin has been associated with several toxic side effects including nephrotoxicity, oxidative damage to liver, cardiomyopathy, allergic reactions, ototoxicity and gastrotoxicity , 24\u201326.Cisplatin resistance to tumor cells can be overcome by adopting techniques Fig.\u00a0 to improOwing to treatment resistance and toxic side effects of cisplatin, implementation of combination therapy has gained much importance with regards to increased sensitivity of cisplatin towards cancer cells when combined with other drugs. The combination of other drugs with cisplatin may result in increased anticancer effect, there by result in reducing the dose of cisplatin and ultimately reduce toxic side effects. Hence, combination therapy may be adopted to evade cisplatin toxicity and resistance to cancer cells.To overcome the occurrence of resistance observed in patients relapsing from cisplatin treatment, cisplatin is commonly used in combination with various chemotherapeutic drugs and is the basis of many cancer treatments . CisplatPaclitaxel, a pro-apoptotic and mitotic agent, is used to treat numerous cancers including ovarian, lung and breast . PaclitaThe combination of doxorubicin and cisplatin has been demonstrated to be effective and well tolerated in cancer patients . In vitrIn aims to enhance the cytotoxic effects of interferon, a group of natural cytokines displaying antineoplastic effects, specifically in advanced hepatocellular carcinoma , interfeCisplatin and capecitabine in combination have been demonstrated in gastrointestinal cancers and have shown promising results , 42. HowMany recent studies reported that combination therapy using natural products, in particular Chinese herbal medicines, is a beneficial therapeutic strategy to overcome the side effects and drug resistance of cisplatin \u201346. NatuThrough the advancements made in our knowledge of tumor biology, the vital role of targeted therapies as either first or second line treatment regimens for cancer has been established. The plethora of data accumulated, especially over the last few years, has suggested the promising potential of targeted therapeutic agents use in combination to enhance and support the anti-tumoral effects of conventional chemotherapies \u201350.Such studies include combination therapy of cisplatin with C-75, which is a selective inhibitor of Fatty Acid Synthase (FASN) . FASN isA recent study demonstrated that silencing of YB-1 protein, expressed in various forms of malignant tumors, sensitized SH-SY5Y neuroblastoma cells to cisplatin and stimulated cisplatin induced apoptosis by down regulating multidrug resistance (MDR)1 protein through NFkB signaling pathway .c-fos, c-jun, bcl-2, c-myc, H-ras etc. resulting in improved activity of cisplatin against cancer cells [Inactivation of oncogenes responsible for cisplatin resistance is indeed a strategy to improve sensitivity of cisplatin towards tumor cells. In vitro approaches using antisense oligo deoxy nucleotides and by ribozymes (RNAs that have the ability of ligation at particular sites) successfully led to the inactivation of target genes like er cells .An example of combination therapy includes delta-tocotrienol with cisplatin where the combination resulted in downregulation of Notch-1 via inhibition of NFkB signaling pathways resulting in controlling non-small cell lung cancer, while reducing the effective dose of cisplatin  suggestiMajor survival pathways and their aberrant activation have been reported to frequently occur and contribute towards the development and progression of tumor growth, including the PI3K (phosphatidylinositol 3-kinase)/AKT (protein kinase B)/mTOR  signaling pathway \u201359. NumePrevious studies explored the in vitro effects of mTOR inactivation and evaluated the using of Torin2, an mTOR inhibitor, in EOC cell lines. Sub toxic doses of Torin2 potentiated cisplatin-induced apoptotic activity in EOC. In vivo studies also revealed that in combination of Torin2 and cisplatin, tumor growth in nude mice was synergistically inhibited. Overall, such studies highlight the importance and potential of targeting the mTOR survival pathway, suggesting that co-treatment of cisplatin and mTOR inhibitors, such as Torin2, may be beneficial for the management of EOC and various other resistant cancers .Previous studies have demonstrated that Akt inhibitor, MK-2206, sensitizes cisplatin towards gastric cancer cell line AGS. MTT assay and apoptosis assay demonstrated that cisplatin followed by MK-2206 resulted in synergistic effect of proliferative inhibition and apoptosis respectively for the combination treatment when compared to the monotherapy. The combination treatment also resulted in cleavage of PARP contributing to apoptosis . The AktMAPKs are a highly conserved family of structurally related serine/threonine protein kinases responsible for coordinating a variety of extracellular signaling pathways, regulating fundamental cellular processes involved in cell growth and survival , 9, 66. Bax (promotes apoptosis) and repressing transcription of Bcl-2 (inhibits apoptosis) [Although previous findings have identified cisplatin to result in ERK activation in several forms of cancer, whether this activation prevents or contributes to cisplatin-induced apoptotic effects remains greatly controversial \u201375. Cispoptosis) .Fig.\u00a02Ilcis and trans forms of cisplatin. DNA damage consequently leads to the stabilization and activation of p73, a pro-apoptotic protein related to p53, forming a complex with JNK, mediating apoptosis , Raf and Ras. Recent studies have reported the combination of MEK inhibitors along with cisplatin in in vitro and clinical trials , 85. MEKThe RSK proteins are the downstream mediators of the Ras\u2013ERK signal transduction pathway. ERK-mediated phosphorylation/activation of RSK plays an important role in regulation of protein translation, cell cycle progression and migration , 12, 13.In humans, four RSK isoforms (RSK 1\u20134) and two structurally related homologs have been identified . AlthougIt is proposed that targeting anti-apoptotic signals associated with the promotion of cell survival may be a promising strategy to optimize the effectiveness of conventional chemotherapeutic agents. RSK mediated signaling involves the phosphorylation and inactivation of pro-apoptotic proteins and the activation of transcription factors and translational machinery that promote cell survival. RSK1 and RSK2 activate pro-survival proteins Bcl-2 and Bcl-xL by post translational phosphorylation and inactivation of the pro-apoptotic protein Bad, enhancing its ability to bind 14-3-3 proteins and preventing its heterodimerization with the pro-survival proteins. RSK1 and RSK2 also phosphorylate CREB leading to increased transcription of Bcl-2 and promote cell survival. Also, RSK1 directly inhibits caspase activity, promoting cell survival. RSK phosphorylates and inhibits GSK3\u03b2 to promote stabilization of cyclin D1 resulting in cell cycle progression and cell-survival . PreviouThis review article depicted the importance of combination therapy in treatment of cancer. Standard chemotherapy with cisplatin has limitations, such as drug resistance and its toxic side effects. However, these limitations can be overcome by administering low doses of cisplatin, which can be achieved in combination with such agents which would increase the sensitivity of cisplatin towards cancer cells. Studies conducted in recent years suggested the importance of administering targeted therapeutic agents in combination with cisplatin to enhance the anti-tumoral effects of cisplatin. It would be beneficial to target the major cell survival pathways that are upregulated in various types of cancers. The MAPK family, including its members ERK, p38, JNK and downstream mediator, RSK, executes major cellular functions like proliferation, survival and differentiation. However, the inhibition of ERK, p38 and JNK either prevents or contributes towards cisplatin induced apoptosis remains controversial; which necessitates focusing on inhibition of RSKs which are the downstream effectors of the ERK/MAPK signaling cascade. RSKs are implicated in various cellular processes and modulate cell proliferation, tumorigenesis and metastasis in various cancers. Therefore, as shown in Fig.\u00a0Although this review focused on cisplatin, the findings discussed here are not only specific to cisplatin but also applicable to a class of platinum derived chemotherapeutic drugs. Platinum derived agents like carboplatin and oxaliplatin are other globally approved anticancer drugs and have similar mechanism of action as cisplatin , 2. CarbIn conclusion, targeting MAPK pathway predominantly in its downstream signaling cascade would be favorable in cancer treatment. A combination therapy of platinum based drugs with a RSK inhibitor would be a beneficial strategy to overcome the resistance and toxic side effects of platinum drugs and ultimately to combat cancer."}
+{"text": "La myosite ossifiante circonscrite (MOC) est une ossification h\u00e9t\u00e9rotopique des muscles stri\u00e9s. Sa localisation au niveau du coude est rare. Elle survient chez le sujet jeune, souvent suite \u00e0 un traumatisme comme elle peut se d\u00e9velopper \u00e9galement en dehors de tout traumatisme. Sa localisation pr\u00e9dominante est au niveau des muscles les plus volumineux de la racine des membres  ou les plus expos\u00e9s aux chocs direct (plus de 40 % des cas post-traumatiques sur le quadriceps). Nous proposons d\u2019illustrer \u00e0 partir d\u2019une observation, les aspects que r\u00e9alise la myosite ossifiante circonscrite en radiologie conventionnelle, en tomodensitom\u00e9trie afin d\u2019\u00e9viter la confusion diagnostique potentielle avec une tumeur osseuse maligne. La myosite ossifiante circonscrite (MOC) est une ossification h\u00e9t\u00e9rotopique des muscles stri\u00e9s. C'est une affection b\u00e9nigne pseudo-tumorale, rare, d'\u00e9tiologie assez mal d\u00e9finie. C'est un processus local r\u00e9actif, de d\u00e9veloppement auto limit\u00e9, qui se produit \u00e0 partir du tissu conjonctif interstitiel et non pas des fibres musculaire stri\u00e9es squelettiques. Le tableau clinique et la biologie ne sont pas sp\u00e9cifiques, ce qui rend \u00e0 l'imagerie un r\u00f4le important dans le diagnostic de la maladie. Ce fut le cas de notre patient dont nous rapportons l'observation.3). Nous avons compl\u00e9t\u00e9 le bilan par une tomodensitom\u00e9trie du coude qui a parl\u00e9e de formations de densit\u00e9 calcique s'\u00e9tend de la face post\u00e9rieure du coude jusqu'au niveau de la partie moyenne du bras  est une pathologie b\u00e9nigne, rare des parties molles, caract\u00e9ris\u00e9e par une prolif\u00e9ration h\u00e9t\u00e9rotopique non n\u00e9oplasique d'os et de cartilage dans les tissus mous \u00e0 distance du p\u00e9rioste. Cette affection est retrouv\u00e9e avec une fr\u00e9quence sensiblement \u00e9gale dans les deux sexes, et concerne le sujet jeune jusqu\u00b4\u00e0 30 ans. La notion de traumatisme violent est parfois retrouv\u00e9e mais souvent il s'agit de microtraumatismes r\u00e9p\u00e9t\u00e9s . L'atteiLa MOC est une affection b\u00e9nigne des parties molles, dont l'\u00e9tiologie et dont l'\u00e9volution est constamment favorable. Le tableau clinique et paraclinique initiale de la MOC fait craindre un processus malin ce qui peut amener parfois a une biopsie chirurgicale. Le traitement reste essentiellement m\u00e9dical, il est rarement chirurgical."}
+{"text": "Galaxea (G. fascicularis) and Montipora  collected from five different locations in the South China Sea using massively-parallel sequencing of 16S rRNA gene and multivariate analysis. The results indicated that microbiome assemblages for each coral species were unique regardless of location and were different from the corresponding seawater. Host type appeared to drive the\u00a0coral microbiome assemblages rather than location and seawater. Network analysis was employed to explore coral microbiome co-occurrence patterns, which revealed 61 and 80 co-occurring microbial species assembling the Galaxea and Montipora microbiomes, respectively. Most of these co-occurring microbial species were commonly found in corals and were\u00a0inferred to play potential roles in host nutrient metabolism; carbon, nitrogen, sulfur cycles; host detoxification; and climate change. These findings suggest that the co-occurring microbial species explored might be essential to maintain the critical coral-microbial partnership. The present study provides new insights into coral microbiome assemblages in the South China Sea.Coral reefs are significant ecosystems. The ecological success of coral reefs relies on not only coral-algal symbiosis but also coral-microbial partnership. However, microbiome assemblages in the South China Sea corals remain largely unexplored. Here, we compared the microbiome assemblages of reef-building corals  However, coral reefs around the world have suffered from declines and extinction risks largely due to bleaching events and emerging/reemerging diseases induced by climate change and anthropogenic disturbances4. The ecological success of coral reefs relies on coral-algal symbiosis, and recent studies using 16S rRNA gene amplicon pyrosequencing have revealed highly diverse and abundant microbes in individual coral colonies9. It is believed that some of these microbes can form partnerships with coral hosts and help them with possible access to those unavailable nutrients and metabolic pathways10. Compared with the well-understood coral-algal symbiosis, current understanding of coral-microbial partnership is rather limited. Although members of Endozoicomonas6 and Prosthecochloris11 have been shown to form potential symbioses with corals, no obligate coral-microbial symbiosis has been addressed to date. A recent advance in calcification for coral symbiotic algae demonstrated that free-living Symbiodinium in culture could form algal-microbial partnership, which facilitated the Symbiodinium calcification12. Taken these evidences, potentially complex coral-algal-microbial interactions in holobionts might facilitate the ecological success of coral reefs.Coral reef ecosystems are considered\u00a0as the tropical rainforests of the sea, nurturing the highest biodiversity of marine life and providing vital ecosystem goods and servicesEukarya, Archaea, and Bacteria, as well as many viruses14. The term coral microbiome is employed herein to collectively refer to the bacteria and archaea in coral holobionts. The characterization of coral core microbiomes reveals that two most abundant bacterial phylotypes affiliating to the genera Propionibacterium and Ralstonia are co-localized specifically with the host\u2019s endosymbiotic algae which are likely to facilitate the success of coral-algal symbiosis10. Coral microbiomes are highly complex and dynamic, usually changing with environmental conditions, host types, and tempo-spatial gradients15. However, we assumed that there would be co-occurring microbial species assembling coral microbiomes which might be fundamental for coral holobionts to maintain the critical coral-microbial partnership. To explore these potential microbial species, co-occurrence network analysis was conducted in this study. Network-based approaches have been widely applied in microbial ecology studies16 such as soil17, coral18, human gut19, marine sediment20, stream biofilm21, activated sludge22, marine biofilm23, and other natural and man-made environments.The coral mucus, tissue, and skeleton all contain large populations of associated microbes from three domains of life, including 28, but very limited knowledge has been gained in understanding the microbiome assemblages for the South China Sea corals. We do not yet clearly understand how coral microbiomes are assembled in the South China Sea and potential co-occurring microbial species making up the coral microbiomes. The aims of this study were (i) to compare the microbiome assemblages using congeneric corals collected from different locations of the South China Sea, and (ii) to explore potential co-occurring microbial species through co-occurrence network analysis. We believe the present study facilitates our understanding of coral-microbial partnership in the South China Sea and serves as a useful reference for comparison with other regional coral microbiome assemblages.In recent years, there has been a growing interest in coral microbiome studiesProteobacteria dominated all of the coral and seawater microbiome assemblages with a relative abundance greater than 50%, among which Alphaproteobacteria or Gammaproteobacteria accounted for the largest proportion , and the Montipora microbiome assemblages were also significantly different among locations LI, CB, LHT, and SB . To further explore the differences in coral microbiome assemblages between host type, location, and seawater comprehensively, thirteen groups  were generated for pairwise comparisons using one-way PERMANOVA. As shown in Table\u00a0P\u2009<\u20090.05, 1.96\u2009<\u2009F\u2009<\u200922.44). To explore the similarities among these thirteen groups, NMDS (stress\u2009=\u20090.13) was conducted and shown in Fig.\u00a034. The eight coral groups were loosely clustered together with four Galaxea groups and four Montipora groups distributed at the top and bottom respectively, indicating that host type exhibited a consistent distribution pattern. However, there was no consistent geographic pattern in distribution for both Galaxea and Montipora groups, suggesting that location contributed less in driving the coral microbiome assemblages. According to the NMDS result, host type but not location showed a consistent distribution pattern for the\u00a0coral microbiome assemblages and the coral and seawater microbiomes were assembled dissimilarly. Therefore host type is suggested to play an even more important role than location and seawater in driving coral microbiome assemblages.The recovered 97% OTU data were used for statistical analyses to examine the differences (PERMANOVA) and similarities (NMDS) in coral microbiome assemblages between host type, location, and seawater. One-way PERMANOVA revealed that the Galaxea and Montipora microbiomes were used for co-occurrence network analysis after removal of the poorly represented OTUs, respectively. We first tested the two OTU datasets and observed non-random co-occurrence patterns, which is consistent with other studies22. A study even demonstrated that non-random community assemblage might be a common feature of all life domains35. As shown in Fig.\u00a0Galaxea and Montipora microbiomes, respectively. The network topological parameters for the Galaxea microbiomes included an average node connectivity of 2.39, an average network distance of 2.18 edges, a network diameter of 6 edges, an average clustering coefficient of 0.37, and a modularity index of 0.73. While the network topological parameters for the Montipora microbiomes contained an average node connectivity of 4.30, an average network distance of 2.76 edges, a network diameter of 6 edges, an average clustering coefficient of 0.48, and a modularity index of 0.62. These topological properties were used to describe the general structures of co-occurrence networks17. Both networks of the Galaxea and Montipora microbiomes had modular structures because each modularity index value was higher than 0.4036. Based on the modularity class, each network could be further divided into several sub-networks, for example, those closely interacted nodes on the left side of each network , Betaproteobacteria (1), Gammaproteobacteria (23), Bacteroidetes (5), Cyanobacteria (3), Thaumarchaeota (2), and Euryarchaeota (1). While the 80 co-occurring microbial species assembling the Montipora microbiomes were classified into Alphaproteobacteria (39), Betaproteobacteria (2), Gammaproteobacteria (27), Bacteroidetes (1), Cyanobacteria (7), Thaumarchaeota (1), Nitrospirae (1), Chloroflexi (1), and Fusabacteria (1). The relative abundance of the 114 co-occurring microbial species and the corresponding seawater samples is shown using a heat map , Betaproteobacteria (1), Gammaproteobacteria (13), Cyanobacteria (1), and Thaumarchaeota (1). These shared co-occurring microbial species included 7 unknown spp., 6 Acinetobacter spp., 3 Endozoicomonas spp., and a species from each of the following 11 genera: Brevundimonas, Brucella, Escherichia, Filomicrobium, Methyloceanibacter, Methylophilus, Nitrosopumilus, Pelagibius, Roseibium, Thalassomonas, and Thioprofundum. These shared co-occurring microbial species assembling the Galaxea and Montipora microbiomes might be prevalent in other corals from the South China Sea.There were 27 shared co-occurring microbial species assembling the  Figures\u00a0 and S3. 37, while the counterpart \u201cstable microbiome\u201d has also been used in related studies38. According to the statistical methods used for data analysis, coral-associated microbes can be assigned into \u201ccore/stable microbiome\u201d and \u201ctransient/sporadic microbiome\u201d37. The \u201ccore microbiome\u201d is described as the stable and consistent components across complex microbiome assemblages from similar habitats which can be determined with one of the five described variations, that is, a core based on shared presence, shared abundance, shared composition, phylogenetic information, or interaction39. Co-occurring microbial species explored here through co-occurrence network analysis are exactly equal to the \u201ccore microbiome\u201d based on interaction, as illustrated in related studies39. In the present study, we collected coral samples with a large set of biological replicates from the five locations in the South China Sea, allowing us to compare the complex microbiome assemblages comprehensively and to explore co-occurrence patterns through network analysis. We further discussed the effects of host type and location in coral microbiome assemblages (the first section below) and potential roles of co-occurring microbial species explored including those functional microbes (the second section below) and those mostly found microbes (the third section below).In recent years, the concept of \u201ccore microbiome\u201d has been introduced in coral microbial ecology studies15. The study on Red Sea corals reveals that the microbiome assemblages vary largely with location and are shaped by host type5. Certain microbes are reported to form species-specific associations with corals40, suggesting the roles of host played in driving coral microbiome assemblages. The study on corals from the Great Barrier Reef shows that certain microbes are associated with corals specifically and that microbiome assemblages differ with the location but not the host type41. These findings support the present study that host type rather than location drives the coral microbiome assemblages, as shown in Fig.\u00a0Montipora were collected for the present study because the shared species were not found in the selected locations except for Galaxea fascicularis. The study using three species of the genus Acropora found that they harbored conserved microbes and suggested that closely related corals of the same genus were assembled with similar microbes41. So it can be assumed that if one of the three studied Montipora species were shared and used for the present study, the general distribution pattern of microbiome assemblages for this species from different locations might be less likely affected in the NMDS ordination. The distribution pattern of the three Montipora species shown in Fig.\u00a0Montipora monasteriata were clustered closely and were relatively distinct from LI-Mo and CB-Mo (from Montipora venosa and Montipora peltiformis respectively).Corals harbor highly diverse microbes that might be assembled similarly or dissimilarly among host types, locations and other potential determinants. The study on Caribbean corals shows that different hosts harbor distinct host-specific microbes and that specificity varies by host type and locationBrevundimonas spp. and Phyllobacterium spp. might fix atmospheric nitrogen gas to ammonia. Potential ammonia-oxidizing archaea like Nitrosopumilus spp. might oxidize the holobiont ammonia to nitrite, and possible nitrite-oxidizing bacteria like Nitrospira sp. might oxidize the holobiont nitrite to nitrate. Finally, potential denitrifying bacteria such as Methylophilus sp. and Nitratireductor sp. might convert the holobiont nitrate to nitrogen gas. These co-occurring microbial species might thus be involved in the complete nitrogen cycle and changes in their population and activity might affect certain process of the nitrogen metabolism in coral holobionts42. It is suggested that they might serve as nitrogen regulators to keep the balance of holobiont nitrogen through providing sufficient bioavailable nitrogen and removing unneeded nitrogen. Carbon dioxide might be fixed into carbohydrates by Symbiodinium and Cyanobacteria through photosynthesis and by the co-occurring microbial species explored such as Nitrosopumilus spp., Thioalkalivibrio sp., and Thioprofundum spp. through chemosynthesis. Like Symbiodinium, the synthesized carbohydrates by the co-occurring microbial species might also serve as the host nutrients or food sources. The study has demonstrated that free-living Symbiodinium can calcify with the aid of microbial partners12, implying that potential co-occurring microbial species explored might also facilitate coral calcification directly or indirectly. Hydrogen sulfide is toxic to a wide range of eukaryotic organisms, including marine invertebrates like coral and sponge, by inhibiting cytochrome c oxidase and a type of catalase44. In addition, hydrogen sulfide generated by sulfate-reducing bacteria might lead to the initiation of coral black band disease, which has been consistently found in lab induction and field observation46. Co-occurring microbial species Thioalkalivibrio sp. and Thioprofundum spp., serving as potential sulfur-oxidizing bacteria, might oxidize holobiont accumulated hydrogen sulfide to sulfate, thus contributing to coral health through detoxification of reduced sulfur compounds. Both DMSP (dimethylsulfoniopropionate) and DMS (dimethylsulfide) are important compounds in the global sulfur cycle as they are closely related to cloud formation and global climate change47. Both reef-building corals and free-living Symbiodinium have been documented to be high producers of DMSP49. The generated DMSP might be degraded into climate-active gas DMS via the bacterial cleavage pathway by co-occurring microbial species explored such as Hoeflea sp., Loktanella sp., Phaeobacter sp., Roseovarius sp., Shewanella sp., and Vibrio sp.47. In summary, co-occurring microbial species explored might play potential roles in host nutrient metabolism; carbon, nitrogen, sulfur cycles; host detoxification; and climate change and might be essential to maintain the critical coral-microbial partnership. However, the real functions need to be tested in future studies. For example, microbial genome recovery using culture-independent methods such as genome binning and single-cell genomics is promising to validate the specific functions on a genome scale.To achieve a better understanding of co-occurring microbial species, potential ecological roles and associations with coral hosts were further explored. As shown in Fig.\u00a0Galaxea and Montipora microbiomes, 16 Acinetobacter spp. and 10 Endozoicomonas spp. were found, of which 6 Acinetobacter spp. and 3 Endozoicomonas spp. were shared between the two corals. This finding corroborates the high occurrence of Acinetobacter spp. and Endozoicomonas spp. in coral microbiomes found in different coral species and regions  and ground in 1\u00d7\u2009PBSE using a mortar & pestle. The resulting coral slurry was centrifuged at 12,000\u2009g for 5\u2009min to collect all pellets for total DNA extraction using the FastDNA\u00ae Spin Kit for Soil . The DNA extracts were used as PCR templates after quality and purity determination through agarose gel electrophoresis and OD260/OD280 ratio measurement. To amplify and sequence the hypervariable V3 and V4 regions of the 16S rRNA gene, a universal primer set, 341\u2009F (5\u2032-CCTAYGGGRBGCASCAG-3\u2032) and 802\u2009R (5\u2032-TACNVGGGTATCTAATCC-3\u2032), was used for PCR amplification70. Unique barcodes of six nucleotides were modified at the 5\u2032 terminus of the forward primer 341\u2009F to allow multiplexed sequencing71. PCR was conducted using the following program: initial denaturation at 94\u2009\u00b0C for 5\u2009min, followed by 30 cycles at 94\u2009\u00b0C for 0.5\u2009min, 50\u2009\u00b0C for 0.5\u2009min, 72\u2009\u00b0C for 1\u2009min, and a final extension at 72\u2009\u00b0C for 5\u2009min. Each PCR reaction was pooled with ~50 ng DNA template, 200\u2009nM of each primer, 25\u2009\u03bcL 2\u00d7\u2009Premix Ex Taq solution , and ddH2O up to 50\u2009\u03bcL. To minimize potential PCR bias, three independent reactions were made for each sample and the products were pooled for purification using the PureLink\u00ae PCR Purification Kit . The purified PCR products were quantified using a Thermo NanoDrop 2000 UV-Vis Spectrophotometer and pooled at equivalent concentrations for subsequent multiplexed sequencing of\u00a016S amplicons. The pooled DNA sample was sent to Novogene  for sequencing on an Illumina MiSeq sequencer with a paired-end (PE) mode and a read length of 300\u2009bp. Because the length of the 16S V3V4 fragment is approximately 450\u2009bp, paired-end 300\u2009bp sequencing is sufficient to form overlaps for full-length amplicon assembly73. The sequencing datasets have been deposited in the NCBI Sequence Read Archive under accession number SRP066229. Totally, there were 59 samples sequenced for the five locations LI, CB, LHT, SB, and DI , CB (5), LHT (4), and SB (5); 2 additional Montipora technical replicates for the location of LHT, and 2\u20133 technical replicates of seawater samples for locations of LI (2), CB (2), LHT (3), SB (3), and DI (3). There were four Montipora samples failed in the DNA extraction or PCR amplification, that is, each one from the locations of CB and SB and two from the location of LHT.A small fraction (~0.5\u2009cm\u2009\u00d7\u20090.5\u2009cm) of each coral specimen fixed in ethanol was first rinsed with 1\u00d7\u2009PBSE  platform75 was used to demultiplex 16S tags into individual samples identified by unique barcodes. Potential PCR chimeras were detected and removed using the ChimeraSlayer76 command in the QIIME. After strict data trimming, the clean 16S tags were recovered and normalized for the downstream analysis. OTUs  at 97% similarity  were clustered and annotated using the QIIME pipeline under default settings. Relative abundance data for community structures were automatically generated from phylum to genus. A 97% OTU matrix was recovered from the generated OTU table of biom file format, and the data were used for species-level permutational multivariate analysis of variance (PERMANOVA), non-metric multidimensional scaling (NMDS), and network analysis. Those poorly represented OTUs (mean relative abundance lower than 0.1%) were removed before the network analysis as previously described17. Briefly, microbial species-level co-occurrence networks were conducted in the R77 environment using the vegan78, igraph79, and Hmisc80 packages. Positive co-occurrence events were considered if the OTUs demonstrated the Spearman\u2019s correlation coefficient higher than 0.6 and statistically significant (P\u2009<\u20090.01)21. The constructed networks were further optimized by the interactive visualization platform Gephi81. The R software was also employed for the heat map plotting to visualize profiles of the co-occurring microbial species in each sample. MEGA 6.0682 was used to construct the phylogenetic tree of 16S rRNA gene. PAST 3.1083 was used for PERMANOVA, NMDS and cluster analysis based on the Bray-Curtis distance. OriginPro 9.0 was employed for interactive scientific graphing. PICRUSt (Phylogenetic Investigation of Communities by Reconstruction of Unobserved States) prediction was conducted following the established pipeline84.The raw data were first trimmed to remove sequencing adaptors, short reads (<300\u2009bp), and low quality reads . To obtain the full-length of 16S V3V4 sequences, the PEAR (paired-end read merger) toolSupplementary Information"}
+{"text": "Designers of undergraduate medical education (UME) need to address the exponentially expanding volume and variability of scientific knowledge, where by didactic teaching techniques need to be augmented by innovative student-centric pedagogical strategies and implementation of milieus, where information, communication and technology-enabled tools are seamlessly integrated, and lifelong information gathering, assimilation, integration and implementation is the ultimate goal. In UME, the basic sciences provide a solid scaffold allowing students to develop their personal critical decisional framework as well as define the understanding of normal human physiology, pivotal for the identification, categorization and management of pathophysiology. However, most medical schools confine themselves to \u201cstagnant curricula\u201d, with the implementation of traditional \u201cteacher centered\u201d pedagogical techniques in the dissemination of the courses pertaining to basic sciences in UME.To tackle the above paucity, we present a novel \u201c6D-Approach\u201d for the dissemination of concepts in basic sciences through mentored journal-clubs. The approach is informed by a teaching principle derived from Constructivism. The technique in which the 6D-approach can be implemented in UME, is shown using an example from a first-year course of Molecular Biology and Principles of Genetics at our medical school. A reflection on the impact of 6D-Approach for students as well as instructors is also presented.The 6D-approach was positively received by the students and the formal feedback for the course: Molecular Biology and Principles of Genetics, where the approach was repeatedly employed, indicated that students expressed satisfaction with the teaching strategies employed in the course, with ~\u200989% of the students in the cohort strongly agreeing with the highest grading score \u201cextremely satisfied\u201d. Further, the approach through the use of mentored journal clubs encourages retention of knowledge, critical thinking, metacognition, collaboration and leadership skills in addition to self-evaluation and peer feedback.Hence, through the 6D-Approach, our attempt is to initiate, advance and facilitate critical thinking, problem-solving and self-learning in UME, demonstrated by graduating accomplished, competent and safe medical practitioners. The complexity of twenty-first century healthcare requires re-thinking current  educational paradigms. In the past emphasis was on gathering facts and understanding the context of these facts: knowledge, presented to students and trainees in textbooks, often taught by lecturers, in large classroom settings in which passive absorption of the shared knowledge was expected. Information or scientific knowledge has expanded at an exponential rate, requiring undergraduate curriculum planners to address the rapidly increasing Velocity, Variability and Volume of data , 2. DidaOver the last few decades, medical curricula are shifting towards a more integrative approach, offering activities that augment critical thinking and problem-solving skills into medical education, a need first foreseen by Abraham Flexner. Flexner advocated the concept that formal analytic reasoning, the kind of rational thinking fundamental to the basic sciences especially the natural sciences, should hold precedence in the intellectual training of physicians , 4. In aHowever, many medical schools globally continue to confine themselves to a so called \u201cstagnant curricula\u201d, with traditional \u201cteacher-centered\u201d pedagogical techniques and an apparent inability, or even resistance to change , 8 thereIn undergraduate medical education (UME), the conventional basic sciences facilitate the so-called ontogenesis of the medical practitioner. They aim to provide a solid scaffold allowing students to develop their personal critical decisional framework as well as define the understanding of normal human physiology, pivotal for the identification, categorization and management of pathophysiology .Further UME should, also encourage activities that augment critical thinking, problem-solving, metacognition, empathy and collaboration skills, to catalyze action and spark innovation. One approach has been that of journal clubs, a staple of continuing education, scholarly meetings in which individuals convene regularly to critically assess/appraise current/recent articles in the scientific literature . InitialCurrently, in UME, pedagogical techniques focus primarily on dissemination of knowledge, but few teaching stratagems aim at providing the medical student with an understanding of how this knowledge can be applied for practicing evidence-based medicine. Further, UME should also provide the foundation for the growth of non-cognitive and metacognitive skills, which is rarely addressed by the teaching strategies implemented in UME, specifically in the delivery of basic science courses. The overarching aim of this research is to design an innovative pedagogical strategy using journal clubs, whereby the student will be informed of the clinical applicability of disseminated knowledge, using published facts and data available in peer-reviewed literature. Such a strategy will also allow instructors to incorporate evidence-based medicine (EBM) teaching, especially in the delivery of basic science courses.In this article, we delineate the \u201c6D-Approach\u201d for the dissemination of concepts in basic sciences through mentored journal clubs. The 6-steps of the approach are shown in Fig.\u2009The significance of linking instructional approaches or strategies to the theories of human learning has been demonstrated by Onyura et al . BehavioIn order to develop a teaching principle that will inform an approach through which journal clubs can be implemented in UME, we employed Constructivist theory as the foundational core, with contributions from the principles of the Humanistic and Experiential Learning (ELT) theories Fig. b. The prThe philosophical roots of Constructivism are founded on Kantian epistemology where Empiricism  and Rationalism (learning happening through practice or principles of basing opinions and actions on reason and knowledge) are integrated . In termHowever, we perceived that to apply a teaching principle founded on Constructivism, a positive non-threatening setting is essential, where the student can \u201cthink\u201d and \u201creflect\u201d on the acquired knowledge, aiding learning autonomy and peer-assisted learning. Hence, in designing the principle, we drew from principles of Humanism  and ELT (learning through reflection on doing) , 23; TheThe acquirement and construction of knowledge is integrated more deeply by engaging in multiple viewpoints and depictions of content, amongst peers addressing a set of common learning outcomes.Didactics, Designate, Distribute, Design, Delivery and Discuss) are summarized in Fig.\u00a0Employing the derived teaching principle, we designed the 6D-approach. The six steps of 6D-approach  main learning objectives; employing effective visuals, analogies, demonstrations, and illustrations to reinforce the main points; making the material available to students prior to delivery; highlighting and summarizing the learning objectives and major points of the lecture clearly.http://www.nlm.nih.gov/bsd/disted/pubmedtutorial/and the PubMed help page http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=helppubmed&part=pubmedhelp.First, the instructor designates two to three small groups  could similarly be circulated using electronic-mail or shared over a cloud service, such as Google-Drive, OneDrive, iCloud etc.Following article selection by the groups, the selected article(s) are made available to other students in the cohort by uploading through the learning management system (LMS) software . This coAn alternative approach is to employ a social media application to host the journal club online, such as that described by Topf et al , 28.This step focuses on the design of presentation by the student groups. While implementing 6D-approach at MBRU, we recommended the students to follow the guidelines of Schmatlz et al., while designing their presentation .The designated student-group(s) delivers the presentation, during which a representative from the group may deliver the entire presentation or different sections of the presentation are delivered by different group members. While it is important that the instructor leave the choice of the presentation style to the students, groups are instructed to be mindful of time constraints and to ensure interactivity with other learners. At MBRU one of the students from the cohort volunteered as the timekeeper to make sure all presentations started and concluded within a given time-frame.Didactic step provided them with a suitable background to allow critical appraisal. While implementing the 6D-Appoach we encouraged the student who is moderating the discussion to have a list of initiating and follow-up questions to promote discussion.As in journal clubs at more advanced levels, it is important to ensure that students have time to discuss the article(s), evaluate its strength and weaknesses and to assess if the concepts delivered in I.Choosing the course: We decided to implement the 6D-approach at MBRU for which we selected the course: Molecular Biology and Principles of Genetics, offered in semester 2 of the first-year medical curriculum at MBRU.The reason we chose this course is, because with the rapid integration of molecular biology and genetics into medicine it has become evident that practicing physicians as well as medical students and clinical researchers, need to be updated on the new developments especially with important findings based on molecular-biology/genetics related biomedical research. The course provides a foundation for understanding the relationship between molecular biology, developmental biology, genetics, genomics, bioinformatics, and medicine over a period of 15\u2009weeks. One of the key aims of this course is to develop explicit associations between basic research, medical understanding, and the perspective of patients. After attending the course, the students should be able translate clinical understanding into analysis at the level of the gene, chromosome and molecule. Further, the topics in the course encompass concepts and techniques of molecular biology and genomics, and the strategies and methods of genetic analysis, including an introduction to bioinformatics.II.Logistics: The entire cohort consisting of 54 students registered for the course, and there were no dropouts over the duration of the course. Seven presentation sessions were organized throughout the course. Students were informed of the dates of presentation at prior to the commencement of the course. On the day of commencement of the course, a designated student representative was instructed to distribute the students into groups, each group containing 3 to 4 students.The representative divided all 54 students into 14 groups. The first 12 groups had 4 students in each group (12 X 4\u2009=\u200948 students) and the last 2 groups had 3 students in each group (2 X 3\u2009=\u20096 students). In each presentation session, 2 student groups delivered their presentation.III.Overview of a typical session: Each student group had 14-days, following the Didactic step (Step 1 of 6D-approach) to decide on the article and prepare for the presentation. Slides corresponding to each presentation were uploaded on the LMS and shared amongst the students at least a day prior to the presentation. Each student group was allotted a maximum time of 25\u2009min to deliver the presentation. This was followed by a question/answer or discussion session of 15\u2009min. As overview of the implementation of 6D approach for the topic titled \u201cDNA Replication and Repair\u201d is shown in Table\u00a0Didactic step focused on defined objectives which were delivered through two lecture sessions. At the end of the, second lecture session the instructor identified two topics for exploring the application of the concepts delivered.Group 1 was Designated to present two case reports with regards to genetic disorders where the DNA repair mechanisms are impaired. These included (A) Fanconi Anemia and (B) Xeroderma pigmentosum.Group 2 was Designated to review the topic \u201cExploitation of DNA repair mechanisms in cancer therapy\u201d.Distributed through LMS.The student articles were Each group Designed their presentations separately, with intermittent help and clarifications from the instructor. The designed presentations were then uploaded and shared among all students using LMS.Both Groups 1 and 2 Delivered their presentation on a pre-decided day, with other students and the instructor as the audience.Following the Delivery of the presentation a student-initiated Discussion session was organized, which critically appraised the articles. The Discussion step of the approach also explored the importance of the concepts delivered in the Didactic step, with regards to their clinical relevance. Case in point, in the Didactic step students were informed that nucleotide excision repair (NER) is the most flexible of all DNA repair mechanisms because the mechanisms ability to eliminate a multifarious structurally unrelated DNA lesions. However, the Discussion step of the approach elaborated that the common denominator of the different types of damage induced by the numerous chemicals to which NER-deficient cells are sensitive seems to be the generation of bulky base adducts which lead to a distortion in helical structure and DNA chemistry relating it to the diseased state Xeroderma pigmentosum.In the present work, our focus was on the design and implementation of the 6D-Approach, an elaborate evaluation of the outcome of the 6D-Approach is still pending and will be addressed in our future work through the design of defined rubrics and questionnaires, employing software module such as Survey Monkey. Hence, the evaluation of the 6D-Approach presented here is only preliminary.The approach was evaluated informally following Pendleton\u2019s approach . A measuThe 6D-apporach was received positively by the students, who exhibited enthusiasm in both organizing and in participating in the event. In each of the seven sessions the students were informally queried with regards to two questions: (A) What were the strengths of the session? (B) What aspects of the session they believed required attention/improvement? Along, with the students the instructor also addressed the above questions on site.Students from different academic backgrounds effectively functioned as a group.Reading habits of students improved significantly from the commencement to the conclusion of the course as was observed through the increase in the depth and content of the questions posed by the students during Discussion. This observation which is in line with the findings of Honey et al .Few points of note observed by the instructor during the sessions:The time allocated for the Discussion was insufficient in all seven sessions. Since these events are scheduled in line with a designated time-slot, one way to address this deficiency will be to host online discussion sessions over Twitter, Snap Chat, WhatsApp etc.Students had difficulty in accessing specific journals (as they weren\u2019t subscribed by the institution). One of the ways to bypass this limitation is to encourage students to refer to articles in open access journals, albeit of repute, as not all open access journals are stringently reviewed , 32.Specific limitations that need to be addressed:Formal student feedback for the course, like other courses offered during the semester, was obtained online, using a MBRU approved questionnaire identical for all courses. The formal feedback for Molecular Biology and Principles of Genetics indicated that students expressed satisfaction with the teaching strategies employed in the course, with ~\u200989% of the students in the cohort strongly agreeing with the highest grading score \u201cextremely satisfied\u201d.With regards to the facilitation of retention of knowledge following the implementation of the 6D-approach, we appraised the final grades of the students. The course had two-formative and one summative assessments. The pass mark and excellence for individual assessment was determined by Angoff-method . Out of The 6D-approach provides a novel strategy to improve UME. Under the following subheadings, we explore the different cognitive and non-cognitive skills that are beneficially affected, when this approach is implemented.Design and Delivery, elaborate rehearsal is facilitated, accelerating \u201cDeep-learning\u201d [Didactic step, in analyzing the details of the article as well as reflecting on the data presented in the article in light of the concepts delivered.  through the 6D approach. TEL can be defined as an innovative pedagogical approach, that aims to combine learning design and leverage digital technologies to deliver active and engaging, student centered learning . Our endDesigners of UME need to address the exponentially expanding volume and variability of scientific knowledge, where by didactic teaching methods need to be augmented by innovative student-centric pedagogical strategies. Such pedagogical strategies need to be informed by focused teaching principles, and should integrate information, communication and technology-enabled tools to promote lifelong information gathering, assimilation, integration. One of these approaches is to employ mentored journal clubs, which are scholarly congregations in which individuals convene regularly to critically assess/appraise current/recent articles in the scientific literature, especially in basic science courses in UME, as these courses provide a platform allowing students to develop their personal critical decisional framework as well as define the understanding of normal human physiology, pivotal for the identification, categorization and management of pathophysiology. Such journal club integrating pedagogical strategies will encourage retention of knowledge, critical thinking, metacognition, collaboration and leadership skills in addition to self-evaluation and peer feedback.Due to the exploratory nature of this educational approach, we decided to publish and share our approach before we embarked on a larger more formally structured program. We therefore are unable to present a well-designed and executed evaluation of the expected improvements in knowledge retention as well as enhanced non-cognitive skills and competences.We explored the benefits of the 6D-Approach in a course on Molecular Biology and Principles of Genetics. The nature of the chosen topic is characterized by mechanistic and structural aspects of medical science. Topics that require the development of other skills and competencies such as analytical prowess, emotional intelligence and emphatic aptness could improve by implementing the 6D-Approach. However, we cannot extrapolate the results from our pilot project to courses of a distinctly different nature.The positive feedback received from the course participants are an indication that the implementation of the 6D technique can incite enthusiasm in students. However, a formal and more robust measurable outcome need to be formulated and analyzed as to provide an overall accepted scientific proof of the expected superior end results.The successful implementations of the 6D approach builds on the technical and practical implementation of the model. However, like all teaching situations, success depends on the enthusiasm, knowledge and communication skills of the course directors - teachers as well.We successfully implemented the 6D-Approach in UME, and the approach was favorably received by the students. However, our study has specific limitations:The 6D-approach through the use of mentored journal clubs encourages retention of knowledge, critical thinking, metacognition, collaboration and leadership skills in addition to self-evaluation and peer feedback. Given the significance of the above traits in medical education, the approach provides a strategy to those who want to implement and strengthen the above traits in UME. Finally, and importantly, mentored journal clubs employed in the implementation of the approach represent an example of the medical education continuum, in particular highlighting the use of a continuing education method in UME."}
+{"text": "Methicillin Resistant Staphylococcus aureus (MRSA), which has been reported to survive on surfaces for several months. Bactericidal activity of copper-TiO2 thin films, which release copper ions and are deposited on glass surfaces and heated to high temperatures, is well known even when illuminated with very weak UVA light of about 10 \u03bcW/cm2. Lately, there is an increased intrerest for one-dimensional TiO2 nanomaterials, due to their unique properties, low cost, and high thermal and photochemical stability. Here we show that copper doped TiO2 nanotubes produce about five times more \u00b7OH radicals as compared to undoped TiO2 nanotubes and that effective surface disinfection, determined by a modified ISO 22196:2011 test, can be achieved even at low intensity UVA light of 30 \u03bcW/cm2. The nanotubes can be deposited on a preformed surface at room temperature, resulting in a stable deposition resistant to multiple washings. Up to 103 microorganisms per cm2 can be inactivated in 24 hours, including resistant strains such as Methicillin-resistant Staphylococcus aureus (MRSA) and Extended-spectrum beta-lactamase Escherichia coli (E. coli ESBL). This disinfection method could provide a valuable alternative to the current surface disinfection methods.Bacterial infections acquired in healthcare facilities including hospitals, the so called healthcare acquired or nosocomial infections, are still of great concern worldwide and represent a significant economical burden. One of the major causes of morbidity is infection with  On the other hand, no enzyme is known with ability to catalyze the \u00b7OH [Bacterial infections acquired in healthcare facilities including hospitals, the so called healthcare acquired or nosocomial infections (HAI), are still of great concern worldwide and represent a significant economical burden ,2. Accor2O2  or super2O2 [2\u2212  cleaning2O2 [2\u2212  with sev2O2 [2\u2212 . This is the \u00b7OH , which c the \u00b7OH , present the \u00b7OH \u201321. This2 irradiated with metal halide lamp was demonstrated by Matsunaga et al. already in 1985, when Escherichia coli were completely sterilized [2 was shown against a wide range of bacteria, fungi, algae, protozoa, viruses, and even bacterial endospores, fungal spores, and protozoan cysts [2 [2 with the energy of the light consistent with the TiO2\u2019s bandgap  and European commission\u2019s recommendation for \u201cclean\u201d food preparation and storage surfaces of less than 10 microbes per cm2, targeted disinfection efficiency is the inactivation of at least 103 microbes/cm2. Indeed, this was achieved recently by Dunlop et al. [2. In another study, efficient suppression of growth of Listeria monocytogenes, which can cause dangerous foodborne illness with high mortality rate [2O2 is formed  and Extended-spectrum beta-lactamase Escherichia coli (E. coli ESBL).Recently, interest in one-dimensional TiOtability . Here we2  and rutile TiO2 powder  oxide, rutile powder < 5\u03bcm) were dispersed in Erlenmeyer flask with KOH to obtain the stock solution of concentration 1 mg/ml. Media and culture materials were obtained from Gibco\u2013Invitrogen Corporation .The spin trap, 5-(Diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide (DEPMPO)  was used as purchased without further purification and stored at -80\u00b0C. The spin-trap stock solutions were always freshly prepared. Ethanol (EtOH) and methanol (MeOH) from Merck AG  were used in Lichrosolv\u00ae gradient grade quality. Titanium dioxide powder TiOL. innocua, which is closely related to pathogenic species L. monocytogenes, E. coli, and S. aureus. Suspension of each reference bacterial strain, was supplied by the Institute of Microbiology and Parasitology, Veterinary faculty, University of Ljubljana. Strain was maintained frozen at -70\u00b0C in sterile vials containing porous beads which serve as carriers to support microorganisms  and kept at -70\u00b0C. The inoculum was prepared in liquid medium  and incubated aerobically for 24 h at 37\u00b0C. After incubation the culture contain approximately 109 colony forming units (CFU) per milliliter. Working suspensions with appropriate concentrations were achieved by several 10-fold dilutions. The amount of the initial CFU/mL of the suspension placed in a petri dish was assessed by covering the suspension with approximately 15 mL nutrient agar. After overnight incubation at 37\u00b0C colonies were counted to estimate the CFU/mL in the original suspension.Antimicrobial properties were tested on non-pathogenic bacterium 2+-doped TiO2 nanotubes (Cu-TiO2NTs) were prepared in several steps: (i) first sodium titanate nanotubes (Na+-TiNTs) were synthesized from anatase powder  and 10 M NaOH (aq) (Aldrich) at T = 135\u00b0C for 3 days under hydrothermal conditions. Exact synthesis procedure is described previously [+-TiNTs were rinsed with 0.1 M HCl(aq) yielding protonated titanate nanotubes (H+-TiNTs), (iii) then H+-TiNTs were dispersed in 0.5 mM solution of Cu2+(aq) (source of the Cu2+ was CuSO4\u00b75H2O (Riedel de Haen)) and stirred at room temperature for 3 hours. By centrifugation the solid material was separated from the solution containing excess copper, and (iv) finally isolated material was dryed for 10 hours.Cueviously , (ii) in\u03b8 range from 10 to 65\u00b0). The elemental compositions of samples were investigated with a field emission scanning electron microscope  equipped with an EDX (energy dispersive X-ray spectrometer) element analysis system. Morphology of the particles in the sample was determined using transmission electron microscope . The specimen for the TEM investigation was prepared by dispersing the sample in MeOH with the help of an ultrasonic bath and depositing a droplet of the dispersion on a lacey carbon-coated copper grid.The powder X-ray diffraction (XRD) pattern was obtained on a Bruker AXS D4 Endeavor diffractometer using Cu K\u03b1 radiation  with spin trapping, which was optimized for measurement of primary radicals generated in the vicinity of the nanomaterial surface. This was achieved by measuring primary hidroxyl radicals in the presence of 30% ethanol with 5-(Diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide spin trap (DEPMPO). EPR spin trapping was applied to measure the generation of reactive oxygen species (ROS) production. On the surface, small pool, proportionate to the size of the sample, was made with silicon paste and was filled with 2 \u03bcl of 0,5 M DEPMPO and 18 \u03bcl of 30% ethanol and irradiated with 290 nm diode for 5 min. The diode was 1\u20132 mm above the surface of the sample. The solution with short-lived radicals being trapped in the form of stable DEPMPO spin adducts was then drawn into the quartz capillary of 1 mm diameter, which was put in the 5 mm wide quartz tube and transferred into EPR spectrometer. All EPR measurements were performed on an X-band EPR spectrometer Bruker ELEXYS, Type W3002180. All measurements were recorded at room temperature using 1 Gauss (10\u22124 T) modulation amplitude, 100 kHz modulation frequency, 20 ms time constant, 15 x 20 seconds sweep time, 20 mW microwave power and 150 G sweep width with center field positioned at 3320 G.The photocatalytic activity of synthesized TiO2NTs was made on polystyrene petri dishes (55.4 cm2). Petri dishses were washed before deposition. They were soaked in 20% NaOH solution, rinsed with distilled water, and finally with ethanol vapor. The suspension of the nanotubes with concentration of 1 mg/ml was processed with ultrasonic liquid processor  prior to the deposition on the slides. Sonication was performed using 419 MicrotipTM probe, 15 min process time, 10 s pulse-ON time, 10 s pulse-OFF time and maximum amplitude (resulting in 52 W of power). The petri dishes were treated with compressed air 3 times for 3 s. 150 \u03bcl of nanoparticle suspension was applied on each petri dish, immediately after compressed air treatment, and smeared evenly. The same number of dishes with nanoparticle deposition and control dishes were prepared for each experiment. On control dishes, only 150 \u03bcl of solution was applied. After the deposition, the dishes were left in the oven at 50\u00b0C for 2 hours. Then they were rinsed with distilled water and put back in the oven at 50\u00b0C for another 2 hours. The amount of deposited material was estimated from EPR signal decrease when soaking the surface with water, alkaline or acidic medium.The deposition of Cu-TiO2NTs coated petri dishes  using a slightly modified test according to standard ISO 22196 , a 24-hour inoculation method [5 CFU/mL) in a petri dish for 24 hours at 4\u00b0C in a refrigerator. Petri dishes were covered with the lid to prevent drying of the suspension during the exposure. The surfaces were illuminated with a LED lamp  for \u201cL. inncoua\u201d series, 9 points (N = 9) for \u201cS. aureus\u201d series, 6 points (N = 6) for \u201cMRSA\u201d series, 14 points (N = 14) for \u201cE. coli ESBL\u201d series, and 9 points (N = 9) for \u201cE. coli\u201d series; error bars represent standard error of the regression coefficient.Height of bars in 2 nanoparticles with high photocatalytic activity and small amount of copper ions incorporated into TiO2 lattice, avoiding leaching of the copper ions from the antibacterial surface. In addition, we wanted to develop TiO2 nanoparticles that would adhere better than spherical particles to achieve better stability on a preformed surface. Being aware that transition metal doping of titanate nanostructures results in a change of surface chemistry as well as in catalytic and photocatalytic properties [2 (in situ doping) [in situ doping it was shown that Cu2+ species may intercalate between titanate layers and preserve structure of the nanotubes, whereas in situ method does not produce well-formed nanotubes [2+ species preferentially adsorbed on the surface of the nanotubes in the form of CuO nanoparticles, while doping at low levels of copper (app 0.02%) also produced well-formed nanotubes with more homogeneous distribution of Cu2+ ions. Although the distribution of Cu2+ could not be resolved by electron microscopy, low temperature electron paramagnetic resonance (EPR) method showed that 70% of EPR intensity could be attributed to intercalated Cu2+ ions, suggesting that around 30% of the Cu2+ ions were probably substitutionally doped into the titanate host structure [The goal of this study was to produce TiOoperties , three doperties . Epitaxioperties  than foroperties  and grow doping)  producedanotubes . Moreovetructure .2 host structure, we prepared TiO2 and Cu2+-doped TiO2 nanotubes by calcination in air at 390\u00b0C (10 h) of pure hydrogen trititanate nanotubes (HTiNTs) and HTiNTs impregnated with Cu2+ [2 nanotubes (TiO2NTs) and Cu-doped2+ TiO2 NT (Cu-TiO2NTs) samples  images show a crystalline structure of TiOanotubes  with no anotubes . Using aanotubes , which ianotubes , but no anotubes  indicati3+ state, whereas in the bulk we can see characteristic octahedrally coordinated Ti4+ EELS spectrum.We also performed electron energy loss spectroscopy (EELS) on the bulk and at the edge of a nanotube OH) spin adduct instead of DEPMPO-OH [2+-doped TiO2 nanotubes (Cu-TiO2NTs) is comparable to the amount generated by undoped TiO2 nanotubes (TiO2NTs) and almost as high as Degussa P25, commercially available spherical TiO2 nanoparticles  spectroscopy. Although this is a highly sensitive method for detecting paramagnetic species including radicals, some of them, like  \u00b7OH radicals are so reactive that they never reach high enough concentrations to be detected by EPR. To overcome this, short-lived radicals must be trapped by nitrone compounds forming stable (long-lived) radical adducts, which can then be easily detected and their identity confirmed through their specific hyperfine splitting pattern. Although it has been shown that for some nanoparticles, such as graphene nanosheets, their photocatalytic activity depends on hydroxyl and superoxide radicals as well as holes , same is anatase . In this anatase . In agreEPMPO-OH . The anaarticles . The pho2 and Cu-doped TiO2 nanoparticles generated similar amount of radicals  using a slightly modified test ISO 22196, a 24-hour inoculation method [L. innocua, E. coli, E. coli ESBL and S. aureus including MRSA. L. innocua has been chosen due to close relativity to L. monocytogenes, which has the highest mortality rate among all known foodborne pathogens and its ability to grew at very low temperatures, even at -1.5\u00b0C [L. innnocua is an example of Gram-positive bacteria, E. coli has been added to our selection as the very common Gram-negative bacteria. S. aureus has been included to the test as a very frequent strain that can cause many infectious with severe complications. In addition, two antibiotic-resistant bacterial strains MRSA and E. coli ESBL have been selected to check the efficiency against the bacteria that are currently among the biggest challenges in clinical medicine and maintenance of clean hospital infrastructure. Starting contamination concentration range between 10 and 10000 has been decided based on a typical number of bacteria per gram of food or cm2 of surface, which has then been renormalized to the surface of our petri dish. The petri dishes innoculated with the predefined number (as defined above) of the previously selected bacteria have been exposed and continuously illuminated with 300 mW/m2 at the same time for 24 hours at 4\u00b0C. Then they were covered with agar and incubated at 37\u00b0C for additional 3 days. The colony forming units (CFU) of microorganisms have been counted on petri dishes without the coating  that has been previously protected by deposition of tested nanomaterial. The whole sample (with nanotubes and bacteria on) has been stained by rhodamine B isothiocyanate (RITC), which strongly binds to proteins on the surface of bacteria and which at the same time enables identification of TiO2 nanotubes by their modulation of the RITC emission spectrum. Namely, fluorescence properties of rhodamine B are known to be strongly affected by TiO2 via self-photosensitized oxidative transformation [2 nanotubes and bacteria on the same spectrally-contrasted image (2NTs coated surface showed remarkably lower fluorescence emission peak position wavelengths (b) span very large range (MAX). Individual L. innocua cells were clearly identified on an uncoated surface by higher fluorescence intensity, very low bleaching rates as well as the blue-shifted spectral peak position .To prove the relation between the disinfection efficiency and the contact/proximity between material and bacteria , we tried to co-localize nanotubes and bacteria on the nanotube coated surfaces. To assess the later, we conducted microscopic visual inspection of inoculated surface , which enables us to discriminate the nanotubes and bacteria based on the shape of objects. The micrographs confirmed that in agreement with fluorescence data shown above, individual bacteria were found mainly in those micro regions without or with much less of the nanomaterial coating. In contrast with the samples without the coating Click here for additional data file."}
+{"text": "Background: As an important downstream factor in the Hippo pathway, yes-associated protein 1(YAP1) has been detected to be elevated in various cancers and demonstrated to play a role in tumor development. Therefore, we evaluated by a meta-analysis the prognostic value of YAP1 in cancer patients.p<0.001) and disease-free survival (DFS) , as well as in studies with recurrence-free survival (RFS) , and disease-specific survival (DSS) . Meanwhile, YAP1 overexpression was also observed to be significantly associated with worse OS in GEPIA .Results: Sixty-eight studies with 8631 patients were identified. The results indicated that YAP1 overexpression predicted unfavorable patient prognosis in studies with overall survival (OS) (HR=1.76, 95%CI: 1.50-2.06, Conclusions: Overexpression of YAP1 showed great association with poorer prognosis in patients with various cancers, particularly liver cancer. Therefore, YAP1 might be an important prognostic marker and a novel target of cancer therapy.Methods: We searched for potential publications in several online databases and retrieved relevant data. Overall and subgroup analyses were performed. Begg\u2019s and Egger\u2019s tests were used to assess publication bias. Online dataset GEPIA was used to generate the survival curves and verify the prognostic role of YAP1 in patients with tumors. Cancer, a health challenge worldwide, is a dominating cause of death mainly in developed countries, with high incidence and low curative rate . In 2018Drosophila by using genetic screens to search the tumor suppressor genes [Cancers are derived from the excessive growth of cancer cells, which activate the mechanisms to promote the cells growth and cancer progression . A varieor genes \u201315. YAP1or genes . Meanwhior genes , 17. Subor genes , 19. In or genes  reportedor genes  considerTherefore, it is critical to investigate the relationship between YAP1 and cancer. The first meta-analysis regarding YAP1 was published three years ago, and reported that the positive expression of YAP1 was associated with poorer prognosis in various cancers . In receIn total, 3208 studies were searched initially, and 101 were considered potentially relevant articles. Among them, 33 studies were excluded . In the process of selecting literature, some studies were excluded due to the questionable data, such as the studies of Han et al., Fan Ye et al., and Wang et al. \u201325. The Of these 68 studies, 53 evaluated the relationship between YAP1 and OS, 19 for DFS, 7 for progression-free survival (PFS), 8 for RFS, and 5 for DSS. Collectively, these 68 included studies contained data for 8631 patients, from 12 countries, who were diagnosed with 18 different types of cancer, such as liver cancer, gastric cancer, and breast cancer, and so on.2=73.7%, p<0.001) and a random-effects model was applied. The result indicated that overexpression of YAP1 would lead to poorer OS  (There was obvious heterogeneity of OS in 53 studies (Ip<0.001) .2=30.7%, p=0.10). The results showed that the overexpression of YAP1 was related to poorer DFS   .2=69.2%, p=0.003; RFS: I2=55.6%, p=0.027), so we used a random-effects model to evaluate the pooled HR and its 95% CI ; for DSS, pooled HR , for insignificant heterogeneity  was calculated using a fixed-effects model. The results suggested that YAP1 overexpression would predict poorer RFS and DSS, but not PFS (As shown in  not PFS .p<0.001), but not in non-Asian patients. In the tumor type subgroup, significant associations were found in liver cancer , esophageal cancer , breast cancer , and bladder cancer . When stratified by method, IHC  and PCR , both showed a significant relationship with poorer OS. When analyzed by YAP1 staining location, total YAP1 expression and nuclear YAP1 expression showed great significance with dismal prognosis in tumor patients . Regarding the type of analysis, univariate analysis  indicated that overexpression of YAP1 was significantly related with elevated OS, as did the multivariate analysis .To explore the source of heterogeneity, we further carried out subgroup analysis according to four possible factors , summarized in p<0.001), as well as in Chinese , was associated with poor DFS. In the subgroup analysis by tumor type, a significant relationship was found between patients with liver cancer  and DFS. Regarding staining location, subgroup analysis suggested that both total YAP1 expression  and nuclear YAP1 expression  were predictors of poorer DFS. According to the subgroup result of analysis type, univariate analysis  and multivariate analysis  showed significant associations with poorer DFS.Based on ethnicity, tumor type, staining location, and analysis type, subgroup analysis was also performed in the studies reporting DFS. YAP1 overexpression in Korean studies , but there was significant bias per Egger\u2019s test (p<0.05). Therefore, considering the efficiency of two tests, we believed that there was publication bias among studies reporting OS. Publication bias was not detected in studies reporting DFS   .p<0.001)  . HoweverThe mechanism involved in the development of cancer is complicated and is generally thought to be related to the dysfunction of cellular signaling pathways. Previous studies suggested the cellular signaling pathways could regulate the growth factors and expression of some vital genes that are associated with cell proliferation, migration, metabolism and junction between cells and so forth \u201395. HippIn our study, overall and subgroup analyses were conducted to provide evidence of the association between the overexpression of YAP1 and patients\u2019 prognosis. The results showed that overexpression of YAP1 predicted poorer OS, DFS, RFS, and DSS. The sensitivity analysis showed that no single study changed the pooled results significantly, which indicated the results of sensitivity analysis were stable. Subgroup analysis was conducted by ethnicity, tumor type, method, staining location, and HR estimation analysis type. According to the results of the subgroup analysis, there were no clear relations between YAP1 with OS in non-Asians, colorectal cancer, lung cancer, renal cancer, and ovarian cancer.To clarify it further, we investigated the association between YAP1 expression and cancer patients\u2019 prognosis in the online database, GEPIA. The results showed YAP1 expression was observed to be significantly related with worse OS, but not with DFS. Additionally, the same correlations were found between YAP1 expression and unfavorable OS and DFS in patients with glioma, as well as shorter OS in esophageal carcinoma patients and better DFS in bladder urothelial carcinoma patients. The differences between this meta-analysis and the online dataset might come from the following reasons. First of all, the detecting methods are thought to be the main reason for the differences. In most of the studies included in our meta-analysis, IHC was used to detect the expression of YAP1 protein, whereas the data of the online dataset were from the results of RNA sequencing. What\u2019s more, in some specific cancer types, the number of patients that included in meta-analysis was much larger than the dataset. For example, although we have summed up the data of cholangio carcinoma and liver hepatocellular carcinoma, the total number of patients with liver cancer studied in the meta-analysis (1629), far exceed the number of the liver cancer patients represented in the online dataset (398). In addition, the cancer types are different. For instance, the breast cancer patients included in our meta-analysis represented various breast cancer subtypes, while the data in GEPIA included data for breast invasive carcinoma (BRCA) only. Therefore, while GEPIA provides valuable insights into gene-specific cancer survival outcomes, it is still being developed, and results obtained from the robust meta-analysis remain more convincing.In the present study, we identified 14 studies that reported the relationship between prognosis of patients with liver cancer and YAP1 overexpression. Our results suggested that YAP1 overexpression predicted poorer OS and DFS. YAP1 was reported to be amplified and overexpressed in many cancers, and patients were suggested to receive further treatment, such as chemotherapy and hepatectomy. Although tumors can be removed by hepatectomy, rapid repair of the liver is crucial after partial hepatectomy, in which YAP1 increases and participates in the promotion of cell proliferation , 105. ThIt is widely accepted that YAP1 is phosphorylated and accumulates in the cytoplasm such that it loses its ability to activate genes that promote cell invasion and proliferation and suppresses cell apoptosis once the Hippo pathway is activated , 107. ThPreviously, Sun et al.  performeDespite our efforts to make an accurate and comprehensive analysis, a few limitations inevitably existed in our study. First of all, some studies did not report clinical data; we had to extract the HR from the Kaplan-Meier survival curves indirectly, which might influence the accuracy of the original data. Secondly, publication bias was observed in our study among the studies reporting OS and we thought it was associated with the studies we included. Most of the studies on the relation between YAP1 and prognosis were based on Asian patients. Meanwhile, the bias might also be due to the non-publishing of studies with negative results. More studies are needed to resolve this limitation. Thirdly, significant heterogeneity between studies with OS, PFS, and RFS were detected. However, we failed to identify the source of heterogeneity by subgroup analysis. Then we minimized the effect by applying a random-effects model. Possibly, it was due to the differences in the methods of surgery and treatment, and baseline YAP1 expression level. Additionally, owing to the lack of clinical data, we could not confirm their contribution to heterogeneity.Overall, our study provides evidence that overexpression of YAP1 is significantly associated with the progression and recurrence of tumors and it might be a useful prognosticator in various cancers. However, future prospective studies with more complete and available clinical information are expected and required to confirm the results.Studies related to YAP1 were searched through PubMed, EMBASE, Web of Science, WANFANG, and CNKI. The main keywords used in the search were as follows: \u201cYes-associated protein 1\u201d, \u201cYAP1\u201d, \u201cHippo\u201d, \u201cNeoplasms\u201d, \u201cNeoplasia\u201d, \u201cTumor\u201d, \u201cCancer\u201d, \u201cMalignancy\u201d, \u201cPrognosis\u201d, \u201cSurvival\u201d and \u201cMetastasis\u201d. The search strategy for PubMed is shown in Studies were included when they met the following criteria: (1) studies explored the association between YAP1 expression and patients\u2019 prognosis; (2) studies declared the number and information of patients; (3) studies measured positive or high YAP1 expression in patients with any carcinoma by immunohistochemistry and polymerase chain reaction; (4) the endpoints were listed which are OS, RFS, DFS, PFS, and DSS; (5) the HRs with their 95% CIs or Kaplan-Meier survival curves were provided; (6) as for the overlapping data, the latest and most complete studies were included.Studies were excluded if they: (1) were published meta-analyses, letters, abstracts, reviews or case reports; (2) did not provide survival information; (3) the data were not related to humans; (4) used data from public databases; or (5) were questionable data or the full text was not found.The information extracted from the included studies were: first author, publication year, patient source, number of patients, tumor type, specimen, methods, YAP1 expression, staining location, median follow-up time, prognostic outcome, HR estimation . We applied the HR with its 95% CI from the text directly or calculated the HR using Engauge Digitizer 4.0 by Tierney et al. through Kaplan-Meier survival curve analysis. The literature and data were selected and abstracted by two authors, and divergence views were resolved in a group meeting.The NOS was used to assess the quality of the literature. Two authors of our group scored all eligible studies independently, and all discrepancies were discussed to arrive at a conclusion. The scores of studies included ranged from 6 to 8, which implied high quality.Gene Expression Profiling Interactive Analysis (GEPIA) is a new online dataset, including 9736 tumors and 8587 normal samples from The Cancer Genome Atlas (TCGA) and Genotype-expression (GTEx) projects. In this study, for further verifying the association between YAP1 and patients\u2019 prognosis with tumors, the GEPIA was used to perform the survival analysis.2 test. Significant heterogeneity was defined by a p value <0.1 or I2 value >50%, based on which we applied a random-effects model; if no significant heterogeneity was found, we used a fixed-effects model. Subgroup analysis was conducted according to ethnicity, tumor type, staining location, and analysis type. In the sensitivity analysis, to estimate the impact of each study on our meta-analysis, we omitted individual studies consecutively. Begg\u2019s test and Egger\u2019s test were performed to assess the potential publication bias. If the funnel plots showed the points distributed asymmetrically on both sides of the axis, the publish bias was regarded to be significant. All statistical tests were bi-directional and a p-value less than 0.05 indicated statistical significance. Data analysis was performed using STATA 12.0 .HRs and their 95% CIs were used to assess the relation between YAP1 expression and prognosis of patients with carcinoma. We evaluated the heterogeneity using the Q test and ISupplementary FiguresSupplementary Table 1Supplementary Table 2Supplementary Table 3"}
+{"text": "MLKL is the most important executor of necroptosis pathway. Recent studies have demonstrated that MLKL could serve as a potential prognostic biomarker for cancer patients. However, most studies reported so far are limited in discrete outcome and sample size.We systematically searched PubMed, Embase, Web of Science and CNKI to obtain all relevant articles about the prognostic value of abnormally expressed MLKL in patients with any type of tumor. Odds ratios or hazards ratios (HRs) with corresponding 95% confidence intervals (CIs) were pooled to estimate the association between MLKL expression and clinicopathological characteristics or survival of cancer patients.A total of 6 eligible studies with 613 cancer patients were enrolled in our meta-analysis. Our results demonstrated that decreased expression level of MLKL was significantly associated with poor overall survival (OS)  and event-free survival (EFS)  in cancer patients. Furthermore, subgroup analysis divided by type of cancer, sample size, follow-up time and Newcastle\u2013Ottawa Scale (NOS) score showed consistent prognostic value. In addition, our analysis revealed that decreased expression level of MLKL was significantly associated with advanced tumor stage, more lymph node metastasis and older age.In conclusion, our meta-analysis suggested that decreased MLKL expression might be a convinced unfavorable prognostic factor that could help the clinical decision-making process. Despite the great development of therapies over past years, cancer is still the major public health problem worldwide . InnumerNecrosis, which is regarded as a kind of unprogrammed cell death previously, is characterized by plasma membrane permeabilization, cellular collapse, swelling of the organelles, mitochondria dysfunction and inflammation in the surrounding tissues . NecrosiMLKL is the most important executor of necroptosis pathway. Plasma membrane translocation of trimerized MLKL protein is essential for necroptosis, leading to permeabilization of endoplasmic reticulum, mitochondria and lysosome . MLKL alThe present review was performed in accordance with the standard guidelines for meta-analysis and systematic reviews of tumor marker prognostic studies , 28. TheStudies included in this analysis had to meet the following inclusion criteria: (1) Patients were pathologically diagnosed with any type of cancer; (2) MLKL expression was determined in human tissues or plasma samples using any techniques; (3) Patients were divided into high and low expression or positive and negative expression groups, the relationship between MLKL expression levels with survival outcome was investigated; (4) Sufficient published data or the survival curves were provided to calculate hazard ratios (HR) for survival rates and their 95% confidence intervals (CI). Exclusion criteria were as follow: studies without usable or sufficient data, studies using non-human samples, reviews, laboratory articles, case reports, letters, unpublished articles and conference abstracts. All eligible studies were carefully screened by two researchers (Binwu Hu and Deyao Shi), and discrepancies were resolved by discussing with a third researcher (Xiao Lv).Two investigators (Binwu Hu and Deyao Shi) extracted relevant data independently and reached a consensus on all items. For all eligible studies, the following information of each article was collected: author, year of publication, tumor type, expression associated with poor prognosis, Newcastle\u2013Ottawa Scale (NOS) score, method of obtaining HRs and the characteristics of the study population , follow up (month)), endpoints, assay method, cut-off value and survival analysis. For endpoints, overall survival (OS), disease-free survival (DFS) and recurrence -free survival (RFS) were all regarded as endpoints. In addition, DFS and RFS were redefined as event-free survival (EFS) in our article. We employed HR which was extracted following a methodology suggested previously to evaluate the influence of MLKL expression on prognosis of patients . If possQuality of all included studies was assessed independently by three researchers  using the validated NOS, and disagreements were resolved through discussion with another researcher (Xiao Lv). This scale uses a star system to evaluate a study in three domains: selection of participants, comparability of study groups, and the ascertainment of outcomes of interest. We considered studies with scores more than 6 as high-quality studies, and those with scores no more than 6 as low-quality studies.2 statistics. A p value less than 0.10 or an I2 value larger than 50% were considered statistically significant. The fixed-effect model was used for analysis without significant heterogeneity between studies . Otherwise, the random-effect model was chosen. To explore the source of heterogeneity, subgroup analysis was preformed through classifying the included studies into subgroups according to similar features. We also conducted sensitivity analysis to test the effect of each study on the overall pooled results. In addition, for the studies from which we could obtain clinicopathological characteristics, we calculated the pooled ORs and performed heterogeneity tests to analyze the relationship between MLKL expression level with lymph node metastasis, tumor stages, age, differentiation grade\u00a0and gender in different types of cancers. Due to the limited number of studies (below 10) included in this analysis, publication bias was not assessed.Statistical analysis was performed using Stata Software 14.0 . Pooled HRs (high/low) and their associated 95% CIs were used to analyze the prognostic role of MLKL expression in various cancers. Pooled odds ratios (ORs) (low/high) and their associated 95% CIs were used to analyze the association between MLKL expression level with clinicopathological parameters. The heterogeneity among studies was evaluated using Cochran\u2019s Q and In\u2009=\u2009361), review (n\u2009=\u200968), patent (n\u2009=\u20096), meeting abstract (n\u2009=\u200960), studies describing non-cancer topic (n\u2009=\u2009163), studies describing non-MLKL topic (n\u2009=\u200956), studies belonging to basic research (n\u2009=\u200966) and studies lacking relevant data (n\u2009=\u20093). Eventually, 6 studies meeting the inclusion criteria were included in this meta-analysis. The screening process and results are shown in Fig.\u00a01. Among these studies, a total of 613 patients were included, with a maximum sample size of 153 patients and a minimum sample size of 54 patients (Mean 102.0). The accrual period of these studies ranged from 2013 to 2017. The regions represented in this study included\u00a0the China (5) and the United States of America (1). Six different types of cancer were evaluated including breast cancer, gastric cancer, cervical squamous cell carcinoma, ovarian cancer, colon cancer and pancreatic adenocarcinoma. Among these studies, OS, DFS and RFS were estimated as survival outcome in 100% (6/6), 17% (1/6) and 33% (2/6) of the studies respectively. DFS and RFS were combined together into EFS, which was regarded as prognostic parameter in our study. To evaluate the expression of MLKL, all studies used immunohistochemistry (IHC) technique. Because the cut-off definitions were various, the cut-off values were different in these studies.According to our search strategy, a total of 789 studies were retrieved. Among these researches, the following studies were excluded: duplicate n\u2009=\u200936, review p\u2009<\u20090.001), which indicated that decreased expression level of MLKL was significantly associated with poor OS in cancer patients . In order to further explore the association between abnormal expression levels of MLKL with OS in cancer patients, subgroup analysis was performed based on the following factors: type of cancer , sample size (fewer than 100 or more than 100), follow-up time (fewer than 100 or more than 100\u00a0months), paper quality (NOS scores <\u20097 or\u2009\u2265\u20097). The subgroup analysis illustrated the same results that the significant association between decreased expression levels of MLKL with poor OS of cancer patients was not altered with all the factors above (Table\u00a0p\u2009<\u20090.001) and digestive system malignancies\u00a0   and more than 100\u00a0months   and NOS scores \u22657   Fig.\u00a0. For sam01) Fig. . For fol01) Fig. . For pap01) Fig. . No signp\u2009=\u20090.787; I2\u2009=\u20090%).Using HRs of Cox multivariate analysis from three studies, we found that decreased MLKL expression was an independent prognostic factor for OS in cancer patients . Furthermore, there was no significant heterogeneity among studies (p\u2009=\u20090.017) . However, due to the limited number of included studies, we did not performed the subgroup analysis.A total of three studies including 368 patients reported the impact of abnormally expressed MLKL on DFS or RFS of cancer patients. In the current study, we defined DFS and RFS as EFS. The consequence displayed that decreased expression level of MLKL was significantly associated with poor EFS in cancer patients , more lymph node metastasis  and older age . However, there was no significant association between decreased expression of MLKL with the differentiation grade  and gender .As shown in Table\u00a0Sensitivity analysis was performed to examine the effects of individual study on the overall results. For OS, the sensitivity analysis identified that result from Li et al. affected pooled HR greatly, indicating that this study was possible to be the main source of heterogeneity. However, after excluding this study, we still observed that decreased expression level of MLKL was significantly associated with poor OS in cancer patients Fig.\u00a0. For EFSNecroptosis is the most well-studied form of regulated necrosis, which could be triggered by death receptors ligands and stimuli that induce the expression of death receptor ligands under apoptotic deficient conditions . AccumulHere we performed current meta-analysis to explore the prognostic value of abnormally expressed MLKL and the relation between MLKL expression levels with clinicopathological characteristics as well as to further reveal the prognostic value of necroptosis in cancer patients. We examined 6 independent studies comprising data from a total of 613 patients. Through systematic analysis, our results demonstrated that decreased expression level of MLKL was significantly associated with poor OS in cancer patients. In addition, the subgroup analysis results displayed that factors including type of cancer, sample size, follow-up time and paper quality did not alter above results. Furthermore, by combining HRs from studies using Cox multivariate analysis, we found that MLKL was an independent prognostic factor of OS in cancer patients. DFS and RFS are important parameters reflecting the progression of tumor. In this article, we defined DFS and RFS as EFS. The prognostic significance of MLKL in EFS was evaluated in 3 studies with 368 patients. The results indicated that decreased expression level of MLKL was significantly associated with poor EFS of cancer patients.As for the clinicopathological characteristics, our analysis revealed that decreased expression level of MLKL was significantly associated with advanced tumor stage, more lymph node metastasis and older age. However, there was no significant association between MLKL expression levels with gender or differentiation grade of patients.Mechanisms underlying the regulatory role of MLKL in tumorigenesis and tumor progression have been extensively investigated. In cancer cells, RIPK3 interacts with and phosphorylates MLKL to promote necroptosis . MLKL isIn our study, a few limitations should be underlined. First, only 6 studies were included in our meta-analysis, and even fewer articles, three articles were included for the EFS analysis, this restricted our ability to evaluate the prognostic value of MLKL in subgroup analysis and might lead to the bias of the results. Second, due to the limited number of included articles, we could not perform the publication bias analysis, which was possible to exist in our meta\u2013analysis. Third, although all studies used IHC to evaluate the expression of MLKL, the cut-off value differed among studies, which might cause bias in the meta-analysis. Fourth, we calculated some HRs through survival curves, it might not be precise enough. Therefore, larger-scale, multicenter and high-quality studies are desperately necessary to confirm our findings.In conclusion, our study revealed that decreased expression level of MLKL was significantly associated with poor OS and EFS in cancer patients. Moreover, the expression level of MLKL was associated with clinicopathological features including TNM stage, lymph node metastasis and age. This is the first meta-analysis to evaluate the relationship between expression levels of MLKL, the critical component in necroptosis pathway, with prognosis of cancer patients. In the future, more relevant studies are warranted to investigate the role of MLKL in human cancer."}
+{"text": "The Meaningful Use (MU) program has promoted electronic health record adoption among US hospitals. Studies have shown that electronic health record adoption has been slower than desired in certain types of hospitals; but generally, the overall adoption rate has increased among hospitals. However, these studies have neither evaluated the adoption of advanced functionalities of electronic health records (beyond MU) nor forecasted electronic health record maturation over an extended period in a holistic fashion. Additional research is needed to prospectively assess US hospitals\u2019 electronic health record technology adoption and advancement patterns.This study forecasts the maturation of electronic health record functionality adoption among US hospitals through 2035.The Healthcare Information and Management Systems Society (HIMSS) Analytics\u2019 Electronic Medical Record Adoption Model (EMRAM) dataset was used to track historic uptakes of various electronic health record functionalities considered critical to improving health care quality and efficiency in hospitals. The Bass model was used to predict the technological diffusion rates for repeated electronic health record adoptions where upgrades undergo rapid technological improvements. The forecast used EMRAM data from 2006 to 2014 to estimate adoption levels to the year 2035.In 2014, over 5400 hospitals completed HIMSS\u2019 annual EMRAM survey . In 2006, the majority of the US hospitals were in EMRAM Stages 0, 1, and 2. By 2014, most hospitals had achieved Stages 3, 4, and 5. The overall technology diffusion model  reached an adjusted R-squared of .91. The final forecast depicted differing trends for each of the EMRAM stages. In 2006, the first year of observation, peaks of Stages 0 and 1 were shown as electronic health record adoption predates HIMSS\u2019 EMRAM. By 2007, Stage 2 reached its peak. Stage 3 reached its full height by 2011, while Stage 4 peaked by 2014. The first three stages created a graph that exhibits the expected \u201cS-curve\u201d for technology diffusion, with inflection point being the peak diffusion rate. This forecast indicates that Stage 5 should peak by 2019 and Stage 6 by 2026. Although this forecast extends to the year 2035, no peak was readily observed for Stage 7. Overall, most hospitals will achieve Stages 5, 6, or 7 of EMRAM by 2020; however, a considerable number of hospitals will not achieve Stage 7 by 2035.We forecasted the adoption of electronic health record capabilities from a paper-based environment (Stage 0) to an environment where only electronic information is used to document and direct care delivery (Stage 7). According to our forecasts, the majority of hospitals will not reach Stage 7 until 2035, absent major policy changes or leaps in technological capabilities. These results indicate that US hospitals are decades away from fully implementing sophisticated decision support applications and interoperability functionalities in electronic health records as defined by EMRAM\u2019s Stage 7. Technology policy in health care has profoundly affected service delivery and operational efficiencies ,2. The pResearch in the health care field has closely linked EHR technology adoption to business and clinical outcomes -10. As aThe Health Information Technology for Economic and Clinical Health (HITECH) Act  was signThe impact of the HITECH Act has been evaluated in the health services literature so that policymakers can assess the extent to which their intended EHR adoption goals are being realized . Indeed,In the research literature that focuses on EHR technology adoption, analyses frequently rely on MU data for measuring current use percentages in a binary fashion . In partConcurrently, hospital planners adopted maturity models that sought to frame EHR implementation as a journey rather than an endpoint. The Healthcare Information and Management Systems Society (HIMSS) Analytics\u2019 Electronic Medical Record Adoption Model (EMRAM)  was deveDiffusion of innovation model produces \u201ctechnology sophistication forecasts\u201d that predict the degree to which a market or sector has and will adopt sequentially higher levels of functionality in the near future ,30. The The purpose of this study was to explore when hospitals will achieve critical EHR functionality. HIMSS Analytics\u2019 EMRAM data and Bass diffusion models were used to assess current EHR capability levels and forecast future diffusion of EHR functionality levels.In this study, we explored US hospitals\u2019 EHR technology adoption and implementation patterns accounting for functionality and application upgrades. We used the HIMSS EMRAM data to observe the granular change or progression of EHR functionality among hospitals. The same dataset was used to train the Bass diffusion model and then predict the EMRAM score  for each hospital. The forecasted scores were aggregated across all hospitals within each future year to depict a national picture of EHR functionality improvements until 2035. We assumed no change in future policies that would affect health IT efforts or EHR functionality . Similarly, no dramatic advancement in the technology itself is modeled  as such innovations would change the diffusion curves.We used the HIMSS Analytics\u2019 EMRAM data since it provides an MU-comparable EHR adoption measure that takes a more granular approach to assessing functionality uptake  27]. EM. EM27]. Modeling the EHR diffusion using an adaptation approach requires two components. First, the technology must track progression through the diffusion stages as set by the \u201cDiffusion of Innovation\u201d theorem, resulting in an \u201cS-curve\u201d to measure the functional form of analysis appropriately . Second,The EMRAM data were used as the basis of BB-01 statistical analyses, with estimates calculated in Microsoft Excel using nonlinear regression estimates. Visual Basic, Solver, and the SAS Model Procedure were also used to train and estimate several parameters used by the Bass model ,35. The On average, approximately 5200 hospitals were represented in the EMRAM data across the years studied (2006-2014). The percentage of hospitals achieving various EMRAM stages varied across years . More thThe overall model produced an adjusted R-squared of .91, suggesting a high model fit. The forecast used EMRAM data from 2006 to 2014 to estimate adoption levels to the year 2035. leapfrogging, suggesting that adopters either skipped Stage 4 or moved concomitantly with technology adoption for both Stages 4 and 5. Based on the analysis, most hospitals will be focused on the higher stages  by the year 2025. It is also clear that Stage 7 will not reach a maximum or plateau by the end of the forecast window . There are two potential explanations for the simultaneous, multistage adoption in the lower EMRAM levels: EHR vendors integrating multiple functions upfront and hospitals being overtly motivated by external factors .First, EHR vendors may have introduced multiple functions at once. As part of the MU program, the US government introduced an EHR vendor certification regime . Its purSecond, hospitals may have wanted to move through the early stages quickly . ConsideAdditionally, hospitals with more recent EHR adoptions may have taken a simultaneous, multistage form in that product vendors began bundling functionalities and clinical capabilities together in a more holistic fashion ,42. SimiThe government\u2019s MU program did not provide rewards or incentives for the later EMRAM stages . As a result, one of the previously noted major external motivations for adopting higher EHR functionalities was not in play. The BB-01 model effectively controls for this change in motivational factors, suggesting that internal motivation measures play a significantly large role in EHR functionality and clinical application adoption . This caThe lack of additional EHR incentives in this period will potentially cause internal factors to become the main driver for hospitals to adopt new EHR features. In this scenario, hospitals should observe the imminent need to request and adopt new EHR functionalities to achieve their higher-order goals . For example, EMRAM\u2019s Stage 6 of EHR maturation requires the full adoption of CDS systems across the entire health care system for a variety of clinical practice guidelines. However, if the desired outcomes of a health system, either cost or clinical outcomes, are not aligned with such decision support enhancements in the underlying EHR platform, the hospitals may not have the internal pressure or desire to adopt the new EHR functionalities. Indeed, a complex series of internal factors may disincentivize such progression through EHR functionalities, specifically in a volatile health care market . Hence, a considerable number of hospitals are forecasted not to reach Stage 6 by 2035  and 3. IStage 7 of EMRAM requires the development of EHR-derived centralized data warehouses along with extensive analytic infrastructure by hospitals. Although the need for data analytics has grown tremendously among health care providers over the last decade , the valAnother major milestone of EMRAM\u2019s Stage 7 for EHR maturation is the interoperability of EHRs among health care providers as well as integration of EHRs with local and regional health information exchanges . The chaBass model  has beenWe assumed no change in future policies or external factors that may affect EHR functionality advancements or health IT adoption generally  . New heaEMRAM does not include EHR adoption data for 2004 and 2005 when health IT policies started to take effect . As thisThis study only focuses on inpatient hospital settings and excludes the potential effect of EHR adoption trends in outpatient setting on hospitals. Future studies should investigate the interaction regarding adopting new EHR functionalities between inpatient and outpatient settings   to an environment where only electronic information is used to document and direct care delivery (Stage 7). According to the forecast, the majority of hospitals will not reach Stage 7 of EHR maturity by 2035, given that there are no major policy changes."}
+{"text": "Chronic Lung Allograft Dysfunction (CLAD) is the main cause of morbidity and mortality after the first year following lung transplantation (LTx). Risk factors of CLAD have been extensively studied, but the association between gram-negative bacteria (GNB) bronchial colonization and the development of CLAD is controversial. The purpose of our study was to investigate the association between post-transplant recolonization with the same species or de-novo colonization with a new GNB species and CLAD. The same analysis was performed on a sub-group of patients at the strain level using Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry technique.p\u2009=\u20090.04) and a lower rate of CLAD-free survival (p\u2009=\u20090.005) compared to patients with GNB recolonization. No conclusion could be drawn from the subgroup MALDI-TOF MS analysis at the strain level.Forty adult cystic fibrosis (CF) patients who underwent a first bilateral LTx in the University Hospital of Marseille, between January 2010 and December 2014, were included in the study. Patients with GNB de-novo colonization had a higher risk of developing CLAD  contains supplementary material, which is available to authorized users. Chronic lung allograft dysfunction (CLAD) is the major factor limiting long term survival following lung transplantation (LTx). The International Society of Heart and Lung Transplantation (ISHLT) evaluates the prevalence of bronchiolitis obliterans syndrome (BOS) at 50% at 5\u00a0years post transplantation .Previously known as \u201cchronic rejection\u201d, the term CLAD was coined in 2010 in order to encompass two distinct identities: BOS and restrictive allograft syndrome (RAS) . The undIn order to study bacterial populations, the use of Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) has shown its efficacy in the field of proteomics for the identification of routinely isolated micro-organisms when compared to conventional phenotypic identification . It was The objective of our study was to evaluate the impact of Gram Negative Bacteria (GNB) colonization of the lung allograft on the development of CLAD, in a population of patients with CF. Patients with GNB de-novo colonization are hypothesized to have a higher risk of CLAD development compared to patients without GNB de-novo colonization.A retrospective observational single-center study was conducted on a population of adult CF patients who underwent a first LTx or heart-LTx at the University Hospital of Marseille, France, between January 2010 and December 2014. Patients who were included in the analysis had at least two culture samples , bronchial aspirates (BA) or broncho-alveolar lavage (BAL)) during the postoperative period .For the subgroup analysis using MALDI-TOF MS technique, patients who did not have at least one specimen available both pre and post-LTx for the same bacterial species were excluded.Currently, there is no consensus regarding the definition of chronic pulmonary colonization . In thisWe also took into account risk factors recognized as associated with CLAD , 22, 23:Written informed consent was obtained from all patients. The study was approved by the local ethic committee (Assistance Publique H\u00f4pitaux de Marseille).All recipients received a standardized immunosuppressive regimen. Induction therapy consisted of intravenous administration of 1.5\u00a0mg/kg/day of rabbit anti-thymocyte globulins given for the first three postoperative days, associated with a high dose of methylprednisolone. Intravenous cyclosporine was administered immediately after LTx and was then switched to oral tacrolimus. Standard triple maintenance immunosuppressive regimen consisted of tacrolimus, mycophenolate mofetil and prednisone.Pneumocystis pneumonia.Postoperatively, recipients received a prophylactic antibiotic treatment according to their preoperative and/or concomitant infectious status, for at least 14\u00a0days. Seropositive CMV recipients received prophylactic IV ganciclovir for the first two postoperative weeks. Higher risk CMV-mismatched recipients (Donor+/Recipient-) were treated systematically for the first 3\u00a0months. Antifungal prophylaxis with voriconazole was used during the first month in case of previous fungal infection present in the 6\u00a0months preceding LTx. Our center did not practice routine prophylaxis against PFTs were performed according to the American Thoracic Society and European Respiratory Society guidelines . Lung fuThe diagnosis of CLAD was made on the PFTs according to the ISHLT\u2019s definition , 5. BOS Sputum samples were collected when clinically indicated. BAL and BA were obtained during a bronchoscopy, following current guidelines . SamplesBacterial colonies were identified using MALDI-TOF MS as previously described . AntimicP values <\u20090.05 were considered statistically significant.Categorical data were expressed in percentages and absolute values. Continuous data were expressed in means and standard deviations. Categorical data were compared using Fisher\u2019s Exact test in the univariate analysis. The multivariate analysis was done using a stepwise, multivariate, logistic regression model. Continuous data were compared using Mann-Whitney nonparametric tests. Survival analysis were performed using the Kaplan-Meier estimate and compared using the log rank test. Two sided During the study period, fifty-five CF adult patients underwent LTx in our transplant center. Of these patients, forty CF lung transplant recipients had at least two bronchial cultures available in the 6\u00a0months pre-transplant and at 1, 6 and/or 12\u00a0months post-transplant and were included in the study. All the patients included in the study were followed for more than a year, 34 patients for more than 2\u00a0years, 26 patients for more than 3\u00a0years and 21 patients for more than 4\u00a0years. A mean follow-up of 1413.5\u00a0days (\u00b1565.8) was reported. Baseline characteristics are reported in Table\u00a0n\u2009=\u200936) were colonized with Pseudomonas aeruginosa (PA). At one month, six months and twelve months post-LTx respectively 57.5% (n\u2009=\u200923), 64.5% (n\u2009=\u200920) and 51.5% (n\u2009=\u200917) were colonized with a GNB.Bacterial airways\u2019 colonization results are reported in Table\u00a0PA strains from these two periods showed that seven patients were colonized with the same PA strain, two patients were both recolonized and de- novo colonized and four patients had a new strain of PA. The two patients colonized with Pandorea pulmonicola (PP) in the study, were colonized with the same strain of PP pre and post-LTx.Of the forty patients included in the study, fourteen patients had at least one MALDI-TOF MS spectrum available pre and post-LTx, enabling the construction of dendrograms , de-novo colonization n\u2009=\u20097 (17.5%) and no measurable GNB colonization n\u2009=\u20095 (12.5%). The incidence of CLAD was highest in the group with de-novo colonization (p\u2009=\u20090.02) , and multivariate analysis  (Table The forty patients who met the aforementioned inclusion criteria were categorized into three exclusive groups: GNB recolonization without de-novo colonization on n\u2009=\u20097 7.5% and p\u2009=\u20090.005). There was no significant difference in overall survival between the 3 groups (p\u2009=\u20090.319)  Fig.\u00a0.Fig. 2Ovn\u2009=\u200914). There were no significant differences in overall survival (p\u2009=\u20090.346) and CLAD free survival (p\u2009=\u20090.107) between the three groups  secondary to lower survival. The results of this study concluded that GNB de-novo colonization is a risk factor for the development of CLAD . Moreover, GNB re-colonization conveyed an improved CLAD-free survival compared to the group GNB de-novo colonization (p\u2009=\u20090.005). These results corroborate findings from Botha et al. and Willner et al. studies, in which an association between GNB de-novo colonization and BOS was found [PA colonization is responsible for chronic airways inflammation, which might lead to the development of BOS [PA leads to loss of function and reduced virulence [The association between BOS and GNB post-transplant colonization has been debated in recent literature \u201311. In tas found , 36. Thet of BOS , 37, 38.t of BOS . Moreoveirulence \u201342.Several limitations have been identified in this study. Most of our pre-transplantation samples available were BSC\u2019s (92.5%), the close follow-up required after a lung transplant allowed us to use BAs and BAL for 80% of our patients at one month and 42% of our patients after one month. However the inconsistent use of BSCs, BAs, and BAL to determine colonization in the airways, increases the possibility of contamination as well as decreases sensitivity . Some stIn our study, at the species level, post-transplant GNB de-novo colonization was a risk factor for the development of CLAD and decreased CLAD free survival, compared to patients with post-transplant GNB recolonization. This stresses the importance of GNB species switching and the resulting immune response as a potential target in the prevention of CLAD. A larger longitudinal study with strain level typing either via MALDI-TOF MS or molecular biology is needed to confirm this association.Additional file 1:Pre and post-lung transplant airway colonization. Pre and post lung transplant airway colonization at the individual level for all forty patients. (DOCX 13\u00a0kb)Additional file 2:Pseudomonas aeruginosa dendrogram. MSP dendrogram performed using Biotyper v 3.0, including 52 spectra of Pseudomonas aeruginosa isolated before, at one month, six months or twelve months after lung transplant for the 13 patients for which data were available. * indicates isolates of the same patient belonging to the same strain. Each color is specific for one patient. (DOCX 120\u00a0kb)Additional file 3:Best post-lung transplant Forced expiratory volume in 1\u00a0s (FEV1) in the three groups: GNB recolonization, GNB de-novo colonization, exempt of GNB. Best post-lung transplant Forced expiratory volume in 1\u00a0s (FEV1) in the three groups: GNB recolonization, GNB de-novo colonization, exempt of GNB expressed in percentage of the expected value. (DOCX 13\u00a0kb)"}
+{"text": "Determine age-specific infection fatality rates for COVID-19 to inform public health policies and communications that help protect vulnerable age groups. Studies of COVID-19 prevalence were collected by conducting an online search of published articles, preprints, and government reports that were publicly disseminated prior to 18 September 2020. The systematic review encompassed 113 studies, of which 27 studies  satisfied the inclusion criteria and were included in the meta-analysis. Age-specific IFRs were computed using the prevalence data in conjunction with reported fatalities 4\u00a0weeks after the midpoint date of the study, reflecting typical lags in fatalities and reporting. Meta-regression procedures in Stata were used to analyze the infection fatality rate (IFR) by age. Our analysis finds a exponential relationship between age and IFR for COVID-19. The estimated age-specific IFR is very low for children and younger adults  but increases progressively to 0.4% at age 55, 1.4% at age 65, 4.6% at age 75, and 15% at age 85. Moreover, our results indicate that about 90% of the variation in population IFR across geographical locations reflects differences in the age composition of the population and the extent to which relatively vulnerable age groups were exposed to the virus. These results indicate that COVID-19 is hazardous not only for the elderly but also for middle-aged adults, for whom the infection fatality rate is two orders of magnitude greater than the annualized risk of a fatal automobile accident and far more dangerous than seasonal influenza. Moreover, the overall IFR for COVID-19 should not be viewed as a fixed parameter but as intrinsically linked to the age-specific pattern of infections. Consequently, public health measures to mitigate infections in older adults could substantially decrease total deaths.The online version of this article (10.1007/s10654-020-00698-1) contains supplementary material, which is available to authorized users. Since the onset of the COVID-19 pandemic in winter 2020, it has been evident that the severity of the disease varies markedly across infected individuals . Moreovecase fatality rate (CFR), the ratio of deaths to reported cases, are fraught with pitfalls in gauging the severity of COVID-19. For example, early case reports from Wuhan noted a preponderance of older people among hospital admissions and a high CFR [Consequently, assessments of the high CFR . Subsequhigh CFR , 6. Noneinfection fatality rate (IFR), the ratio of fatalities to total infections, thereby facilitating the identification of vulnerable segments of the population and informing key policy decisions aimed at mitigating the consequences of the pandemic [To provide more accurate assessments of the spread of COVID-19, researchers have conducted seroprevalence studies in numerous locations , 8. SuchFor example, as shown in Table Nonetheless, divergences in study design and reporting have hampered comparisons of seroprevalence and IFRs across locations and demographic groups. For example, a number of studies have analyzed a representative sample of the general population, while other studies have made use of \u201cconvenience samples\u201d of residual sera collected for other purposes (such as laboratory tests or blood donations) \u201314. Someurgently needed to inform policymaking.\u201d [While the NYC data indicate a population IFR of about 1%, seroprevalence estimates from other locations have yielded a wide array of population IFR estimates, ranging from about 0.6% in Geneva to levels exceeding 2% in northern Italy. Such estimates have fueled intense controversy about the severity of COVID-19 and the appropriate design of public health measures to contain it, which in turn hinges on whether the hazards of this disease are mostly limited to the elderly and infirm. Indeed, a recent meta-analysis noted the high degree of heterogeneity across aggregate estimates of IFR and concluded that research on age-stratified IFR is \u201cmaking.\u201d This paper reports on a systematic review and meta-analysis of age-specific IFRs for COVID-19. We specifically consider the hypothesis that the observed variation in IFR across locations may primarily reflect the age specificity of COVID-19 infections and fatalities. Based on these findings, we are able to assess and contextualize the severity of COVID-19 and examine how age-specific prevalence affects the population IFR and the total incidence of fatalities.To perform the present meta-analysis, we collected published papers and preprints on the seroprevalence and/or infection fatality rate of COVID-19 that were publicly disseminated prior to 18 September 2020. As described in Supplementary Appendix B, we systematically performed online searches in MedRxiv, Medline, PubMed, Google Scholar, and EMBASE, and we identified other studies listed in reports by government institutions such as the U.K. Parliament Office . Data waWe restricted our meta-analysis to studies of advanced economies, based on current membership in the Organization for Economic Cooperation and Development (OECD), in light of the distinct challenges of health care provision and reporting of fatalities in developing economies . We alsoOur meta-analysis encompasses two distinct approaches for assessing the prevalence of.COVID-19: (1) seroprevalence studies that test for antibodies produced in response to the virus, and (2) comprehensive tracing programs using extensive live-virus testing of everyone who has had contact with a potentially infected individual. Seroprevalence estimates are associated with uncertainty related to the sensitivity and specificity of the test method and the extent to which the sampling frame provides an accurate representation of prevalence in the general population; see Supplementary Appendix C. Prevalence measures from comprehensive tracing programs are associated with uncertainty about the extent of inclusion of infected individuals, especially those who are asymptomatic.Blood donors. Only a small fraction of blood donors are ages 60 and above\u2014a fundamental limitation in assessing COVID-19 prevalence and IFRs for older age groups\u2014and the social behavior of blood donors may be systematically different from their peers [ir peers , 18. Their peers .Dialysis centers. Assessing seroprevalence of dialysis patients using residual sera collected at dialysis centers is crucial for gauging the infection risks faced by these individuals, of which a disproportionately high fraction tend to be underrepresented minorities. Nonetheless, the seroprevalence within this group may be markedly different from that of the general population. For example, a study of U.K. dialysis patients found seroprevalence of about 36%, several times higher than that obtained using a very large random sample of the English population [pulation , 22. Simpulation , 23.Hospitals and urgent care clinics. Estimates of seroprevalence among current medical patients are subject to substantial bias, as evident from a pair of studies conducted in Tokyo, Japan: One study found 41 positive cases among 1071 urgent care clinic patients, whereas the other study found only two confirmed positive results in a random sample of nearly 2000 Tokyo residents  [s. 0.1%) , 25.Active recruitment. Soliciting participants is particularly problematic in contexts of low prevalence, because seroprevalence can be markedly affected by a few individuals who volunteer due to concerns about prior exposure. For example, a Luxembourg study obtained positive antibody results for 35 out of 1807 participants, but nearly half of those individuals (15 of 35) had previously had a positive live virus test, were residing in a household with someone who had a confirmed positive test, or had direct contact with someone else who had been infected [infected .To assess prevalence in the general population, a study should be specifically designed to utilize a random sample using standard survey procedures such as stratification and weighting by demographic characteristics. Other sampling frames may be useful for specific purposes such as sentinel surveillance but not well-suited for assessing prevalence due to substantial risk of systemic bias. Consequently, our meta-analysis excludes the following types of studies:Our critical review has also underscored the pitfalls of seroprevalence studies based on \u201cconvenience samples\u201d of residual sera collected for other purposes. For example, two studies assessed seroprevalence of Utah residents during spring 2020. The first study analyzed residual sera from two commercial laboratories and obtained a prevalence estimate of 2.2% (CI: 1.2\u20133.4%), whereas the second study collected specimens from a representative sample and obtained a markedly lower prevalence estimate of 0.96% (CI: 0.4\u20131.8%) , 28. In Our meta-analysis incorporates data on COVID-19 prevalence and fatalities in countries that have consistently maintained comprehensive tracing programs since the early stages of the pandemic. Such a program was only feasible in places where public health officials could conduct repeated tests of potentially infected individuals and trace those whom they had direct contact. We identify such countries using a threshold of 300 for the ratio of cumulative tests to reported cases as of 30 April 2020, based on comparisons of prevalence estimates and reported cases in Czech Republic, Korea, and Iceland; see Supplementary Appendices D and E . StudiesAccurately measuring total deaths is a substantial issue in assessing IFR due to time lags from onset of symptoms to death and from death to official reporting. Symptoms typically develop within 6\u00a0days after exposure but may develop as early as 2\u00a0days or as late as 14\u00a0days . More thFigure\u00a0As shown in Table Therefore, we construct age-specific IFRs using the seroprevalence data in conjunction with cumulative fatalities 4\u00a0weeks after the midpoint date of each study; see Supplementary Appendix F. We have also conducted sensitivity analysis using cumulative fatalities 5\u00a0weeks after the midpoint date, and we flag studies as having an elevated risk of bias if the change in cumulative fatalities between weeks 4 and 5 exceeds 10%.By contrast, matching prevalence estimates with subsequent fatalities is not feasible if a seroprevalence study was conducted in the midst of an accelerating outbreak. Therefore, our meta-analysis excludes seroprevalence studies for which the change in cumulative fatalities from week 0 to week 4 exceeds 200%.meta regress procedure in Stata v16 [To analyze IFR by age, we use meta-regression with random effects, using the tata v16 , 40. In Analyze whether the metaregression coefficients exhibit any significant differences across three broad age categories ;Analyze whether the results are sensitive to exclusion of the oldest age group in each location , given that such groups may span a relatively wide age range and hence not fully capture their vulnerability to the virus;Conduct out-of-sample analysis using small-scale seroprevalence studies as well as studies not included in the metaregression due to overlapping geographical regions;Compare the actual population IFR in each location  with the population IFR predicted by the metaregression .To assess the robustness of the metaregression results, we conduct several forms of sensitivity analysis:Finally, publication bias is assessed using Egger\u2019s regression and the trim-and-fill method.As shown in fig. Representative samples from studies of England, France, Ireland, Italy, Netherlands, Portugal, Spain, Geneva (Switzerland), and four U.S. locations  [ke City) , 118\u2013127Convenience samples from studies of Belgium, Sweden, Ontario (Canada), and eight U.S. locations  [Seattle) , 128\u2013130Comprehensive tracing programs for Australia, Iceland, Korea, Lithuania, and New Zealand [ Zealand \u2013135.Consequently, our meta-analysis encompasses 27 studies of 34 geographical locations, of which 28 are included in our metaregression and 6 are used for out-of-sample analysis. The metaregression observations can be categorized into three distinct groups:The metaregression includes results from the very large REACT-2 seroprevalence study of the English population . Thus, tp\u2009>\u20090.10), and the trim-and-fill method produced the same estimate as the metaregression.Data taken from included studies is shown in Supplementary Appendix I. Supplementary Appendix J assesses the risk of bias for each individual study. As indicated in Supplementary Appendix K, no publication bias was found using Egger\u2019s test  and I2\u2009=\u200997.0, confirming that the random effects are essential for capturing unexplained variations across studies and age groups. The adjusted R2 is 94.7%.where the standard error for each estimated coefficient is given in parentheses. These estimates are highly significant with t-statistics of \u221244.5 and 40.4, respectively, and As noted above, the validity of this metaregression rests on the condition that the data are consistent with a Gaussian distribution. The validity of that assumption is evident in Fig.\u00a0This specification of the metaregression also assumes that the intercept and slope parameters are stable across the entire age distribution. We have confirmed the validity of that assumption by estimating alternative specifications in which the parameters are allowed to differ between three distinct age categories . The estimated parameters are similar across all three age categories, and the null hypothesis of parameter constancy is consistent with the metaregression data. We have also confirmed that the metaregression results are not sensitive to exclusion of open-ended top age groups. (See Supplementary Appendix L for details.)Figure\u00a0As shown in Fig.\u00a0The IFR is central to our understanding of the public health impact of the COVID-19 pandemic and the appropriate policies for mitigating those consequences. In the absence of effective therapies or vaccines, such policies will primarily involve non-pharmaceutical interventions (NPIs). NPIs may include relatively mild measures (such as prohibitions on large gatherings) or more draconian restrictions such as shelter-in-place edicts, popularly known as \u201clockdowns.\u201dUnfortunately, public debate on these issues has been hampered by diverging assessments of the severity of COVID-19. For example, some early seroprevalence studies  yielded miniscule estimates of population IFR similar to those of seasonal influenza. Such estimates implied that strict NPIs would be completely irrational given the limited benefits and severe economic and social costs. With the dissemination of many more seroprevalence studies over recent months, a wide array of hypotheses have been mooted to explain the diverging implications for IFR, including regional variations in the quality of treatment or the extent of T-cell immunity to other betacoronaviruses.By contrast, our critical review identifies the key characteristics of seroprevalence studies that can be used to provide reliable assessments of IFR. Indeed, once we focus on this group of studies , our metaregression reveals a remarkably high degree of consistency in the implications for age-specific IFR. Moreover, our results indicate that most of the variation in population IFR across locations reflects differences in the extent to which vulnerable age groups were exposed to the virus.One key implication of our findings is that the incidence of fatalities from a COVID-19 outbreak depends crucially on the age groups that are infected, which in turn reflects the age structure of that population and the extent to which public health measures limit the incidence of infections among vulnerable age groups . Indeed,To gauge the benefits of age-stratified public health strategies for COVID-19, we have constructed two illustrative scenarios for the U.S. trajectory of infections and fatalities (see Supplementary Appendix N). Each scenario assumes that U.S. prevalence rises to a plateau of around 20% but with different patterns of age-specific prevalence. In particular, if prevalence becomes uniform across age groups, this analysis projects that total U.S. fatalities would rise to nearly 900 thousand and that population IFR would converge to around 1.3%. By contrast, a scenario with relatively low incidence of new infections among vulnerable age groups would be associated with a much lower number of fatalities (about 350 thousand) and a correspondingly lower population IFR of about 0.5%.A further implication of our results is that the risks of infection to the middle aged cannot be neglected. This is important for pandemic management strategies that aim to avoid large influxes of patients to healthcare. Indeed, it is likely that an unmitigated outbreak among middle-aged and older adults could have severe consequences on the healthcare system.Public health communications can be helpful for persuading individuals to take steps to mitigate the risk of infection for themselves as well as others with whom they have direct contact . For this purpose, it is helpful to contextualize the magnitude of age-specific IFRs for COVID-19 relative to annualized fatality rates for other routine activities; that annual timeframe reflects the premise that effective vaccines and/or treatments for COVID-19 would hopefully become widely available sometime within the next year or two.In particular, Table Our critical review highlights the benefits of assessing prevalence using large-scale studies of representative samples of the general population rather than convenience samples of blood donors or medical patients. Conducting such studies on an ongoing basis will enable public health officials to monitor changes in prevalence among vulnerable age groups and gauge the efficacy of public policy measures. Moreover, such studies enable researchers to assess the extent to which antibodies to SARS-CoV-2 may gradually diminish over time as well as the extent to which advances in treatment facilitate the reduction of age-specific IFRs.Our critical review also underscores the importance of methodological issues in assessing IFR. For example, the raw prevalence results reported by a national study of Italy would imply a population IFR of about 2.3%, whereas test-adjusted prevalence implies a substantially higher IFR of 2.7%. Likewise, a few recent studies have excluded all deaths occurring in nursing homes and retirement communities and have obtained estimates of population IFR that are markedly lower than our estimates based on all confirmed COVID-19 fatalities, whereas assessments of IFR based on measures of excess mortality are broadly similar to our estimates , 145\u2013147Our metaregression results are generally consistent with the study of Verity et al.  and FergOur findings are well-aligned with a recent meta-analysis of population IFR and indeed explain a high proportion of the dispersion in population IFRs highlighted by that study . In contThe exponential pattern of our age-specific IFR estimates is qualitatively similar to that of case fatality rates (CFRs). However, the relative magnitudes are systematically different, reflecting the extent to which asymptomatic or mildly symptomatic cases are much more common in younger adults than in middle-aged and older adults. For example, the ratio of CFR to IFR is about 15:1 for ages 30\u201349, about 7:1 for ages 50\u201369, and about 5:1 for ages 70\u201379\u00a0years (see Supplementary Appendix R).A potential concern about measuring IFR based on seroprevalence is that antibody titers may diminish over time, leading to underestimation of true prevalence and corresponding overestimation of IFR. This concern is particularly relevant for seroprevalence studies conducted many months after the outbreak was contained . Howevernot seroprevalence) for five countries that have maintained comprehensive tracing programs since the onset of the pandemic, namely, Australia, Iceland, Korea, Lithuania, and New Zealand. As shown in Fig.\u00a0Moreover, a key feature of our metaregression analysis is that we also utilize age-specific IFR data based on RT-PCR results Supplementary file2 (XLSX 56 kb)Below is the link to the electronic supplementary material."}
+{"text": "Staphylococcus aureus to circumvent it, and the impact of deletion mutants on the fitness, infectivity, and persistence inside the host. This information finally converges in an overview of the current development of inhibitors targeting the different stages of iron uptake, an as-yet unexploited target in the field of antistaphylococcal drug discovery.Nutritional immunity is a form of innate immunity widespread in both vertebrates and invertebrates. The term refers to a rich repertoire of mechanisms set up by the host to inhibit bacterial proliferation by sequestering trace minerals . This strategy, selected by evolution, represents an effective front-line defense against pathogens and has thus inspired the exploitation of iron restriction in the development of innovative antimicrobials or enhancers of antimicrobial therapy. This review focuses on the mechanisms of nutritional immunity, the strategies adopted by opportunistic human pathogen  Borrelia burgdorferi . IsdA, IsdB, IsdC, and IsdH  are bouS. aureus for virulence and proliferation in mouse model ..S. aureuS. aureus, weakening the host antimicrobial response, and, more importantly, are unable to adapt their iron acquisition strategy in response to the inflammation-dependent restriction [S. aureus as a commensal or a pathogen [Staphylococcal fitness in host invasion is partly attributed to the proficiency in iron acquisition through multiple mechanisms. Each of them relies on protein complexes, employing at least one ABC-transporter importing iron across the membrane, an ATPase and a membrane-anchored lipoprotein ,201. Thitriction . In addipathogen .In the present section, we review the iron-related systems that can be targeted, or exploited, for the development of possible antimicrobials and the strategies investigated up to now. The section has been divided into paragraphs that report targets belonging to pre-iron-uptake systems (secretion of hemolysins), iron-uptake systems (siderophores and iron mimetics), and post-iron-uptake systems (heme degradation).S. aureus undergoes a transformation from harmless to virulent for the host organism [agr) locus, part of a QS system [agr operon consists of four genes (agrBDCA) that encode for the respective AgrBDCA proteins. For a more detailed description of this QS system please refer to [The production and secretion of hemolysins is part of a more generic mechanism, called quorum-sensing (QS), by which organism ,206. HlaS system . The agrrefer to ,209,210.S. aureus (MRSA) infections. Indeed, the combination of an antibiotic with an antivirulence agent constitutes a potential new mode of treatment that may alleviate the antibiotic resistance crisis. Administration of quorum-sensing inhibitors (QSI) or quorum-quenching agents (QQA) leads to inhibition of virulence factors expression, which makes bacteria less aggressive and more susceptible to natural immunity. Antivirulence agents are neither bactericidal nor bacteriostatic and in principle should be less susceptible to resistance. Many efforts have been done in that direction and, even though there is currently no antivirulence agent in clinical use, several studies show the usefulness of these compounds, alone or as adjuvants in conventional antibiotic therapy for methicillin-resistant agr operon and the toxin production can be achieved by targeting any component of the system. The most exploited strategies are reported hereafter.Ideally, inhibition of the agr operon, and the bacterial promoter has been extensively studied. The AgrA C-terminal (AgrAC) DNA-binding domain LytTR was identified as a druggable site, with the aim to inhibit the engagement with the bacterial DNA. In the same study, a screening of 500 fragment compounds led to the identification of three molecules able to inhibit AgrA DNA binding activity in the range of 10\u22124 and thus can be considered a good starting point for further drug development [The interaction between the cytoplasmic protein AgrA, acting as a transcription factor of elopment ,212,213.A series of biarylhydroxyketones was discovered in 2013 to inhibit in vitro MRSA-triggered erythrocyte hemolysis by 98% at a concentration of 1 \u00b5g/mL ,215. TheS. aureus virulence inhibitor) as an inhibitor of the agr-specific transcription regulation of the major virulence factors, such as Hla, with promising outcomes. Interestingly, the compound was also shown to not be subjected to tolerance or resistance mechanisms. The same group also investigated a series of nine polyhydroxyantraquinones, produced by Penicillium restrictum, able to bind the AgrAC DNA binding region, among which \u03a9-hydroxyemodin (OHM) was found to reduce dermonecrosis in mouse model of MRSA skin infection [agr activation through AgrA inhibition [In 2014, Sully and colleagues  identifinfection ,221. Thehibition . Finally, mimetics of the autoinducing peptides (AIPs), which are fundamental for assessing bacterial population density and interact with AgrC receptors ,224, havOne concern in the clinical use of QSI is that their efficacy is related to the immune system integrity , meaning that in immunocompromised patients the only use of QSI could not be enough to overcome an infection. Few hemolysin inhibitors have been reported till now, with mechanistic details mainly explained by means of in silico studies. According to their mechanism, hemolysin inhibitors may be divided into two groups: (i) hemolysin monomer binders, which hinder the oligomerization process, and (ii) direct blockers of the pore formed by the oligomerized Hla subunits. Natural compounds such as baicalin, cyrtominetin and oroxylin A ,232,233,A number of iron-chelating agents, clinically useful in treating iron-overload conditions, have been evaluated for their ability to inhibit bacterial growth in multidrug-resistant bacteria, mainly by competing with bacterial siderophores for available iron.\u00ae, Novartis), a hexadentate hydroxamate siderophore derived from Streptomyces pilosus, was one of the first compounds to be investigated as an inhibitor of the in vitro growth of various Staphylococci, including S. aureus. However, Staphylococci appeared to give different responses to DFO treatment, with the growth of many S. aureus strains being actually enhanced [DFO , a bidentate 3- hydroxypyridin-4(1H)-one  chelator -pyridinone , linked to a triaza-macrocyclic backbone scaffold to give a hexadentate chelator, was also evaluated for its inhibitor activity against Gram-negative and Gram-positive bacteria, among which . aureus .S. aureus possesses to supply iron deficiency, including the expression of the Isd system proteins.Despite their unquestionable ability to chelate and thus eliminate the free iron as a source of bacterial nutriment, these compounds exhibit relevant MICs decrease only in combination with antibiotics. This could be attributed to the many redundant mechanisms that Siderophores have been envisaged as \u201cTrojan horses\u201d, for their capacity of entering bacteria and deliver therapeutic agents conjugated to the siderophore molecule . Even ifStreptomyces strains [This strategy has been first exploited by the same bacteria for delivering antibiotics in competing organisms able to internalize xenosiderophores. These conjugated compounds, constituted by a siderophore and a covalently bound antibiotic, were identified for the first time in 1947 in  strains , before  strains . Albomycins, the first discovered sideromycins, present a ferrichrome-like trihydroxamate siderophore linked to a seryl-thioribosyl pyrimidine. They inhibit seryl-tRNA synthetase and have a broad-spectrum activity towards both Gram-positives and Gram-negatives, with MIC in the order of 5 ng/mL ,256. TheStreptomyces violaceus strain [S. aureus is not so high and the compound has to be administered at shorter intervals with respect to other antibiotics as vancomycin and rifamycin, possibly because of the instability of the ester linkage [Salmycins were later isolated from a s strain . They ars strain . Again, s strain . Unfortu linkage . Salmyci linkage ,260. TheFerrimycins, finally, are constituted by a ferrioxamine B moiety, conjugated to an antibiotically active group, and specifically target Gram-positives .As mentioned, despite the quite different nature of the antibiotic moiety, both albomycins and salmycins are transported by the same siderophore transport proteins. S. aureus and E. coli, because of permeability issues [E. coli and Salmonella strains [Sideromycins are equally expressed and potent in Gram-negatives, in which the outer membrane does not represent a permeability barrier, but actively facilitates their transport. For many antibiotics, in fact, the diffusion through the outer membrane is so poor that the MIC reaches toxic levels. Exploiting the siderophore transport system often leads to a 100-fold MIC reduction . Indeed,y issues . As well strains .As for other antimicrobials, the same bacteria can become resistant to sideromycins, generally because of mutations at the level of the siderophore receptor or of the TonB transport complex. The capability of these conjugates to easily cross the bacterial membrane was soon exploited in antimicrobial research, and synthetic compounds mimicking natural sideromycins were designed using a \u201cTrojan horse\u201d strategy. These molecules contain three components, a natural or mimetic iron-chelating siderophore, a linker, and an antibiotic. With respect to canonical antibiotics, these siderophore\u2013antibiotic conjugates generally present an improved selectivity, since each bacterium produces and uses different types of siderophores. A higher specificity goes in the direction of more responsible use of antibiotics, because of a reduced risk of antibiotic resistance. Other possible advantages are related to the enhanced antibacterial potency and the selectivity for pathogenic strains over non-pathogenic ones, accomplished by the transformation of Gram-positive antibiotics in Gram-negative antibiotics, which might render multiple drug-resistance bacteria more susceptible .Thanks to the structural tolerance displayed by siderophore transporters, artificial siderophores, and conjugates quite different from the parent structures can be transported as the natural siderophores in the targeted bacterial cells. However, the best strategy for the development of a successful synthetic conjugate relies on the replication of natural siderophores ,267. AccS. aureus [Staphylococcus and Enterococcus spp., when compared to the 5-fluorocytosine active metabolite alone [Among the first to design semisynthetic siderophore\u2013antibiotic conjugates, Zahner and coworkers joined a sulfonamide antibiotic to ferrioxamine B analogs, obtaining compounds with a limited spectrum of action against . aureus . Later, . aureus . Differete alone .S. aureus [Enterococcus faecium, S. aureus, Klebsiella pneumoniae, Acinetobacter baumannii, P. aeruginosa, and Enterobacter species). Moreover, the mono- and bis-hydroxamate fluoroquinolone conjugates also had a reduced activity spectrum and a reduced potency with respect to ciprofloxacin, while the trihydroxamate derivative showed selectivity towards Gram-positive S. aureus, maintaining a similar potency in terms of MIC (1 mM with respect to 0.5 mM). The compound, thus, is one of the very few synthetic siderophore\u2013antibiotic conjugates capable of maintaining the activity of the original antibiotic. The narrower spectrum of action must be attributed to the trihydroxamate siderophore, also known as desferridanoxamine, better recognized, with respect to the mono- and bis derivatives, by FhuD1 and FhuD2, the promiscuous membrane-anchored hydroxamate siderophore binding proteins of S. aureus. Gram-negatives have more selective siderophore binding outer-membrane proteins, possibly unable to recognize these derivatives. The lack of activity of \u03b2-lactam conjugates could be attributed to the location of the biological target. As the compounds are transferred by active transport into the cytoplasm, a target as PBPs located in the plasma membrane is not the ideal one. Thus, when targeting Gram-positives with siderophore\u2013antibiotics conjugates, cytoplasmic targets should be preferred. The capability of the trihydroxamate-ciprofloxacin derivative to reach the target by means of active transport was also demonstrated by the dependence of the antibacterial activity on iron concentration and the presence of siderophores competing for the same transport mechanism. As stated by the authors, two main advantages can be associated with the development and use of trihydroxamate-ciprofloxacin derivatives. First, the selectivity of the compounds towards S. aureus makes them ideal candidates for targeted antimicrobial chemotherapy, which limits the exposure of other bacteria to ciprofloxacin and, thus, is less prone to generate multidrug resistance. Moreover, the compound intrinsic toxicity towards mammalian cells is reduced because siderophore transporters are absent in eukaryotic cells. On the other side, the limits associated with \u201cTrojan horse\u201d compounds could be represented by a reduced efficacy in treating intracellular infections because of the incapability of passing eukaryotic cell membrane. Also, the use of a very selective treatment implies an early detection of the infective agent, with respect to the use of broad-spectrum antibiotics.Inspired by natural salmycins, Wencwicz and colleagues designed a series of linear hydroxamate siderophore-fluoroquinolone conjugates to target . aureus . The autS. aureus, that is SA [In the same period, Milner and coworkers attempted to join the fluoroquinolones ciprofloxacin and norfloxacin to the same siderophore produced by at is SA . With reS. aureus. The control compound was, instead, not active, suggesting it was no more recognized by the membrane siderophore receptors. MIC analyses confirmed a moderate antibacterial activity against all the strains with the exception of E. faecium. However, in general, the authors observed a decreased activity with respect to the parent antibiotic and attributed it to the compounds being poor substrates for esterases. To overcome the risk that the conjugates could be cleaved by extracellular hydrolases, the same authors proposed similar conjugates bearing a linker cleavable by ferric reductases, instead of esterases or phosphatases [When designing a new siderophore-conjugate, Ji and Miller chose DFO as siderophore, ciprofloxacin as antibiotic and a trimethyl-lock based linker, specifically designed to promote the antibiotic release by means of esterase or phosphate-mediated hydrolysis . The triphatases . Upon trThe failure of \u201cTrojan horses\u201d strategies could be attributed to the same specific uptake systems that make these conjugates particularly active. Indeed, highly modified conjugates could result more difficult to be transported than native compounds. The presence in bacterial cells of several redundant iron-uptake systems, able to switch off when needed without affecting the cell survival, represents an additional failure reason, as well as the necessity to further release from the conjugate the antibiotic, by means of peptidase- or esterase-mediated cleavage . FortunaGallium-based molecules have been also classified as \u201cTrojan Horses\u201d for their capacity of being recognized by the iron transport system. Good antibacterial activities have been shown by simple gallium salts as nitrate (GaN) or maltolate (GaM) ,278,279,Fe(III) and Ga(III) have a significant chemical similarity, concerning ionic radius, electronegativity, ligand affinities, and coordination geometry . AccordiDespite the high similarity, however, Ga(III) cannot be reduced in physiological conditions; thus, any biological process involving an iron redox reaction, as electron transport or oxidative stress defense, can be disrupted by the iron-gallium substitution ,285. AlsP. aeruginosa and Gram-positives as S. aureus. It is well distributed in the body thanks to its binding and transport by transferrin, and easily taken up by bacteria through the iron transport system. Moreover, it has been proved capable of disrupting and preventing the formation of biofilms [S. aureus. Gallium nitrate has been extensively tested towards both Gram-negatives as biofilms . More cobiofilms , pyridonbiofilms , acinetobiofilms , and nonbiofilms . The comYersinia enterocolitica and Mycobacterium smegmatis, while that including zinc and ruthenium demonstrated prominent activity against S. aureus [The most promising MPs were first described by Stojiljkovic and colleagues in 1999 . Once ta. aureus . The mosS. aureus (MSSA) and MRSA [S. aureus colony variants [Richter and coworkers recently demonstrated that the combined treatment with Ga-PPIX and the iron chelator deferiprone significantly reduces the growth of methicillin-susceptible and MRSA , and smavariants . Apart fvariants ,290,291.variants , which svariants .Along with Ga-PPIX, also Ga-deuteroporphyrins, Ga-mesoporphyrins, Ga-hematoporphyrins, Ga-octaethylporphyrins, and Ga-porphyrins have provided interesting activity towards MRSA . BacteriS. aureus and A. baumannii strains. In particular, GaN and GaM showed a bacteriostatic effect, while Ga-PPIX had a bactericidal activity on some strains. Interestingly, the latter demonstrated to be susceptible to the number of bacterial heme uptake systems and the presence of serum albumin, able to bind a variety of ligands, including heme, and to inhibit their activity. They also confirmed the susceptibility of all derivatives to the iron concentration (enhanced by iron deprivation) [S. aureus strains. Overall, being active even in the presence of serum albumin, GaN and GaM derivatives were the most effective in in vivo mimicking conditions. It is interesting to note that Ga(III) derivatives already proved to be effective in infections caused by Gram-negatives [More recently, the antimicrobial activity of Ga-containing compounds was confirmed by Hijazi and coworkers towards the ESKAPE pathogens, including reference strains and clinical multidrug-resistant (MDR) ones . The autivation) , and of egatives ,294,295.S. aureus [S. aureus heme detoxification system. Upon exposure to heme toxic levels, in fact, the heme sensor system HssRS is activated and the heme-regulated transporter HrtAB is expressed [Apart from Ga-PPIX, Mn-PPIX and Zn-PPIX also exhibit antimicrobial activity against . aureus . As alrexpressed . Howeverxpressed .+ substituted porphyrin were evaluated against S. aureus and E. coli strains in dark and light conditions, showing a lower activity in dark conditions. This supported the photodynamic mode of action of the compounds [Photoactive porphyrins have been also used as photosensitizers for the photodynamic inactivation of microorganisms, based on the generation of singlet oxygen species and free radicals, upon the activation of the porphyrins by visible light . A catioompounds . Their eOverall, Ga-derivatives might represent a powerful resource for the development of new antimicrobials, to be administered alone or in combination with conventional antibiotics, as recently demonstrated by Richter and coworkers .In principle, small molecules inhibiting the iron-uptake system may target either siderophore receptors or hemophores. SA and SB receptors  bind their ligands with nanomolar affinity, so that competitive inhibition is precluded. As for xenosiderophores receptors, up to now, no work has been carried out for investigating whether SstD, or FhuD1 and FhuD2, are druggable with small molecules. Nonetheless, staphyloferrins and xenosiderophores receptors have been addressed as promising antigens for staphylococcal vaccines  see Sec.S. aureus, Bacilli, and Listeria. However, deletion of SrtB has only mild effects on S. aureus virulence . SimilaRecent studies ,110,325 Biologic products against hemolysins make up the second substantial part of biopharmaceuticals against the staphylococcal iron system and consist of either immunotherapeutics or vaccine candidates. Significantly, in vivo tests demonstrated avirulence of staphylococcal strains deficient in hemolysin production .S. aureus infections [S. aureus-induced pneumonia in mice. Furthermore, anti-Hla mAbs have been successfully combined with last-generation antibiotics, such as vancomycin and linezolid, enhancing or synergizing their activity [The literature focuses on neutralizing antibodies against hemolysins (Hla in particular) as a possible platform for the development of active or passive immunization therapies against fections ,330,331.fections  demonstractivity ,331. Expactivity . IBT-V02activity .S. aureus-induced nosocomial pneumonia, namely suvratoxumab , that proved to be safe and effective in a phase II study (NCT02296320) [Currently, there are two vaccine candidates against Hla as promising adjunct treatments for 2296320) , and tos2296320) .The interaction between Hla and a human-derived antigen-binding fragment (Fab) has been characterized by Foletti et al. , who repOverall, other hemolysins aroused far less interest. These include a neutralizing antibody targeting Hlb reported by Pooja and coworkers  and a biClostridium difficile. The main issue affecting the development of effective new antimicrobials is related to the presence of several redundant iron-uptake systems that can switch on and off according to external stimuli, without affecting bacteria survival. Targeting the pre-iron-uptake system through QS inhibitors could represent a valuable strategy, in particular, if antivirulence agents are administered in combination with antibiotics. Still, QS inhibitor activity is strictly related to the human immune system integrity. Hemolysin inhibitors, even if preventing hemolysis, do not have anti-staphylococcal activity and iron chelators are, again, effective only in combination with antibiotics, since S. aureus can acquire iron in different ways. The iron uptake system has been widely investigated as a possible target, mainly through the design of Trojan horses, originally developed by bacteria to counteract competing organisms. These molecules could be ideal candidates for their high selectivity and reduced toxicity. Even if highly modified conjugates have resulted difficult to be transported, the available chemical space is large and more effective combinations can be evaluated in the future. Similarly, also Gallium derivatives can be recognized by the iron transport system and have proved their antimicrobial activity when administered alone or in combination with conventional antibiotics. Siderophores and free iron uptake are not the only targetable systems. Indeed, IsdB and IsdH hemophores represent another essential way of iron acquisition and, accordingly, a promising and yet unexplored resource for the design of new antimicrobials affecting heme scavenging. In the post-iron-uptake metabolism, heme oxygenases, in particular IsdG, offer interesting perspectives too. Above all, biopharmaceuticals represent, today, the most advanced and concrete possibility. Apart from the yet unexplained failure of V710, the monovalent vaccine against IsdB, two vaccine candidates, suvratoxumab and tosatoxumab, targeting hemolysin H1a, have proved their efficacy and safety in phase II clinical trials, and represent the most promising perspective in the fight against S. aureus. Finally, the development of new in vitro/in vivo models to test the efficacy of molecules in the early stages of development will be of great value to allow the identification and the fast progression in the discovery pipeline of the more promising candidates.Targeting the iron-acquisition systems shows promise in the discovery of new anti-staphylococcal agents since the approach should in principle allow to decrease the chance for resistance development and selectively hit pathogenic bacteria without disturbing host microbiota or promoting the insurgence of opportunistic infections by"}
+{"text": "BackgroundThe frequency of radiological surveillance after curative colorectal cancer resection has long been a controversial issue with the need to balance potential harm from ionizing radiation and the financial burden of intense surveillance against advantages of early detection of recurrent disease.\u00a0NICE guidelines issued in 2018 suggested having two surveillance computed tomography (CT) scans within three years of surgery without specifying the timing or the interval.AimTo examine whether an evidence-based flexible approach based on individual patients\u2019 risk factors can add value to surveillance protocols.\u00a0Reaching a targeted protocol that can maximize early detection of metastasis without consumption of resources and most important without compromising patient safety.MethodologyA retrospective study involving five years of data of patients who underwent curative colorectal cancer resections.\u00a0Data extracted after patients completed their three-year surveillance CT scans, CT reports retrieved together with post-operative histology reports, and a detailed database was constructed.ResultsOf 179 patients included, 66 developed recurrence . Recurrence increased from 23.5% in T1 to 66% in T4 (P=0.0001). The median time to recurrence 23 months in T4 disease compared to 36, 42 and 43 months for stages T1, T2 and T3, respectively (P=0.0001). A similar incremental increase in recurrence noted from 22% in the N0 stage to 73.5% in the N2 stage (P=0.0001); the median time to recurrence of 14 months in N2 patients compared to 45 and 33 months for stages N0 and N1, respectively (P=0.0001). Recurrence correlated well with positive extramural vascular invasion (EMVI) status, (71.7% versus 19.3% P=0.0001) being detected significantly earlier in EMVI positive group\u00a0at 17 versus 45 months (P=0.0001).ConclusionFlexible protocol for radiological surveillance after curative resection of colorectal cancer, based on known pathological prognostic factors, is likely to be more effective in maximizing resource utilization as well as improving patient outcomes. Regular surveillance of patients following curative resection of colorectal cancer is crucial to maintain a high standard of follow-up care, ensuring early identification of any local recurrence or metastatic disease to enable further treatment to be offered.\u00a0Long-standing debates around an appropriate follow-up protocol have involved, striking a balance between potential benefits of early detection of local or distant recurrence and the cost as well as potential harm to patients from ionizing radiation from multiple CT scans on the other hand.NICE guidelines  issued iOur departmental protocol, introduced in 2001, involves a clinical examination and carcino-embryonic antigen (CEA) levels every six months for five years, annual CT-TAP for three years and colonoscopy at three and six years post-operatively after all curative resections for colorectal cancer.AimsTo examine whether an evidence-based flexible surveillance protocol, tailored to individual patients\u2019 risk factors can be developed to maximize early detection of local or distant recurrence, without compromising patient safety.MethodologyThis was a retrospective study involving five years\u2019 data of colorectal cancer resections (2010-2014) at Prince Charles Hospital; data were\u00a0collected after the patients had completed their three annual CT-TAP, as per department protocol. The search was extended by a further year to capture any delayed investigations for a variety of reasons such as service pressures, human errors, patient delay due to unforeseen circumstances, etc.A detailed database was constructed which included specific details of known prognostic factors in the post-operative pathology report including tumour staging (TNM classification - UICC 7th edition), extramural vascular invasion (EMVI) status and apical LN involvement as well as data from surveillance CT scans, to identify incidence and timing of any local or distant recurrences.Eligibility criteria and selectionInclusion CriteriaThe study population was derived from adult patients who underwent colorectal cancer resections between January 2010 and December 2014 at our hospital.Inclusion criteria were: (i) patients who had no evidence of metastatic disease at the time of surgery (stage M0), (ii) the primary treatment was performed with curative intent, (iii) the post-operative histology confirmed complete surgical resection (R0), (iv) patients who were deemed fit enough to undergo follow-up and engaged in the follow-up protocol either until the completion of annual CT-TAP for three years or until they were diagnosed with a local or distant recurrence.The main outcome measures were\u00a0the primary outcome was the identification of local or distant recurrence.Statistical techniquesData analysis was carried out using IBM SPSS\u00ae . Identification of any correlations between known prognostic risk factors and recurrences was carried out using Kaplan Meir and Chi-square tests and a P-value of <0.05 was considered significant.Due to the inclusion criteria, only 179 patients were included in the study. There were 103 males and 76 females (M:F = 1.35:1) with a median age of 63 (range 30-96) years.The operative procedures performed included 57 total mesorectal excisions, 29 left-sided segmental resections, 78 right-sided segmental resections and 15 multi-segmental resections. The operations were performed by three colorectal surgeons in the department. Out of the 179 procedures, 18 were performed as open approach (10%) and 161 as a laparoscopic approach.Out of the 179 patients, 66 (36.9 %) developed recurrence, of which 7 were local and 59 were distant recurrences.\u00a0Correlations with known poor prognostic indicators confirmed the following findings.T-stageThe incidence of recurrence increased from 4 out of 17 (23.5%) in T1, one of which was a local recurrence, 7 out of 26 (26%) in T2 , 20 out of 82 (24.4%) in T3  and 35 out of 53 (66%) in T4 .\u00a0The incidence of recurrence in T4 tumours was found to be higher than any of the other T stages, achieving statistical significance (P=0.0001).\u00a0The median time to recurrence was\u00a023 months after the operation for patients with T4 disease (CI 95% \u201c17.905-28.95\u201d),\u00a0compared to 36, 42 and 43 months for stages T1, T2 and T3, respectively.\u00a0This earlier recurrence in patients with T4 tumours is statistically significant  in the N0 stage  to 19 out of 45 patients (42.2%) in the N1 stage  and 25 out of 34 patients (73.5%) in N2 stage, 2 of which were local recurrences.\u00a0Patients with stage N2 disease were at a significantly higher risk of recurrence (P = 0.0001) when compared to patients with N0 and N1 disease.\u00a0The median time to the occurrence was\u00a0also significantly earlier (P=0.0001) at 14 months (CI 95% \u201c9.102-18.898\u201d) after the operation for patients with N2 disease, compared to 45 and 33 months for stages N0 and N1, respectively .\u00a0Recurrence occurred in 43 out of these 60 patients (71.7%), compared with 23 out of 119 (19.3%) in the EMVI negative group. This difference was statistically significant (P = 0.00001).\u00a0Recurrence was also seen to occur significantly earlier in the EMVI positive group\u00a0at 17 months\u00a0(CI 95% 7.24-26.76) versus 45 months\u00a0(CI 95% 41.74-48.28) in the EMVI negative group  resected (the closest to the root of mesentery) and mark it with a stitch for the pathologist. This LN is clearly reported in the pathology report as apical LN. Apical LN tumour infiltration was noted in 17 (9.5%) patients.\u00a0Of these, 10 (58.8%) went on to develop recurrence compared with 56 out of the 162\u00a0apical LN negative patients 34.6%). This difference reached statistical significance (P = 0.049). Time to recurrence in the apical LN positive group was also significantly earlier (P = 0.006) at 12 months\u00a0(CI 95% \u201c7.966-16.034\u201d) compared with 39 months (CI 95% \u201c36.166-42.504\u201d)\u00a0in the apical LN negative group .\u00a0T4 tumour stage, N2 disease, apical LN involvement and EMVI positivity were all independent predictive factors, not only of recurrence but also of early recurrence; however, due to the small sample size, a multi-factorial analysis did not yield meaningful results.Our study has demonstrated a high risk of recurrence in T4 tumours, N2 stage, EMVI and apical LN involvement, and these findings are in broad agreement with many other publications .\u00a0Though The follow-up after colorectal surgery (FACS) trial ,\u00a0a randoA large series 8529 cases) published from the USA , showed 529 casesWe believe that the reason why so many trials have not shown any significant advantages of intensive follow-up is that they have included all cases rather than grouping patients by their prognostic features.\u00a0Our study, presented here, has focused on finding a more tailored approach based on the post-operative histopathological features.\u00a0We have been able to demonstrate that the post-operative histological tumour staging, the presence or absence of EMVI and apical LN involvement can be used as guides to be able to categorize patients into low-intensity or high-intensity surveillance cohorts.\u00a0We feel that such a more structured approach for follow-up of patients after curative colorectal cancer surgery is necessary, not only to improve early detection of treatable recurrence but also to maximize effective utilization of resources.\u00a0Further studies are needed to explore this concept.This study has demonstrated that having a more targeted approach with the intensive follow-up being offered to patients who are at higher risk of recurrence.\u00a0Patients with post-operative histopathological adverse prognostic factors including T stage 4, N stage 2, involvement of apical LN and EMVI positive have higher incidence together with early occurrence of metastasis and targeted, more intense approach to this cohort will lead to earlier detection of recurrences while maximizing resource utilization."}
+{"text": "Miscommunication during patient handover can be a major cause of preventable medical errors. Emergency traumas are situations where high stress and cognitive load make communication more difficult. Simulation allows for junior learners to practice emergency scenarios in a low-risk setting. This technical report outlines a simulation\u00a0involving patient handover in emergency trauma scenarios. The intended group of learners are first-year surgery and emergency medicine residents. The scenarios were developed based on the learning objectives of communication, collaboration, and information transfer. Using a high-fidelity simulation mimicking a tertiary care facility, the skills performed in these scenarios can be applied to everyday practice. According to a report by the Joint Commission, an estimated 80% of serious medical errors are related to miscommunication during patient handover . In a suIndividuals of different backgrounds and knowledge bases are involved in the treatment of patients. These individuals must collaborate with one another using their expertise in order to reduce any knowledge gaps. This makes clear and efficient communication vital, as necessary information must be transmitted to the person assuming care. A medical professional that assumes care also has to rely on working memory, as they have the responsibility to gather and retain this information . Tools sThe intended groups of learners are first-year surgery and emergency medicine residents in a tertiary care facility. These groups of learners often face situations where they will receive patients with little to no knowledge about their condition. It is their responsibility to collect any vital information and transfer this information to the next person that will assume the care of the patient.This technical report outlines a novel simulation that involves both patient handover and an emergency trauma situation. Descriptions of simulation exercises involving emergency trauma situations already exist , but theLearning objectives for this simulation are related to developing skillsets involving communication, collaboration, and information transfer and gathering in a scenario related to emergency trauma.CaseRun 1: A 23-year-old male involved in an all-terrain vehicle (ATV) accident just arriving in the trauma bay. The paramedic will be briefed to give handover to the trauma leader (Learner #1). Primarily abdominal injury. Learner #1 will hand over to Learner #2.Run 2: A 30-year-old male fell from a ladder onto a fence. The paramedic will be briefed to give handover to the trauma leader (Learner #1). Primarily abdominal injury. Distractor inserted (Jehovah's Witness Card) during the handover from Learner #1 to Learner #2.Run 3: A 30-year-old male ATV accident without a helmet. The paramedic will be briefed to give handover to the trauma leader (Learner #1). Primarily head injury. Distractor will be patient coning during the handover from Learner #1 to Learner #2.In this technical report, educational context, inputs, processes, and expected products are elements that will support this simulation-based scenario. These elements are organized into a modified context, input, process, and product (CIPP) program development and evaluation model .ContextThe appropriate setting for this simulation is a hospital or a university-based simulation facility. This type of facility most accurately models a tertiary care facility in which residents would receive their training and face an emergency trauma situation. This is a hybrid simulation taking into consideration both technical and communication skills\u00a0.InputEquipmentTables PersonnelTable ProcessPre-BriefingLearners will be provided with an orientation of the simulation lab and the available equipment. A general overview of the simulation will be explained. The formative nature of this simulation and how it will only be used for teaching purposes without a pass/fail system will be explained.The SimulationThe scenario begins with a standardized patient brought into the room with a paramedic confederate briefing Learner #1 on the situation. One nurse confederate will also be at hand in the room to provide any assistance the learner may require. The first stage involves information gathering, with Learner #1 integrating the information obtained from the paramedic confederate, as well as assessing the patient, using the Advanced Trauma Life Support (ATLS) protocol. A handover checklist\u00a0developed based on the acronym SBAR  is present in the room as a cognitive aid, but its use is optional. Upon the collection of necessary information and stabilization of the patient, Learner #2 enters the room. Care of the patient is transferred from Learner #1 to Learner #2. During this transfer, Learner #1 communicates any necessary information regarding the patient to Learner #2. This also gives Learner #2 the opportunity to ask any relevant questions before assuming the care of the patient. After Learner #2 assumes care, a second scenario arises and Learner #2 will need to rely on the information given to them by Learner #1 to treat the patient. Upon completion of the simulation run, the learners will be debriefed. The learners will each complete three different scenarios, with expected improvement after each run. The scenarios of these runs are detailed in Figures DebriefingFeedback will be based upon the debriefing with good judgment principles that involve self-reflection . This wiProducts/outcomesLearning Objective 1: CommunicationHandover between Learner #1 and Learner #2 will be assessed based on adherence to principles of SBAR (see Appendix A) and the prehospital trauma handover checklist developed by Harmsen et al. . PracticLearning Objective 2: CollaborationCollaboration will be assessed using the Communication and Teamwork Skills (CATS) assessment instrument (see Appendix B). The checklist will include aspects like:- How well Learners #1 and #2 collaborated during transfer .- How well learners function within a team setting with nursing confederates.Learning Objective 3: Information GatheringInformation gathering will be assessed by two faculty members observing the simulation and will take into consideration:- How well Learner #1 examined the patient using an ATLS protocol- How well Learner #1 integrates information relayed by the confederate into their care- How well Learner #2 integrates knowledge attained from Learner #1 into their careAs junior learners inevitably face emergency trauma situations during their residency training, having the opportunity to practice in a controlled environment can ensure that they gain proficiency. In particular, emergency trauma handover poses a unique challenge due to the limited time frame, stress, and cognitive load associated with these events.Learners are not only responsible for diagnosing and stabilizing a patient in a timely manner, but they must also be able to retain pertinent information and communicate this knowledge. As a result of these challenges, vital patient information is often omitted during the process of handover. Knowledge gaps during handover is a risk to patient safety and can result in poor health outcomes. However, with repeated practice with simulation scenarios, junior learners can better prepare themselves for real-life situations by utilizing skills they have attained.This simulation teaches these skills based upon the three learning objectives and is supplemented by feedback from experienced faculty members. Furthermore, this simulation closely mimics real-life situations such that these skills are practical. By introducing aspects like distractors during handover that further increase the difficulty of handover, junior learners will be more prepared for random events that can complicate cases in real-life emergency departments.This technical report describes the design of a simulation scenario on patient handover during emergency trauma situations. Simulation provides an opportunity for junior learners to practice key skills related to handover in a controlled setting. These skills are based on the learning objectives of communication, collaboration, and information gathering. As the simulation mimics real-life scenarios, the skills attained can be practiced during encounters in the emergency department."}
+{"text": "Cycas micronesica K.D. Hill as a model that represents the global issues of conservation science and invasion biology. In Guam, several non-native insect invasions began in 2003 and have combined to threaten the island population of this cycad species. In this article, we summarize the history of reported invasions and the reported non-native insect herbivores that have recently increased the threat status. We also discuss the interactions among herbivores that threaten the sustainability of C. micronesica on the island of Guam.Effective conservation of endangered plant species requires identifying their greatest threats to formulate management protocols. Invasive species are a result of global change and are a major threat to biodiversity. We used the island cycad Cycas micronesica by reviewing the history of previously reported invasions and providing an update of recent invasions. Then, we prioritize the threat status of each herbivore and the interactions among them. Plant damage was initiated in 2003\u25002005 by the non-native Aulacaspis yasumatsui Takagi armored scale, Erechthias sp. Meyrick leaf miner, and Luthrodes pandava Horsfield butterfly, which elicited unprecedented irruptions of the native Acalolepta marianarum Aurivillius stem borer and increased herbivory by feral pigs (Sus scrofa L.). The combined impact of these five consumers represents the greatest sustained threat to the cycad tree species. Mitigation of the damage caused by phytophagous non-native species is urgently needed to conserve this unique gymnosperm tree.Invasions of non-native species can threaten native biodiversity, and island ecosystems are ideal for studying these phenomena. In this article, first, we report on the invasive species that combine to threaten the island cycad  The loss of biodiversity through extinction events has occurred since the beginning of life itself. The extinction of some organisms creates a crucial opportunity for the proliferation of other organisms that can exploit the changing environment. Biologists study this phenomenon and are particularly interested in the consequences of a relatively new extinction driver, that is, human activity during what is described as the Anthropocene epoch ,2,3,4,5.Conservation biology has evolved into a well-studied discipline that recognizes that anthropogenic mitigation actions must counteract this extinction crisis . The intThe unprecedented volume of movement of species from their native range into new geographic locations is one of the main extinction drivers in the Anthropocene. Although this would appear to increase biodiversity, the negative impact of the new invasions on native organisms may actually decrease biodiversity ; therefoCycas micronesica provides a model case study that fully integrates conservation science with invasion biology. Improvements in the conservation of biodiversity require the support of adaptive management, both regionally and internationally. This can only be achieved if new knowledge from local research is shared to inform global conservation issues [The recent history of the island cycad known as n issues . Here, wn issues  to beingn issues .Aulacaspis yasumatsui Takagi armored scale in Guam in 2003 [C. micronesica population. The C. micronesica trees exhibited no signs of leaf herbivory prior to this invasion  is the third termite taxon that we have observed consuming C. micronesica stem tissue in habitat. The C. micronesica trees\u2019 damage does not appear direct, as the feeding we observed was restricted to dead vascular tissue. However, the lowest strata of the infested trees\u2019 structural integrity are compromised by the termite activity, which increases the plant population\u2019s threat by reducing the tree\u2019s ability to withstand tropical cyclone force winds.The termite Hermetia illucens L. larvae were observed consuming C. micronesica sarcotesta tissue from seeds heavily infested with A. yasumatsui. This insect\u2019s primary larval food is detritus or feces, including in Guam [A. yasumatsui infestations on the seed integument. in Guam ; therefoRattus rattus L. subspecies diardii were observed on numerous occasions on the island of Rota, feeding on sarcotesta tissue of mature C. micronesica seeds. The seeds were still attached to living, intact megasporophylls in every case.C. micronesica stems was observed being consumed by the hermit crab Coenobita sp. Latreille on numerous occasions. An open wound often results in exposed cortex parenchyma when pigs begin to feed on standing, living C. micronesica stems. This crab is a secondary herbivore of the exposed parenchyma tissue after the initial pig damage.The cortex parenchyma of fallen Satsuma sp. Adams was observed feeding on young expanding C. micronesica leaves. This snail herbivory was a one-time observation.The mollusc C. micronesica herbivores  was released in Guam in 2005 [A. yasumatsui individuals are so small that they can infest locations on Cycas organ surfaces that the relatively large predator cannot access [A. yasumatsui infestations are uncontrolled on leaf, stem, and root surfaces near the ground [A. yasumatsui are less attractive to the predator when the infested leaves are from seedlings rather than adults [A. yasumatsui damage.The scale predator  in 2005 . We havet access ,40. Secoe ground . Third, n adults . These aCoccobius fulvus Compere & Annecke (Hymenoptera: Aphelinidae) and Aphytis lingnanensis Compere (Hymenoptera: Aphelinidae) parasitoids have been conducted in Guam to control A. yasumatsui, and to date we have been unable to verify that these animals have established; however, a fortuitous establishment of the scale parasitoid Arrhenophagus chionaspidis Aurivillius (Hymenoptera: Encyrtidae) was observed in 2013. This parasitoid avoids female armored scales, and preferentially parasitizes male armored scales; therefore, the parasitoid\u2019s effectiveness is limited by this behavior.Numerous introductions and releases of Cycas specialist L. pandava does not pose a lethal threat when it is the only herbivore, but general plant vigor is damaged. This butterfly was in the Northern Mariana Islands of Rota, Saipan, and Tinian for many years before being identified in one northern Guam locality in 2005. The population spread throughout Guam along with A. yasumatsui, with both insects often entering new localities at the same time. The fact that the larvae require young, expanding tissue is both a limitation to the herbivore\u2019s population success and why the herbivory is so damaging to plant vigor. The larvae require leaf tissue for greatest performance, but we have observed larvae feeding on young cataphyll, megasporophyll, and microstrobili tissue when no expanding leaves are available. All Cycas species grow with ephemeral flushes of leaves separated by quiescent periods that may last more than one year. The entire expansion phase of C. micronesica leaves is about 30 d [L. pandava adult finds a leaf in the first week of expansion, the larvae will consume the entire leaf. If a gravid female adult finds a leaf in the second week of expansion, leaflets may be partially or entirely consumed, but rachis and petiole tissue is not consumed. Ovipositions that occur after two weeks may lead to partial leaflet herbivory but not complete leaflet consumption.The out 30 d . If a grC. micronesica and L. pandava co-exist. In a common garden setting, C. micronesica is among the most damaged of Cycas species by this butterfly [Cycas species is one plant trait that determines resistance to L. pandava herbivory [C. micronesica is slow compared to Cycas species that are resistant to this specialist herbivore. Damage to C. micronesica in Guam and Rota is primarily due to the host tree\u2019s lack of resistance [These behavioral phenomena of the host and this butterfly herbivore cause highly heterogeneous spatiotemporal herbivory levels within areas of occupancy where utterfly . The speerbivory . Leaf masistance ,44.Erechthias throughout Guam causes this leaf miner to be the most damaging of the secondary threats. This leaf herbivore cannot kill the host tree when it is the only herbivore present, but general plant vigor is damaged. The initial outbreak was observed in one locality in southern Guam in 2003. To date, this leaf miner has not been observed on Rota. In relatively healthy trees, all of the youngest C. micronesica leaves are undamaged by this leaf miner  was introduced to Guam in the 1700s as a wild hunting game [C. micronesica species recovery, the opposite is true. The deer herbivory is restricted to leaves and megasporophylls and seasons with severe drought, which are rare in Guam. In contrast, the pig herbivory includes leaves, stems, and seeds and is chronic. The recent shift in pig behavior such that standing stems are eaten despite no signs of previous stem borer damage indicates that pig damage has emerged as a new acute threat to the tree species\u2019 survival.The Philippine Deer  for hundreds of years until the invasion of a second non-native organism (A. yasumatsui) caused a change in the tree\u2019s seed dispersal behavior.In the absence of vidually ,69. The C. micronesica population from historical anthropogenic destruction is the steep topography of the coastal areas of occupancy. The relatively recent conversion of habitat to agriculture, military, and urban uses has decimated all native tree species\u2019 populations on most of the northern Guam terrain, which is characterized as a tectonically uplifted plateau. The steep topography of the coastal regions has effectively protected C. micronesica from this historical and ongoing anthropogenic damage. This form of topographic protection of threatened gymnosperms has been identified as a benefit to biodiversity in other geographic regions [One factor that has protected Guam\u2019s  regions . The recC. micronesica, the native interdependencies that have been disrupted by the recent anthropogenic activities include the host tree, the mutualist pollinator Anatrachyntis, the stem borer A. marianarum, the native mycorrhizae species that colonize the tree\u2019s roots, and the native nitrogen-fixing Nostoc species that colonize the tree\u2019s coralloid roots.The influence of how coalitions of organisms influence conservation decisions is not restricted to planning the mitigation of coalitions of non-native threats. Coalitions of native organisms should also guide in defining targeted goals for threatened plant species recovery. Indeed, conservation of native species interactions should be a goal of all restoration plans, not just conserving the species . For C. Cycas micronesica has been listed on the United States Endangered Species Act (ESA) for five years [C. micronesica recovery should embrace the need to conserve all native species that rely on the tree while eradicating or mitigating the non-native biotic threats.ve years . The dirve years . The ultAnatrachyntis pollinators may pose the greatest dilemma for decision-makers. A biological control program for A. yasumatsui may use aggressive exploratory and permitting methods because there are no native armored scale species in the Mariana Islands that may suffer collateral damage. In contrast, a biological control program for the non-native Erechthias leaf miner carries great risk to the native Anatrachyntis pollinator population, as these two genera are closely related. Parasitoids may use cues from leaf-mining activity that are more challenging to find from mining activity [C. micronesica microstrobili production within an area of occupancy to provide brood sites to enable regeneration [Anatrachyntis populations, which can reduce future pollination services. If the pollinator population crashes in some or all of the Guam and Rota areas of occupancy, C. micronesica populations\u2019 potential to passively recover would be minimal. Some of the justification for the in situ conservation management plan in northern Guam [C. micronesica trees within the management plots to sustain regeneration and ensure the pollinator remained viable for future C. micronesica species recovery.Conservation of the activity , so a paneration . Less frern Guam  was to eA. marianarum population in Guam is just as important for conservation as the C. micronesica population; therefore, attempts to implement the biological control of the stem borer to protect the tree population are unjustified. This native beetle poses no threat to the tree population in the absence of stressors such as the list of non-native herbivores in A. marianarum as an acute threat to the tree. Inadvertent conservation of the stem borer in the coming years is ensured because the population of Guam\u2019s unhealthy C. micronesica trees will sustain the beetle\u2019s population until recovery of the plant population ensues.The C. micronesica roots and the Nostoc and mycorrhizae mutualists should embrace a holistic approach. One of the greatest benefits of in situ conservation is that ecological interactions and mutualisms may be conserved even if they have not been studied and are not understood [C. micronesica Nostoc and mycorrhizae mutualists have not been adequately studied. Efforts to remove all non-native plant and animal species from habitats may allow the native root symbionts to restore the native mutualisms passively. For example, the prevalent invasive trees that have altered the Mariana Islands\u2019 landscapes have been shown to change chemistry and mineralization biology of soils compared with adjacent native trees [C. micronesica in northern Guam was exclusion of ungulates with fencing and removal of all non-native plant species [Carica papaya L., Leucaena leucocephala (Lam.) de Wit, and Passiflora suberosa L. were present in high densities. Continued management of these plots must remain vigilant to ensure all non-native organisms remain excluded, enabling the passive conservation of the native edaphic mutualisms disrupted by the non-native ungulates and plants. This process may be slow, and the non-native plants\u2019 sustained exclusion will be required for an extended period [C. micronesica trees in northern Guam is a degraded Vitex parviflora Juss. forest, a conservation approach that has been discouraged because of the altered soil biology in the degraded site that may damage the rescued trees\u2019 health [Conservation of the relationships among derstood . The Guave trees ,77. For  species , as Carid period ,79,80. Td period . Many spd period . The priThese complicated conservation issues indicate that species experts are needed to carefully monitor the interactions among the native species that rely on each other whenever conservation interventions are implemented to mitigate the non-native phytophagous insects that have invaded Guam. A loss of the functioning mutualisms among native species may precede the loss of any single organism . These rC. micronesica conservation.Effective mitigation efforts require the identification of the primary threats by species experts. Regardless of funding and effort costs, preemptive actions will not be successful if conservation decisions fail to address the identified primary threats. Following are recommendations for continued research to support in situ C. micronesica plants on Guam and Rota were not threatened prior to 2003 when the invasions were initiated. This case study illuminates that the best approach for sustaining an unthreatened status of native tree species at risk of herbivory is to prevent the invasions before they occur. The 2020 level of threat and the number of non-native herbivores throughout the indigenous range of C. micronesica occur in the sequence Yap < Palau < Rota < Guam. Why are the C. micronesica populations in Yap and Palau persisting with no threats, but the populations in Guam and Rota suffering from acute threats? Numerous differences among these four geopolitical states may account for the disparity in contemporary threats, including volume of human travel, funding for adequate customs inspections, style of oversight by the federal governments, and conservation ethos of the residents [The esidents ,88. The A. yasumatsui is achievable but will require an integrated multi-year plan for funding the cumulative process. We have collected parasitoids from Cycas leaves infested with A. yasumatsui in the Philippines and Thailand where the armored scale does not threaten its host Cycas trees, only to learn that they have not been described. The binomial is required to obtain an import permit, so these efficacious parasitoids cannot be introduced to Guam or Rota because they are unknown to science. An integrated funded program is needed to collect these parasitoids by cycad biologists, have them described by taxonomists, secure import permits with the new binomial, collect live animals by entomologists, then introduce and release them in Guam and Rota. This was proposed in 2017 as an urgent action for conserving C. micronesica [Successful biological control of ronesica .L. pandava butterfly suffers from the probability of collateral damage to native butterflies. Biological control experts should be employed to determine if a highly specific biological control program is achievable.The development of a biological control program for the C. micronesica experts [C. micronesica stems and roots may improve management decisions and provide knowledge that can be used in other islands where threatened native tree species must contend with frequent tropical cyclones. This abiotic stress will persist in Guam and Rota, and conservation managers must integrate the stress into all decisions.Conservationists are unable to turn off the tropical cyclones that define life on Guam and Rota. The use of guy wire anchors for the trees within in situ conservation plots to protect the trees from catastrophic wind damage was a crucial component of the management plan developed by four international  experts . The val experts . More reA. yasumatsui infestations have become less widespread and less severe in the past two years . Many of Guam\u2019s infestations have taken on the gestalt appearance of A. yasumatsui outbreaks within the native range of the scale, indicating newly efficacious biological control may have become established. The most recent parasitoid survey throughout Guam\u2019s C. micronesica habitats was conducted in late 2017 and early 2018 and revealed A. chionaspidis remained the only parasitoid reared from C. micronesica leaves infested with A. yasumatsui. Studies to determine what appears to be fortuitously controlling A. yasumatsui on Guam in the past two years are urgently needed. We propose three possibilities. First, a new and unidentified parasitoid may have invaded Guam in recent years. Second, either C. fulvus or A. lingnanensis may have established following our many releases of these two parasitoids, but only recently irrupted to a population density such that effective biological control of A. yasumatsui was initiated. Third, the A. chionaspidis population known to be widespread in Guam may have become more efficacious in controlling the scale. Indeed, the combination of A. yasumatsui on C. micronesica in the Mariana Islands\u2019 climate has never been available to this parasitoid, and perhaps these interacting factors combine to improve the parasitoid\u2019s biological control abilities.Guam\u2019s C. micronesica from the ESA should be tailored for Guam and Rota separately, as the approach for species recovery may not be similar for the tree populations on the two islands. Due to lack of funding, we have not been able to observe the C. micronesica status on Rota in recent years. A C. micronesica species expert should be funded to assess Rota\u2019s threat status, as many of the Guam threats were never identified on Rota during the ten years that we were actively studying the C. micronesica population on Rota prior to the loss of funding. A parasitoid survey should be conducted on Rota, and the parasitoids from Guam should be released on Rota if needed. More research on the frequent Rota C. micronesica seed gnawing by R. rattus may be useful for informing global island conservation efforts [The development of efforts to de-list  efforts .C. micronesica by these non-native insects has caused reductions in stem carbon dioxide efflux [The consequences of non-native insect herbivores on a native host tree are not restricted to mortality. We have reported that chronic herbivory of e efflux , has sele efflux , has come efflux , and hase efflux . Additioe efflux ,93. The O. rhinoceros has exhibited a recent host shift from Cocos nucifera L. to C. micronesica throughout Guam [A. yasumatsui and L. pandava likely included a single subspecies from a single geographic origin. Both of these specialist insects have a wide indigenous range comprised of distinct subspecies from different geographic regions. The potential for a second invasion of a second subspecies remains as a potential threat from both of these insect species. If this were to occur, these two specialists\u2019 genetic diversity would increase, which would subsequently increase the level of damage to the host tree. These developments will go unnoticed if species experts continue to be excluded from the conservation actions.The observation that out Guam  illuminaC. micronesica as an excellent model for research designed to exploit the islands as natural experiments to more fully understand the role of fragmentation on biodiversity [Continued research and conservation actions overseen by species experts are needed to fully understand this case study\u2019s relevance to global conservation issues. An updated threat assessment by the IUCN is needed, as the last assessment was ten years ago . A speciiversity . The funC. micronesica. We reported an update on more threats that have not been previously reported to illuminate how single threats may interact to form a coalition of synergistic threats. As conservation decision-makers need multi-threat assessments such as ours to develop the most effective conservation actions [A. yasumatsui, Erechthias, L. pandava, and S. scrofa damage. We have estimated more than 70 years of demographic depth have been lost from Guam due to selective mortality of the smallest individuals [We used a summation of the previously reported threats to the cycad known as  actions , we use  actions . Our rep actions ,14,15,16ividuals . If thes"}
+{"text": "Presented here is a scalable and aqueous phase exfoliation of graphite to high yield and quality of few layer graphene (FLG) using Bovine Serum Albomine (BSA) and wet ball milling. The produced graphene ink is tailored for printable and flexible electronics, having shown promising results in terms of electrical conductivity and temporal stability. Shear force generated by steel balls which resulted in 2\u20133 layer defect-free graphene platelets with an average size of hundreds of nm, and with a concentration of about 5.1 mg/mL characterized by Raman spectroscopy, atomic force microscopy (AFM), transmittance electron microscopy (TEM) and UV-vis spectroscopy. Further, a conductive ink was prepared and printed on flexible substrate (Polyimide) with controlled resolution. Scanning electron microscopy (SEM) and Profilometry revealed the effect of thermal annealing on the prints to concede consistent morphological characteristics. The resulted sheet resistance was measured to be  Printable electronics have received increasing attention due to their broad applications, such as roll-to-roll R2R) printed solar cells , micro eR printedGraphene-based inks have shown promise in fulfilling the aforementioned needs ,12. GrapAt this time, it is evident that the Hummer\u2019s method causes single carbon atom defects and nano-sized holes, due to over-oxidization of the carbon framework . As grapDirect Liquid Phase Exfoliation (LPE) of graphite into graphene has been broadly reported to be desirable for inkjet printing, particularly so for electrophysiology and cell-based studies ,26,27,28ications . Furtherications . Since mications ,26,27,33ications ,35. HoweIn order to overcome this issue, Paton et al. successfully exhibited the usage of an edible protein, bovine serum albumin (BSA), as a stabilizing agent in the LPE process . Kumar eIt has been long known that other mechanical activation techniques, such as ball milling, can be considered as a promising process for modifying carbon nanostructures . To the \u00a0mg\u00a0mL\u22121 , and 0.3\u00a0mg\u00a0mL\u22121 , in DMF.\u00a0mg\u00a0mL\u22121 . Edge-ca\u00a0mg\u00a0mL\u22121 . In consIn the course of printing, a high concentration of graphene in solution serves a threefold purpose. Firstly, the more concentrated the graphene solution, the more viscose the ink, which in practice should reach to approximately 10\u00a0cP . Adverse10\u00a0cP .Herein, we initially propose the combination of BSA as an exfoliating/stabling agent, with the sheer force of continuous low speed wet ball milling for achieving scalable and stable water-dispersed graphene nanosheets with high yield. Next, through utilizing an electrostatic field, the inkjet printing of binder-free graphene solution on flexible substrate is demonstrated. To end with, the resultant conductivity of the printed circuit is characterized, and its stability after submergence in water is exhibited. In another study by our group, the biofunctionalization of the biosensors produced via this process is exhibited .It is shown that rat neuronal cells can be cultured in-vitro on graphene biosensors, and these sensors can enable the sensing of electrical signals on a cell membrane. This study aims to provide a practical guideline for the production of highly concentrated graphene and the patterning of circuits for use in biosensing and other applications in flexible electronics.Graphite crystallites  at \u00a0=\u00a0\u03b1A/L) .Raman spectra of the thin films on alumina membranes were acquired using a BWTEK Voyage confocal Raman system , with a CW laser (Excelsior-532-150-CDRH Spectra-Physics) as the energy source operating at the wavelength of TEM images were recorded using a JEOL JSM2100 STEM  at 200 kV accelerating voltage. Graphene samples were diluted to Z-value  and (4), were satisfied to obtain a printable ink. These equations confirm that the ink is not too viscous to clog the needle or the junctions, and not too diluted to splash while maintaining high flow rates. The physical properties of graphene ink itself are critically influential in controlling the resolution and consistency of the printed patterns. Hence, two nondimensional properties: Reynolds number and Weber number (Equations (1) and (2)) were utilized to govern the printability of the ink . Consequnk drops . Suggestelow 100 .(1)Re\u00a0=Z-value of A high flow rate is essential for achieving superior conductivity outcomes for single printing passes. The measured  on a Cu-grid  images were collected using dried, deposited graphene  . M is a n et al.  (EquatioWe used atomic force microscopy (AFM) to analyze the thickness of the graphene nanosheets after stabilization with BSA. The height-profile images of the graphene sheets revealed that they had a thickness of 7.6 \u00b1 2.3 nm, suggesting the presence of few-layer graphene sheets in aqueous dispersion F. Note tA significant factor in the process of exfoliating graphene from pristine graphite is the scalability of the production method. The bio-graphene presented here preserves its qualities, such as consistent yield, lateral size, conductive characteristics and being devoid of defect. In order to increase the batch volume, the number of balls was chosen based on the ratio of the overall surface area of the balls to the volume of the solvent  using a custom designed printing setup demonstrated schematically in The chosen PI substrate was ubstrate . Due to The resolution of prints is controlled by: the inner diameter of the chosen needle, the positive pressure behind the ink-filled syringe (represented by the flow rate injected by the syringe pump), needle speed along the printed lines and the electric potential difference between the needle\u2019s tip and the substrate  neuronal cells live compatibly and adhere to the graphene patterns  in the b"}
+{"text": "Anaplastic lymphoma kinase (ALK) positive large B-cell lymphoma (ALK+ LBCL) is an extremely uncommon non-Hodgkin lymphoma (NHL) with a distinctive histomorphologic, immunophenotypic and cytogenetic profile. It is unlike the more common ALK-positive anaplastic large cell lymphoma, although the latter shares the ALK rearrangement pathognomonic for this entity. ALK+ LBCL is an underrecognized entity since it is rare and unfamiliar, and shares morphologic and immunohistochemical features with a variety of other neoplasms that can result in misdiagnosis. This lymphoma exhibits plasmacytoid morphology and negativity for classical\u00a0immunomarkers of B- and T-cell lineages, and CD30; however, it expresses terminally differentiated B-cell/plasma cell markers such as CD38, CD138, and MUM-1. Precise identification of this entity is pivotal because of its aggressive behaviour, poor response to standard chemotherapy regimens and the potential for the development of\u00a0novel\u00a0targeted therapy.\u00a0A high index of suspicion on morphology and an extensive immunohistochemistry armoury are required for the veracious\u00a0detection of this lymphoma, especially at extranodal sites. The purpose of bringing forth this present case, an extranodal neoplasm with plasmacytoid morphology at vertebral location in a young adult, is to highlight the diagnostic perils and pitfalls, the clues to unravel the quandaries and thus, the incredible utility of histopathological examination and immunohistochemical analysis in attaining the unerring diagnosis. Anaplastic lymphoma kinase positive large B-cell lymphoma (ALK+ LBCL) is an unusual variant of diffuse large B-cell lymphoma (DLBCL) accompanied by exclusive Anaplastic lymphoma kinase (ALK) rearrangements. A tyrosine kinase receptor belonging to the superfamily of insulin receptors, ALK contributes significantly to the development of the brain, and regulates the proliferation of nerve cells .\u00a0ALK+LBCAn 18-year-old male presented with dull aching abdominal pain for four months and weakness of both lower limbs for three months, accompanied by fever and weight loss. Magnetic resonance imaging (MRI) scan of the thoraco-lumbar spine performed\u00a0for the complaints of abdominal pain and limb weakness revealed a T2/ST1R hyperintense homogenously enhancing mantle of soft tissues involving the dorsolumbar pre- and para-vertebral regions with epidural, subpleural extension and rib destruction, with the epidural component\u00a0at D8-D10 levels causing significant cord compression. Bulky bilateral axillary lymphadenopathy measuring 8.6x6.2x12 cm, bulky prevascular lymphadenopathy measuring 2.9x7.7x9.4 cm, along with multiple nodes in the mediastinum and bilateral supraclavicular region were also discovered. Biopsy of the epidural space-occupying lesion at D8-D10 level had been performed and the preliminary histopathology report from a private diagnostic laboratory was small round cell tumor, probably non-Hodgkin lymphoma (NHL). The patient was then referred to our tertiary cancer centre, and the outside reported tissue paraffin blocks and slides were reviewed. Appraisal of the histopathology slides revealed fibrocollagenous tissue infiltrated by sheets of mostly medium-sized atypical cells separated by fine fibrovascular septa imparting an alveolar pattern. The cells were predominantly plasmacytoid possessing eccentric round to oval nuclei and moderate to abundant cytoplasm. Marked pleomorphism was noted at places, with some cells displaying large irregular hyperchromatic nuclei and scant cytoplasm. Focal necrosis was evident  was broad, encompassing plasmacytoma, small round cell tumors such as rhabdomyosarcoma and Ewing sarcoma, poorly differentiated carcinoma (metastasis), neuroendocrine tumor, melanoma and a host of NHLs, including anaplastic large cell lymphoma (ALCL), DLBCL, plasmablastic lymphoma (PBL) and primary effusion lymphoma (PEL). An elaborate panel of immunohistochemical (IHC) antibodies was executed to attain the diagnosis, which divulged the atypical plasmacytoid cells to be positive for CD138 (diffuse and strong), Mum1 and CD45, and negative for EMA, desmin, CD30, CD20, CD3, CD43, CD56, FLI1, S100, HMB45 and CD79a , and eosinophilic appearance of the cytoplasm. Necrosis, brisk mitosis and a starry-sky appearance may be spotted ,13,14.In view of the morphologic overlap with a variety of neoplasms, immunophenotypic analysis is crucial for the diagnosis. The atypical cells are habitually negative for B cell markers - CD20 and CD79a, but parade unflinching positivity for CD138, Mum1 and CD38, compatible with a postgerminal center phenotype, thus simulating a plasma cell neoplasm. However, the most distinctive IHC profile is the positivity for the ALK protein, staining 100% of tumors, with most demonstrating a restricted granular cytoplasmic staining pattern, while the rest display cytoplasmic, nuclear and nucleolar ALK staining ,15. CD45The characteristic cytogenetic abnormality is the ALK locus translocation on chromosome 2, and the commonest rearrangement is the t2;17), leading to a CLTC-ALK fusion protein. The t2;5) translocation is identified in relatively minor cases, resulting in the NPM-ALK fusion ,9,16. ALA medley of neoplasms enter the differential diagnoses, hematopoietic and non-hematopoietic as well, especially at extranodal sites. ALK+ LBCL occasionally forms nests and can display round cell morphology and thus, mimic poorly differentiated carcinoma, rhabdomyosarcoma, Ewing sarcoma, neuroendocrine tumors and melanoma; most of these can felicitously be distinguished on the basis of IHC findings, and usually do not cause much headache to the pathologists. However, at times IHC can be fallacious: EMA and occasionally focal cytokeratin are expressed in ALK+ LBCL while the customary B-cell and T-cell markers are not embodied. To boot, CD138 is not specific to the hilt; apart from labelling ALK+ LBCL and plasma cell neoplasm, it also stains the tumor cells in most carcinomas. Hence, in such scenarios, additional IHC with different cytokeratins, other lineage markers, Mum1 and ALK protein ought to resolve the conundrum.The more arduous quandaries relate to the hematopoietic neoplasms, the most formidable being ALK+ ALCL and plasmacytoma; B cell NHLs disclosing an immunoblastic/plasmablastic morphology, particularly PBL, PEL, HHV8 positive large B cell lymphoma and DLBCL, NOS round off the list. Clinical presentation, sinusoidal template of involvement, and expression of markers like ALK, CD45, EMA (83% in ALK+ ALCL and 93% in ALK+ LBCL), and CD4 (40-70% in ALK+ ALCL and 40-75% in ALK+ LBCL) ,17 are nThe natural course of ALK+ LBCL is dismal, which is not disparate from other large B-cell lymphomas with plasmablastic differentiation. Promising results have not been reaped with standard lymphoma therapeutics, including CHOP or CHOP-derived chemotherapy with/without radiation and stem cell transplant . This lymphoma being primarily CD20 negative, rituximab is not expected to churn out quantifiable benefits ,13,15,16To conclude, ALK+LBCL is a rare aggressive\u00a0B-cell NHL with discrete morphologic, immunophenotypic and cytogenetic/molecular findings. In a tumor of plasmacytoid/immunoblastic morphology presenting at nodal/extranodal sites exhibiting immunopositivity for ALK, plasma cell markers and sometimes CD4 and EMA and occasionally focal and weak cytokeratin, and negative staining for both B and T cell markers, CD30 and even CD45 at times, the possibility of this neoplasm should be considered. Owing to the extreme rarity of this entity and consequently lack of a high index of suspicion, morphologic overlap with other hematopoietic and non-hematopoietic neoplasms, unusual immunoprofile and seldom regular employment of ALK IHC, the diagnosis\u00a0remains challenging and may be missed. Accurate\u00a0recognition of this entity is of paramount importance, as the promise of a targeted therapy bestows a fascinating recourse for patients with this malady."}
+{"text": "Women with pre-existing morbidity arising from medical conditions or previous caesarean section are at higher risk of adverse pregnancy outcomes compared to women without such morbidity. Women often face complex pregnancy-related decision-making that may be characterized by conflicting maternal and perinatal priorities. The aim of this systematic review and meta-analysis was to assess randomised controlled trials of decision aids to evaluate whether they are effective at reducing decisional conflict scores and to evaluate what type of decision aids are most effective for women with pre-existing morbidity in pregnancy.We searched Medline (via Ovid), Embase (via Ovid), CINAHL (via EBSCO) from the earliest entries until September 2021. We selected randomised controlled trials comparing patient decision aids for women with pre-existing morbidity with usual clinical practice or a control intervention. Study characteristics and Jadad risk of bias was recorded. Meta-analysis by pre-existing morbidity type was performed using Stata 17 and the data was presented with a Forest Plot. Random effects models were used to calculate summary estimates if there was substantial clinical or statistical heterogeneity and post mean DCS scores were described in a sensitivity analysis and presented as a line graph, to improve clinical interpretation of results.. A narrative synthesis of the selected studies evaluated what type of decision aid works and for in what circumstances.Ten randomised controlled trials, which reported data from 4028 women, were included. Patient decision aids were evaluated in women with pre-existing morbidity who were undertaking pregnancy-related decision-making. Patient decision aids reduced decisional conflict scale scores by an additional \u2212\u20093.7, 95% Confidence Interval\u2009\u2212\u20095.9% to \u2212\u20091.6%) compared to the control group. Women with pre-existing medical conditions were more conflicted at baseline and had greater reductions in decisional conflict scale score , in contrast to those with previous caesarean section . There was limited evidence on the effect of decision aids on health outcomes. Decision aids reduced unwanted variation in decision-making support across maternity settings.Patient decision aids are effective tools to support personalised care planning and informed decision-making in women with pre-existing morbidity. Women with pre-existing medical morbidity were more conflicted at baseline and were more likely to benefit from decision aids. Adoption of aids in this population may lead to improve adherence and health outcomes, warranting further research.The online version contains supplementary material available at 10.1186/s12884-022-04402-x. Personalised care is one of the principal objectives of maternity care in the United Kingdom, and internationally. Care centred on the woman, based around her needs and decisions, and where she has genuine choice, informed by unbiased information, should be provided to women with pre-existing morbidities and those without . Women eShared decision-making is a model of care where clinicians and patients share the best available evidence when faced with the task of making decisions; and where individuals are supported to consider options, to achieve informed preferences . In pregA recent review of patient decision aids in a wide range of clinical scenarios across obstetrics and gynaecology has identified that aids are useful in reducing decisional conflict scores (DCS) in women . HoweverThe aim of this systematic review and meta-analysis was to establish in women with pre-existing medical and surgical morbidity affecting pregnancy whether patient decision aids are effective at reducing decisional conflict scores, improving knowledge and health outcomes. The aim of the narrative synthesis was to evaluate what type of decision aids are most effective in what circumstances.http://www.crd.york.ac.uk/PROSPERO/ reference number (CRD42018109005). No ethical approval was required.The study protocol for this systematic review was developed in line with the PRISMA-2020 checklist and registered on the PROSPERO database  and reviewing thesis titles from https://www.worldcat.org/.. References of studies that underwent full text review and relevant review articles were also searched using the snowballing approach. No language restrictions were applied. The study protocol (including the literature search strategy) is detailed in supplementary file\u00a01.A comprehensive search using Medline (via Ovid), Embase (via Ovid), CINAHL (via EBSCO) from Medline 1946, Embase 1947 and CINHAL 1956 until 07/09/2021 was performed. Search strategies were adapted to each database. Searches of exploded title, abstract and keywords \u201cPregnancy\u201d or \u201cPrenatal Diagnosis,\u201d or \u201cPartuition\u201d (Medline and CINAHL) or \u201cBirth\u201d (Embase) were combined with \u201cDecision Support Techniques\u201d (Medline and CINAHL) or \u201cDecisions Support Systems\u201d (Embase) or \u201cDecision Making\u201d. The randomised controlled trial filter was then applied to the search results.\u201d. The Cochrane Trials Register was accessed via Google Chrome and searched using title, abstract and keywords \u201cPregnancy\u201d and \u201cDecision support\u201d. Medline and Embase searches were performed individually and then combined in a single Ovid database which was automatically deduplicated. Integration and deduplication of the combined search with the search results from CINHAL and The Cochrane Trials Register was done by hand in Microsoft Word. No unpublished studies were identified by searching for trials registered on Randomised controlled trials of patient decision aids for women with pre-existing medical or surgical conditions undertaking pregnancy related decision-making was the focus of the systematic review with meta-analysis. Only patient decision aids that were developed with reference to internationally agreed-on criteria that includes information regarding the health condition; the interventions available and the evidence base for them; the possible benefits and harms; probabilities and uncertainties; and the provision of a method for clarifying and communicating the patient\u2019s values were included . Beyond The titles, abstracts and selected full texts generated from the literature search were independently screened by authors R.J.W. and L.M.W. Data from the trials that met all inclusion criteria were manually extracted and entered a standard extraction table independently from full texts by R.J.W. and L.M.W. The authors were not masked to the results of the study or authors. Where two articles published results from the same study, individual pertinent outcomes were extracted from both articles without repetition of data extraction. The viewers independently assessed each trial\u2019s methodologic quality using the Jadad criteria, a standardized tool that assesses quality and risk of bias of randomized trials . The criDecisional conflict was chosen as the primary outcome as it is a patient-oriented indicator of the decision-making process. Decisional conflict is measured using the validated decisional conflict scale (DCS) . The scaResults were grouped and meta-analysed by pre-existing morbidity type  using Stata 17 and the data was presented with a Forest Plot. Egger\u2019s test for publication bias was performed using Stata 17 and analysed by pre-existing morbidity type . Random effects models were used to calculate summary estimates if there was substantial clinical or statistical heterogeneity, as is recommended in the Cochrane Handbook . Where iA narrative synthesis approach  that desTitles and abstracts of 1311 papers were screened, and 60 relevant studies were selected for full manuscript review Fig.\u00a0 14\u201323].\u201323.14\u201323Each trial\u2019s methodologic quality and risk of bias was assessed by using the Jadad criteria  Table\u00a033 and scop\u2009=\u20090.919) . The overall I-squared was 59% (p\u2009=\u20090.008). Moderate heterogeneity [All ten of the studies adopted the O\u2019Connor DCS and were included in the meta-analysis . However19) Fig. . Heterogogeneity  across togeneity , 23. Iniogeneity . FurtherThe ten randomised controlled trials included in the review all reported knowledge as one of their predefined outcomes. Eight out of ten studies used a validated knowledge questionnaire, , 20, 21 Half of the studies included in the systematic review included health outcomes measures, including two pre-existing medical conditions studies both of which addressed anti-depressants use in pregnancy , 18 and Both paper-based decision aids and computerised decision aids were used across the ten randomised controlled trials. Five studies trialled paper-based decision-aids between 2008 and 2019 , 19, 22 The women recruited to the studies included in this systematic review received care in both primary and secondary care settings, from midwives, obstetricians, and physicians. Some women with perinatal mental health conditions, and some women who had had a previous caesarean section birth, received care from a specialist antenatal clinic. Two of the three studies investigating the effect of decision aids on decisional conflict in women who had had a previous caesarean section compared the decision aid to specialist service and found no difference between the two arms , 23. WomThe systematic review has demonstrated that patient decision aids modestly reduce decisional conflict in women with pre-existing morbidity making pregnancy related decisions (2\u20137%). Women were also more knowledgeable about their condition following use of a decision aid compared to usual care that includes information brochures. Although reductions in decisional conflict were modest, on average they helped women move towards a less conflicted state, which coupled with increases in knowledge scores suggests women were more likely to make informed pregnancy related decisions. Decision aids were most beneficial when provided to women with pre-existing medical conditions who were making decisions about the safety of medication and pregnancy, as these women were generally more conflicted at baseline and had greatest reductions in mean difference. There is evidence to suggest that paper-based aids are as effective as computerised aids, but there is only limited evidence on electronic personalised decision aids. Decisions aids are likely to reduce variation amongst women making pregnancy related decisions regardless of their model of care, by standardising decision-making support across different healthcare settings.The strength of this study includes its comprehensive search strategy and its focus on women with pre-existing morbidity who face particularly complex pregnancy decisions. Its systematic review with meta-analysis and sub-analysis of change in pre and post DCS score enables readers to understand not just the overall effectiveness of decision aids but also the clinical importance of these results. The narrative synthesis approach also enables readers to understand which type of decision aid works and in what circumstances do they work, leading to a more nuanced approach to pregnancy decision aid implementation in women with pre-existing morbidity.The findings of this review are limited by the inherent risk of bias across all the studies. The Jadad risk of bias assessment  found th25 Policy makers should work towards ensuring women with common medical conditions such as hypertension and diabetes have access to contemporary pregnancy decision aids to support personalised care and support planning.Implementing personalised care, centred on the woman, her baby and her family, based around their needs and their decisions, where they have genuine choice, informed by unbiased information is a high priority for national and international policy makers . DespiteDespite benefits associated with implementation, mean post DCS score remained high for some women in many of the studies. This is perhaps unsurprising, as all of the decision aids included in this review were delivered outside of the consultation and were not designed to facilitate personalised in-consultation care planning . Low-cosIn addition to improving personalised care there is a national priority to improve the safety of pregnancy and birth. Recurrent reviews into maternal and perinatal mortality have identified that women with pre-existing morbidity are at greater risk of poor pregnancy outcomes . There aIn conclusion, patient decision aids support personalised care planning and informed decision-making in women with pre-existing morbidity, although the effect may be modest. Women with pre-existing medical morbidity are likely to benefit from decision aids because of high levels of existing conflict and the effectiveness of decision aids in this group. Further, adoption of aids in this population may lead to improve adherence and health outcomes. It is likely that the adoption of a co-design approach to the development of decision aids would result in more effective tools, as women themselves are best placed to identify their decisional needs. Future research should also consider whether in-consultation aids better support personalised care and informed decision-making.Additional file 1.Additional file 2.Additional file 3.Additional file 4.Additional file 5."}
+{"text": "To explain the concept behind the Castrop toric lens (tIOL) power calculation formula and demonstrate its application in clinical examples.The Castrop vergence formula is based on a pseudophakic model eye with four refractive surfaces and three formula constants. All four surfaces  are expressed as spherocylindrical vergences. With tomographic data for the corneal front and back surface, these data are considered to define the thick lens model for the cornea exactly. With front surface data only, the back surface is defined from the front surface and a fixed ratio of radii and corneal thickness as preset. Spectacle correction can be predicted with an inverse calculation.Three clinical examples are presented to show the applicability of this calculation concept. In the 1st example, we derived the tIOL power for a spherocylindrical target refraction and corneal tomography data of corneal front and back surface. In the 2nd example, we calculated the tIOL power with keratometric data from corneal front surface measurements, and considered a surgically induced astigmatism and a correction for the corneal back surface astigmatism. In the 3rd example, we predicted the spherocylindrical power of spectacle refraction after implantation of any toric lens with an inverse calculation.The Castrop formula for toric lenses is a generalization of the Castrop formula based on spherocylindrical vergences. The application in clinical studies is needed to prove the potential of this new concept. The first intraocular lens (IOL) power calculation formulae were published in 1967 by Fyodorov . Since tWith most of the classical calculation concepts, toric intraocular lenses (tIOL) can be easily calculated in 2 separate steps for the two cardinal meridians of the cornea: in the first, we calculate the lens power for the flat corneal meridian, and in the second, the respective lens power is calculated for the steep corneal meridian , 23. ThiAs an alternative, if we restrict the model to linear Gaussian optics, we could use spherocylindrical vergences instead of spherical vergences, or equivalently we could use 4 \u00d7 4 matrix algebra instead of 2 \u00d7 2 matrices to address spherocylindrical surfaces with random axes , 32. SucIn principle, such a vergence  or matriThe purpose of this study was to show how the Castrop formula as a vergence-based calculation concept with paraxial simplifications with a \u201cthick lens\u201d cornea and a \u201cthin lens toric intraocular lens\u201d could be generalized for prediction of the power of toric lens implants as well as the prediction of postoperative refraction. In this generalization, all surfaces should be considered spherocylindrical with astigmatic axes at random orientations.cor using a linear regression as described by Cooke et al.  for calculation of IOL power,ViIOL for prediction of refraction after IOL implantation,VCA 1, ACA1, PCA2, ACA1+90\u00b0, PCA2\u2212PCA1, 0.5\u00b7(PCA1+PCA2), (PCA2\u2212PCA1) cos(2\u00b7ACA1), (PCA2\u2212PCA1) sin(2\u00b7ACA1)],,,VCPA , andVR.For the calculation of the power of a toric lens implant, we calculate the vergence in front of (V3) and behind (V3\u2032) the tIOL. According to Eq.  the follV0 for objects at infinityV0\u2032Add_Surface V1Transform_Vergence V1\u2032 Add_Surface V2C)Transform_Vergence V3 A)Transform_Vergence /nV3\u2032 V/(ALcor-ELP), 0\u00b0, nV/(ALcor-ELP), 90\u00b0, 0, nV/(ALcor-ELP), 0, 0] for objects at infinityThe calculation concept for the prediction of spherocylindrical spectacle refraction after implantation of a toric lens is displayed in Fig. The predicted refraction at the spectacle plane (PREF) is extracted from V0, VR, and V0\u2032:The applicability of this step-by-step approach is shown with 3 calculation examples: the first to calculate the power of a toric lens with curvature data from the corneal front and back surface, the second with curvature data from the corneal front surface and consideration of SIA and CPA, and the third to predict the refractive outcome at the spectacle plane in the situation of example 2 with a preset toric lens.This article does not contain any studies with human participants or animals performed by any of the authors.1 = 7.9 mm/ACA1 = 10\u00b0 and RCA2 = 7.6 mm) and back surface curvature (RCP1 = 6.8 mm/ACP1 = 20\u00b0 and RCP2 = 6.6 mm) and the corneal thickness (CCT = 550 \u03bcm) are available from tomographic measurement. Axial length, anterior chamber depth, and lens thickness are measured as AL = 23.7 mm, ACD = 3.5 mm, and LT = 4.1 mm, respectively. With a toric lens implant, we wish to end up with a refraction at the spectacle plane (VD = 12 mm) with TRS = \u22120.1 dpt, TRC = \u22120.1 dpt at TRA = 90\u00b0. As formula constants, we used C = 0.424, H = \u22120.312 mm, and R = 0.077 dpt. Surgical-induced astigmatism was disregarded (SIAC = 0) in this example, and as we measured both corneal surfaces no correction for the corneal back surface astigmatism was necessary (CPAC = 0).For this 1st example, we have assumed that the corneal front  according to Eq.  readVTRTable 1 = 7.9 mm/ACA1 = 10\u00b0 and RCA2 = 7.6 mm, and from the fixed ratio of corneal front to back surface curvature of 7.77/6.4, we obtain RCP1 = 6.51 mm/ACP1 = 10\u00b0 and RCP2 = 6.26 mm and CCT = 500 \u03bcm. For axial length, anterior chamber depth, lens thickness, and target refraction at spectacle plane, we have used the same values as in example 1. Again, the following values for formula constants were used: C = 0.424, H = \u22120.312 mm, and R = 0.077 dpt. In this example, the surgically induced astigmatism was set to SIAC = 0.20 dpt at incision axis 95\u00b0, and in the absence of tomographic measurements for the corneal back surface, a correction for corneal back surface astigmatism with CPAC = 0.27 at 90\u00b0 has been used.For this 2nd example, we have assumed that the corneal front surface is measured and that the corneal back surface and central corneal thickness data are not available. Again, RCAVCP VSIA VCPA For the corneal back surface, the SIA, and the CPA correction, the respective vergences (without units) according to Eq.  readVCPTable For this 3rd example, we predict the spectacle refraction after implantation of a tIOL. The data for corneal front surface curvature, SIA, CPA, AL, ACD, and LT from example 2 are used. Again, for corneal back surface, we have assumed a fixed front to back surface curvature ratio of 7.77/6.4, and for corneal thickness we use CCT = 500 \u03bcm. For the toric lens, we have assumed a spherical equivalent power of the tIOL with iIOLEQ = 21 dpt and an absolute cylinder power of iIOLC = 1.5 dpt implanted in an axis iIOLA = 105\u00b0 (iIOLS = 20.25 dpt).For the implanted lens, the respective vergence (without units) according to Eq.  readsViIOL = Table Today, there are several options available for correcting corneal astigmatism. In addition to classical spectacle correction and correction with soft or rigid contact lenses, there are surgical options such as corneo-refractive surgery in terms of PRK or LASIK, transverse or arcuate shaped corneal incisions using either a guided diamond knife or a femtosecond laser, toric phakic or add-on lenses for the phakic or pseudophakic eye, or toric capsular bag lenses implanted during normal cataract surgery. Spectacle correction with cylindrical lenses has the disadvantage of image distortion in terms of different magnifications in the principal meridians. Furthermore, if the spectacle correction is not perfectly aligned (decentration or tilt) or for peripheral vision, there are additional distortion effects such as coma, which may deteriorate the retinal image performance significantly . In geneTherefore, surgical procedures\u2014if properly aligned\u2014are beneficial for a stable correction of corneal astigmatism. Refractive procedures at the cornea are typically restricted to low or moderate cylinder values; e.g., corneal incisions allow correction of corneal astigmatism of up to 2.5 or 3 dpt. Excimer laser correction (PRK or LASIK) has good predictability and long-term stability of the refractive outcome for low and moderate cylinder values only. For larger corneal astigmatism values of 4 dpt up to 12 dpt and more, toric capsular bag or additional lenses yield a predictable option for refractive correction with a high patient satisfaction.There are several options for calculation of toric lenses in clinical routine: more or less all manufacturers of toric lenses have developed online calculation features. Some of the online calculators require the biometric data for the eye and provide the spherical power (rarely) or spherical equivalent power (according to EN ISO 11979:2008) of the toric lens together with the absolute cylinder power and the implantation axis. Others require the spherical equivalent power of the lens and the corneal astigmatism as input data and derive the torus of the lens implant. In addition, some manufacturers offer a calculation service where the biometric data are entered in a calculation form and professional optometrists or opticians calculate the toric lenses individually. Furthermore, there are some manufacturer-independent calculation platforms such as the Barrett toric calculator where the biometric data are entered by the surgeon and various options for toric lenses  are derived.Calculating toric lenses with classical concepts such as standard formulae has strict limitations. If a toric lens calculation is split into 2 separate calculations of \u201cspherical\u201d lenses, we must take care that the estimated lens position does not depend on the corneal curvature. Otherwise, we obtain different effective lens positions, one for the flat and one for the steep meridian of the cornea, which does not make sense! This is at least true for the SRKT, the Hoffer-Q, and the Holladay1 formula \u20137, 9, 10If a sufficient number of tomographical data points are available, including either height or curvature data from the corneal front and back surface, the design data of the toric lens implant , the pupil outline, and the positions of the cornea and toric lens relative to the visual axis, then full aperture ray tracing strategies are a powerful tool for toric lens calculation \u201322. Ray Another task in toric lens power calculation is the surgical-induced astigmatism. More or less all calculation concepts offer the option to deal with an equivalent-neutral consideration of an astigmatic vector at the corneal front surface plane, which should consider the systematic vector change of corneal power due to cataract incision , 27. ThiAnother issue is the corneal back surface astigmatism. In the last decade, several papers have been published on corneal back surface astigmatism , 28\u201332. The present paper is based on the Castrop calculation strategy for spherical lenses. The Castrop formula is based on 3 formula constants. According to the Olsen formula, the C constant , 18 reflThe calculation concept as shown here is not restricted to any particular number of spherocylindrical surfaces. All surfaces and vector corrections are considered with cylinder axes oriented at random. Therefore, for calculation of a toric lens, the target refraction, the corneal front surface, and CPA or cornea front and back surface as well as SIA could act as crossed cylinders. The resulting toric lens power compensates all these cylinder values to obtain a plano vergence for V0 (for objects at infinity) or a spherical vergence V0\u22600 (for objects at finite distances). For reverse calculation with any toric lens implanted in any orientation, this concept yields the predicted spherocylindrical refraction at the spectacle plane. The basic principle of this calculation concept has already been shown in previous papers.The Castrop toric lens power calculation strategy is restricted to linear Gaussian optics . This simplification allows a straightforward calculation of the power of a toric lens or a prediction of spectacle refraction after implantation of any toric lens. If\u2014instead of data such as sphere or cylinder or cylinder axis\u2014the surface data of ALL refractive surfaces are available, a full aperture ray tracing strategy could be set up, which may yield more precise results for larger pupil sizes or for peripheral vision. However, such a full aperture ray tracing scheme is much more complex than any vergence- or matrix-based calculation concept. Another limitation of this study is that we restricted the model to a centered optical system with all surfaces aligned to the optical axis. Effects of decentration or tilt of refractive surfaces cannot be addressed with this calculation concept.In conclusion, the Castrop formula for calculation of toric intraocular lenses and prediction of spectacle refraction of any toric lens is based on the Castrop formula for spherical lenses with 4 refracting surfaces. It uses spherocylindrical vergences throughout and can consider an arbitrary number of spherocylindrical surfaces with random cylinder axes. The effective lens position is derived from the phakic anterior chamber depth and lens thickness and 2 formula constants C and H, and a 3rd formula constant R is used to adjust the refraction at the spectacle plane. If tomographic data of both corneal surfaces are available, the Castrop formula considers the curvatures of both surfaces and corneal thickness. If corneal back surface data are unavailable, corneal thickness is preset and the back surface curvature is derived from front surface curvature using a fixed front to back surface curvature ratio, and a vector correction for back surface astigmatism could be included. Surgically induced astigmatism is considered a vector at the corneal front apex plane."}
+{"text": "The rheological characterisation of liquids finds application in several fields ranging from industrial production to the medical practice. Conventional rheometers are the gold standard for the rheological characterisation; however, they are affected by several limitations, including high costs, large volumes required and difficult integration to other systems. By contrast, microfluidic devices emerged as inexpensive platforms, requiring a little sample to operate and fashioning a very easy integration into other systems. Such advantages have prompted the development of microfluidic devices to measure rheological properties such as viscosity and longest relaxation time, using a finger-prick of volumes. This review highlights some of the microfluidic platforms introduced so far, describing their advantages and limitations, while also offering some prospective for future works. Rheology is defined as the science of deformation and flow , and it There are several types of rheometers, depending on both the imposed flow and the working principle ,2. For iThe advent of microfluidics has made a significant impact across several applications spanning different fields . MicroflThe goal of this review is to provide an overview of the microfluidic rheometers developed over the years, together with their advantages and limitations. In fact, several microfluidic devices have been introduced to measure rheological properties such as shear-viscosity, longest relaxation time, first normal stress difference and extensional viscosity, for several dilute polymer solutions and for several biofluids, proving their advantages over conventional rheometers. This review does not describe microfluidic devices where complex fluids have been used for a diverse set of applications but rather microfluidic devices for the measurement of the rheological properties of complex fluids. The review is structured as it follows: Before reviewing the advancements in microfluidic devices for rheological applications, some essential concepts of rheology, which may not be familiar to the reader, are briefly discussed. The interested reader can look at the detailed rheology book edited by Macosko  or the oA, having a gap H, such that x-direction. Because of the no-slip conditions on both the stationary and the moving plate, adjacent layers of fluids move with different velocities, leading to the so-called shear flow . At llongest relaxation timex, and y. Assuming that the fluid is subjected to small deformations, it is possible to write . . 110]. CMicrofluidic devices were also integrated with SAXS apparatus, similarly to the SANS, to study the flow behaviour of several complex liquids c. The fiAn overview of the microfluidic devices reviewed here together with their working principles is reported in s review  allowed z et al.  and Kosez et al. , have nollenging . One appllenging ,120,121.While microfluidic rheometers in shear flow still require improvements, extensional microrheometers have been widely used for the analysis of single molecules, as well as for the study of bioparticles. In this case as well, Rheosense commercialised the e-VROC, which is the extensional version of the m-VROC for shear flow. The rheological community has been very engaged in developing extensional microfluidic rheometers, thus leading to a portfolio of devices whose results were strongly linked to the well-established polymer physics framework . SimilarFuture efforts should also look into machine learning and artificial intelligence, which has recently been the focus within the microfluidic community at large . Integra"}
+{"text": "Recombinant human growth hormone (rhGH) therapy is an effective treatment for children with growth disorders. However, poor outcomes are often associated with suboptimal adherence to treatment.The easypod connected injection device records and transmits injection settings and dose data from patients receiving rhGH. In this study, we evaluated adherence to rhGH treatment, and associated growth outcomes, in Latin American patients.Adherence and growth data from patients aged 2-18 years from 12 Latin American countries were analyzed. Adherence data were available for 6207 patients with 2,449,879 injections, and growth data were available for 497 patients with 2232 measurements. Adherence was categorized, based on milligrams of rhGH injected versus milligrams of rhGH prescribed, as high (\u226585%), intermediate (>56%-<85%), or low (\u226456%). Transmission frequency was categorized as high (\u22651 per 3 months) or low (<1 per 3 months). Chi-square tests were applied to study the effect of pubertal status at treatment start and sex on high adherence, and to test differences in frequency transmission between the three adherence levels. Multilevel linear regression techniques were applied to study the effect of adherence on observed change in height standard deviation score (\u2206HSDS).P<.001). Adherence level had a significant effect on \u2206HSDS (P=.006). Mean catch-up growth between 0-24 months was +0.65 SD overall (+0.52 SD in patients with low/intermediate monthly adherence and +0.69 SD in patients with high monthly adherence). This difference translated into 1.1 cm greater catch-up growth with high adherence.Overall, 68% (4213/6207), 25% (n=1574), and 7% (n=420) of patients had high, intermediate, and low adherence, respectively. Pubertal status at treatment start and sex did not have a significant effect on high adherence. Significant differences were found in the proportion of patients with high transmission frequency between high , intermediate , and low  adherence groups (The data extracted from the easypod Connect ecosystem showed high adherence to rhGH treatment in Latin American patients, with positive growth outcomes, indicating the importance of connected device solutions for rhGH treatment in patients with growth disorders. Adherence to long-term pharmacological treatments, such as growth hormone (GH) therapy for growth disorders, is an area with great potential for improvement ,2. Poor Adherence to GH treatment in the real-world setting has always been difficult to monitor, given the use of unreliable proxy methods such as patient testimony or records of prescriptions filled/vials counted ,5. FurthAutomatic transmission of injection data provides a more accurate insight into real-world adherence patterns and enables HCPs to potentially eliminate poor adherence as a reason for a suboptimal response to GH treatment ,9,10. ThAs different health care systems and variations in clinical practice around the world may affect adherence locally, the deployment of a connected injection device for the treatment of growth disorders across different countries allows the study of behavioral adherence patterns across populations and longitudinally for thousands of patients. This substantial compendium of patient-generated data has also been applied in the context of understanding patterns in diabetes thanks to glucose monitoring technologies ,13. In tGlobal analysis of real-world data obtained from connected injection devices for GH treatment has shown that children with high adherence were most likely to regularly transmit data, and that prepubertal children showed higher adherence than older children and adolescents . This anThe Latin American region is one of the fastest growing regions in terms of the adoption of digital health ,19. ThisIn this analysis, we included children with growth disorders from 12 Latin American countries  and assessed the effects of pubertal status at treatment start, sex, and engagement with treatment on their adherence. An analysis of longitudinal records for a total of 13,553 children in the global database was recently published , but herThe easypod electromechanical injection device, in combination with the easypod Connect web-based platform, as part of an ADSS , electroThis was an exploratory descriptive analysis study during which 4 years of adherence data were analyzed from 6207 pediatric patients with 2,449,879 prescribed injections of rhGH delivered via easypod to treat growth disorders, and who were transmitting data to the easypod Connect system between January 2007 and December 2020. These patients resided in 12 Latin American countries . To avoiEligible patients from each of the participating countries had been enrolled in the database and attended at least one visit, according to local routine clinical practice. Diagnoses and decisions on treatment were made at the discretion of the physician responsible for each patient, following standard endocrinologic practice for each of the participating countries. Prior to enrollment, patients/caregivers reviewed and voluntarily signed an informed consent form materializing their agreement for data collection, storage, and use of their child\u2019s pseudonymized data to create aggregated statistical and general adherence reports.For height, we selected patients with at least two measurements. Adherence  was categorized as high (\u226585%), intermediate (>56%-<85%), or low (\u226456%) for patients on either 6 or 7 injections per week, the two possible regimens for treatment with rhGH. The dosage and frequency of rhGH therapy as per easypod settings were defined by HCPs and data transmissions were initiated by the child, parent/caregiver, or HCP. Adherence was assessed overall and monthly, and explored by puberty status at treatment start , sex, and transmission frequency  in the selected patients with available adherence data for \u22653 months between January 2016 and December 2020. Transmission frequency was calculated in each adherence category as a proxy measure of the patient\u2019s engagement in disease management using easypod. No imputation was made for missing data or withdrawal from the study.Height data were available for 497 patients with 2232 measurements; this included 355 patients from Argentina, 64 from Brazil, 70 from Guatemala, and 8 from Mexico. Height standard deviation scores (HSDS) were calculated using the World Health Organization references [Descriptive statistics were used to describe differences over time in adherence , puberty status , and sex. Chi-square tests were applied to test differences in high adherence between girls and boys, and between prepubertal and pubertal girls and boys. In addition, a Chi-square test was applied to test differences in high frequency transmission between the high, intermediate, and low adherence groups. Multilevel linear regression techniques were applied to study the effect of adherence level on \u2206HSDS between all observed growth measurements, adjusted for the time intervals between them.Treatment with rhGH via easypod was conducted according to local practice. This real-world, retrospective analysis of the data set was performed in accordance with the informed consent form, signed by caregivers of children and adult patients materializing their agreement for data collection, storage, and use of their pseudonymized data to create aggregated statistical and general adherence reports.Any requests for data by qualified scientific and medical researchers for legitimate research purposes will be subject to the healthcare business of Merck KGaA\u2019s Data Sharing Policy. All requests should be submitted in writing to the healthcare business of Merck KGaA\u2019s data sharing portal . When thComplete data were available for 6207 patients, where \u201coverall\u201d is defined here (and throughout) as the total number of patients who received rhGH, started treatment at age 2-18 years, and for whom data were available. Transmission data were available for 5086 patients. Patient demographics according to adherence rates are presented for this data set in Overall, 68% (4213/6207) of patients were in the high adherence category, 25% (n=1574) were in the intermediate adherence category, and 7% (n=420) were in the low adherence category. Furthermore, at each time point, there was a higher proportion of patients in the high adherence category than in the intermediate and low categories combined . High adP<.05) higher in prepubertal boys compared with pubertal boys.There were no significant differences in high adherence between boys  and girls  or between prepubertal and pubertal patients . P<.001).In addition, there were significant differences in the proportion of patients with high transmission frequency between the adherence groups; 59% (2018/3404), 46% (608/1331), and 35% (123/351) in the high, intermediate, and low adherence groups, respectively  in the Dominican Republic to 82% (943/1144) in Brazil .P=.006). Mean catch-up growth between 0-12 months was +0.39 SD overall, with +0.27 SD in patients with low/intermediate monthly adherence and +0.42 SD in patients with high monthly adherence. Mean catch-up growth between 0-24 months was +0.65 SD overall, with +0.52 SD in patients with low/intermediate monthly adherence and +0.69 SD in patients with high monthly adherence. Mean catch-up growth within the subgroup of patients (n=40) aged <8 years and HSDS <\u20132 at start was +1.03 SD. It was not possible to stratify this subgroup of patients by low and high adherence due to the small sample size.The data extracted from the easypod Connect ecosystem showed that children receiving rhGH therapy demonstrated high adherence over 4 years in a real-world setting in 12 Latin American countries.Use of easypod Connect, which showed high adherence in patients with high transmission rates, could be regarded as a proxy of usability and utility of the system by patients and HCPs. In such large-scale deployments of digital health systems, usability cannot be easily measured using questionnaires as this would pose a major user/participant burden. Instead, transmissions and data usage can be used toward identifying patterns of usage of the system and adherence to treatment. Indeed, in this regard, a framework has been proposed to guide future implementation and research on the use of digital health tools to support patients with growth disorders who require GH therapy . These iWe observed that patients who transmitted their data tended to have higher adherence; therefore, a potential hypothesis to further explore is the impact of patients knowing that their adherence data is being observed and is useful to their HCP. Studies in other therapeutic areas have found that factors related to the Theory of Planned Behavior can predict ~50% variance in adherence . Future The selected data from this regional study are broadly consistent with recently published data from the global easypod Connect database, which demonstrated that, of 13,553 children, 71% were in the high adherence category , and witIn both the preliminary Latin American study and other analyses, the proportion of children with high adherence declined over time, and factors such as sex and nominal age at puberty appeared to have only a small effect on adherence rates ,17. SimiPatient attrition over time may be due to a number of factors. Patients may have been switched to a different rhGH, may have continued taking somatropin rhGH using a pen injector, may have continued taking rhGH using easypod but without performing any further data transmission, or may have stopped treatment completely. Minimizing patient attrition is an important consideration in long-term clinical trials, and determining the predictors of attrition is key to identifying patients at risk of missed visits or dropout; such patients may be excluded from a trial or efforts may need to be made to prevent their subsequent dropout once enrolled . This isIndeed, previously reported individual cases of patients receiving rhGH have indicated that direct access to adherence monitoring by HCPs followed by intervention can make a difference to a patient\u2019s management and motivation -32. The Optimizing adherence might also be achieved through the use of structured and active interventions from HCPs, patient/caregiver support programs , and/or The strengths of our study include the large data set from a real-world study conducted across 12 countries in one geographic region with substantially diverse and dynamic health care systems, and the fact that the data are derived from a connected injection device, which offers more reliable data compared to data based on the declarations of patients or their parents/caregivers. Comparable insights into patient adherence from other large-scale patient registries for rhGH treatment are not available due to the lack of alternative electronic devices similar to easypod.Further work is required to assess whether patient/caregiver engagement, as measured by the rate and frequency of data transmission with easypod, is associated with better long-term adherence and clinical outcomes for patients. It would also be interesting to investigate whether or not there is a correlation between the frequency of dose adjustments and the adherence/transmission rate.Limitations of the study include patient attrition, summarized above, and the fact that the change in adherence rate over time varied from child to child, perhaps due to changes in individual treatment plans or other actions taken by the HCP or child/family. Furthermore, not all data were available for all patients over the same treatment duration, as would be expected in any observational study. Differences between the 12 participating countries in terms of socioeconomic factors, such as reimbursement and/or out-of-pocket expenses for GH therapy or free access to medication, may also have affected individual or local adherence rates. Similarly, differences in prescribing habits , patients\u2019 visits to the clinic where growth response is checked and dose might be adjusted to maximize response to treatment, and refill of prescriptions from country to country may also affect adherence rates. Other potential limitations include the lack of additional information on diagnosis and clinical background, limited data on growth outcomes that do not allow assessment of the full catch-up growth pattern, and lack of patient-reported data such as reasons for discontinuation or interruption of treatment, or predefined actions taken by PSPs  in the vP<.001 for patients overall) and height velocity (P<.001) [Finally, usage of the system  can be a proxy of usability and utility; however, usage has been to a large extent influenced by the way the system was introduced. All of these areas require more research. Our analysis showed less catch-up growth (\u20130.17 SD over 24 months) in patients who continue to have low/intermediate adherence compared with patients with high adherence, and this difference increased over time. This difference could be translated into centimeters, resulting in 1.1 cm less catch-up growth between 0-24 months for a patient with an average age for the group 10 years). This is in agreement with the literature, which shows that greater adherence leads to improved growth, with poor adherence adversely affecting growth outcomes ,10,40. I years. T(P<.001) .In terms of future research opportunities, analysis of the potential differences between countries with preset additional data collection points would be of interest. The value of this data can be enhanced by complementing it with self-reported height data entered via a patient app. Future research may also allow for automatic self-measurement of height using novel augmented reality technology on mobile phones. Finally, integration with EHRs may facilitate clinical workflows, as has been demonstrated in other therapy areas .Analysis of the data extracted from easypod Connect showed high adherence to rhGH treatment in Latin American patients, with positive growth outcomes. Thus, our study indicates the potential value of using a connected injection device to monitor and study adherence at an international level. It shows that through our validated method of recording adherence with easypod, we can address an unmet need in rhGH therapy, enabling HCPs to accurately identify patients for whom interventions to improve adherence would be beneficial to improve their growth and other clinical outcomes, particularly as the proportion of children with high adherence declined over time, which is consistent with previous findings. The data also show that children who were most adherent to treatment were more likely to transmit their injection data results regularly and have larger catch-up growth than those who were less adherent. This association between adherence and transmission of data may indicate that sharing data with HCPs has a positive impact on adherence rates, and further studies to confirm this are needed. High adherence and transmission rates may reflect the positive use of the system and could be regarded as indirect indicators of usability and utility of the system by patients and HCPs."}
+{"text": "The indiscriminate consumption patterns worldwide have brought in its wake severe problems like pollution and global warming, and this has ultimately called for green products awareness and consumption. The main purpose of this study was to assess the effect of university students\u2019 awareness of green products on their green purchasing intentions. The specific objectives were to identify whether awareness, price, availability, value and quality influence university students\u2019 intention to purchase green products, and to investigate how awareness, price, availability, value and quality predict university students\u2019 intention to purchase green products. A structural equation modeling was used to analyze data collected from an online survey of 478 students. Results show that green perceived quality has the utmost significant positive impact on university students\u2019 green purchase intentions; however, green perceived availability had the slightest impact on university students\u2019 intention to purchase green products. The study is the foremost to conclude that green product awareness impact on university students green purchase intentions is greatly driven by price, high value and extraordinary quality. However, availability is not a critical influencing factor when it comes to green purchase intentions of university students. The implications of study, limitations and further research are discussed. The unsustainable patterns of consumption in the world today have caused severe environmental problems such as pollution, natural resources depletion, growing greenhouse gas emissions and global warming , 66, 69.s stipulates \u201cresponsible consumption and production patterns\u201d by 2030 [.Green products have become the most reliable resolution for environmental sustainability in many developed countries . The val by 2030 .Similarly, there is a remarkable rise in environmental awareness. This surge in environmental awareness has equally increased the pressure on consumers to consider the environmental consequences of their activities in the last few decades, and this is due to increase in media attention, upsurge in pressure group events, greater consciousness of environmental harms and the growth in individuals\u2019 awareness to green products , 52, 54.Ghana, an emerging market in sub-Saharan Africa, over the past years has not considered many proposals intended to minimize environmental impact in the process of reducing waste, provision of access to green, quality life and maximizing efficiency. As a result, it is vital for Ghanaian consumers to alter their consumption and production forms, for if they fail to change their actions, there will be irreparable destruction that will befall the environment . Thus, aRQ1: Does awareness, price, availability, value and quality influence university students\u2019 intention to purchase green products?RQ2: How do awareness, price, availability, value and quality predict students\u2019 intention to purchase green products?Against this background, the aim of this study is to assess the effect of university students\u2019 awareness of green products on their green purchase intentions. The specific objectives of this paper are as follows: 1) to identify whether awareness, price, availability, value and quality influence university students\u2019 intention to purchase green products and 2) to find out how awareness, price, availability, value and quality predict university students\u2019 intention to purchase green products. Consequently, the following research questions (RQs) will seek to be answered:This study is significant because, first, not much study on green products awareness has been conducted in emerging countries, especially in a sub-Saharan African (SSA) country. Second, to the best of the author\u2019s knowledge, no study on green products awareness has been carried out among university students in an emerging economy such as Ghana and SSA. Third, through this study, policies could be formulated which could eventually aid in achieving tremendous green consumer behavior standard of living in order to protect the earth\u2019s disappearing resources. Besides, this study\u2019s findings will serve as evidence of practices that university students\u2019 adopt in their bid to protect the environment and prevent its degradation. Additionally, this research can influence and enhance marketing and place emphasis on the exact factors, thereby helping to develop a more optimistic approach toward green purchase intentions. Similarly, an enquiry into the impact of green product awareness on consumers\u2019 green purchase intention could offer valuable evidence to advertising agencies regarding advertising materials along with channels to be used to propagate the message of green product consumption. Moreover, the awareness of the influencing factors will certainly allow students to uphold environmental sustainability through an appeal to consumers in a more appropriate/precise manner. This can result in improved usage of environmentally friendly products that uses fewer resources and causes less harm to the environment.\u201cenvironmental sustainability as the third aim beyond consumers\u2019 satisfaction and company profitability.\u201d The major challenges of green marketing are the lack of public consensus on what encompasses green [Green marketing has become very important in contemporary marketplace. Green marketing comprises all-inclusive marketing activities of packaging, product modification and manufacturing processes that are done in an environmentally harmless manner while meeting the needs of customers\u2019 . Pagliaces green , the neees green . Ensurines green .The concept of the \u201cgreen consumer\u201d has become the pivot around which marketing strategies relating to the environment have been concentrated by marketing professionals and scholars . Green cGreen products (GPs) are commodities which normally bear characteristics such as energy efficient, recyclable, low emitting, healthy products and the likes . GPs areH1: Green products awareness is positively influenced by university students green purchase intentions.Price criterion usually serves as the main hindrance to green products purchase and that green consumers\u2019 are only ready to pay a premium for a product if they realize that its attributes, designs and functions are beneficial to them, their families and posterity , 46, 63.H2: Green perceived price is positively influenced by university students green purchase intentions.It is believed that awareness is created before the availability of a product. A study confirmed that the rate of awareness of green products (GPs) is higher than availability of the green products . It is nH3: Green perceived availability is positively influenced by university students green purchase intentions.of a product/service between what is received and what is given based on a consumer\u2019s environmental desires, keeping judgments, and needs to make a product environmentally friendly.\u201d In sum, GPV is the consumer\u2019s imagination of the accumulated benefits from the usage of a green product. Of recent, GPV is very crucial and that it has a positive effect on marketing and environmental performance of a product and increases its purchasing intention [Green perceived value (GPV) refers to a green product\u2019s overall characteristics, benefits and performance in consumers thought processes. In short, GPV denotes a customer\u2019s evaluation of the entire benefits that will be accrued to him or her from green products . Accordintention . Therefontention , 72. Accntention , there intention . Also, cntention . Again, ntention . BesidesH4: Green perceived value is positively influenced by university students green purchase intentions.One of the important determining factors that impact the purchasing of green products is quality . PerceivH5: Green perceived quality is positively influenced by university students green purchase intentions.\u201cindications of how hard people are willing to try, of how much of an effort they are planning to exert, in order to perform the behavior\u201d [It must be noted that consumers\u2019 intention indeed plays a crucial part in marketing strategies. Purchasing intention refers to whatever consumers\u2019 ponder and plan to purchase. Consumers\u2019 behavioral intention is referred to as the possible behaviors which induce consumers commitment or decision to purchase a particular product . Similarehavior\u201d , p.181. ehavior\u201d . A changehavior\u201d . The conehavior\u201d . To enhaehavior\u201d , 11. It ehavior\u201d . Outstanehavior\u201d , 38. In ehavior\u201d , 67. Andehavior\u201d .A quantitative research method was used in this paper so as to test the relationships among the variables of importance . The quaIn this study, the target population comprised students of the five departments of a University\u2019s Faculty of Business and Management Studies. These five departments were: accountancy, marketing, secretaryship and management studies, communication studies, and procurement and supply chain management. The students of these five departments resided in the university\u2019s halls of residence, the suburbs of the Municipality and other towns near the university. The ages of these students ranged from 18 to 31 plus. The ages of the undergraduate students ranged from 26 to 31\u2009+\u2009because, in the university where the study was conducted and as in all universities in Ghana, there are students who are matured, older and advanced in age. These students are already holding Diploma and Higher National Diploma certificates and are already working in various organizations. As a result, the university gives them the opportunity in the evenings and weekends to do a two year top-up leading to bachelor\u2019s degree. Again, the university has a policy of giving admission to students aged 25\u2009+\u2009who are classified as \u201cmatured students\u201d to study for a four-year Bachelor degrees. The study was conducted from October 15, 2019, to January 27, 2020. Data were collected by the use of an online survey.The study targeted the entire population of students 1,109) at the Faculty consisting of five departments. The study attempted to use all the students for the research. However, after engaging/contacting them there were 550 who were ready and willing to be part of the study. Out of the 550 who were given the questionnaire via email, 478 returned their questionnaire and, thus, used in this study. This gave a response rate of 86.9%. This response rate is quite in line with past email-based research reply rates , 49. The,109 at tThese questionnaires were developed using the Google forms platform which were later distributed via emails to the participants. It is worth mentioning that the questions were simple, carefully modified and concisely worded to avoid ambiguity and formatted to avoid errors . MoreoveThe thirty 30) Likert-type measurement items that were used in this research were adopted and modified from previous studies. This was done in order to improve the study\u2019s quality, validity and reliability. With regard to all the variables, GPAW, GPP, GPA, GPV, GPQ and GPI, the questionnaires were adapted from studies such as 0 Likert-, 81. It The analysis of the data collected was done by using SPSS version 23.0 and structural equation modeling (SEM) using SmartPLS 3.0. Anderson and Gerbing  proposedThe SmartPLS estimates were used to test the hypotheses through the measuring of the path, strength and the significance level of the path coefficient. Figure\u00a0The ages of the respondents were from 18 to 21\u00a0years (181) which represents 37.9%, 22\u201325\u00a0years (253) which is 52.9%, 26\u201330\u00a0years (32) with 6.7% and over 31\u00a0years (12) with 2.5%. There were 220 (46%) males and females 258 (54%) students from different departments. With regard to education level, 142 (29.7%) were diploma students, 202 (42.3%) were higher national diploma students, whereas 134 (28%) were bachelor degree students. Table The SmartPLS-SEM is very robust, hence, its use in this study in order to better justify the hypotheses with deeper testing, integrity and contribution to green product awareness and green consumerism literature being studied . To get Having carefully scrutinized the outer loadings, the following items: two factors of green perceived availability , three factors of green perceived price , one factor of green product awareness (GPAW_3\u2009=\u20090.542), two factors of green perceived value , two factors of green perceived quality  and two factors of green purchase intention  were deleted because they had low loadings which fell short of the threshold (<\u20090.60) as recommended by . When thThe SmartPLS algorithm was used to compute the outer loadings composite reliability (CR) and the average variance extracted (AVE). Thus, using the AVE, CR and outer loadings, the convergent validity was measured. The results showed that the AVE stretched from 0.597 to 0.736 and this meets the criterion for the convergent validity. As shown in Table In the measuring of the discriminant validity, the Fornell\u2013Larcker benchmark of comparing square root of AVE with the constructs\u2019 correlations was used. The AVE square roots of the entire constructs were beyond the squared correlations among the constructs, and this indicates satisfactory discriminant validity . This isThe second aspect of PLS-SEM was to build and assess the structural model (SM). In brief, it is the supposed causation between the dependent construct and the independent constructs and this SM has been used to test the hypotheses in this study. To assess the SM, the algorithms and bootstraps were re-calculated and this occurred after some of the items/indicators have been deleted. The r-square, f-square, multicollinearity, standard deviations, the t-values, the p-values and the path coefficients were all completed, and these results are subsequently presented in Tables 2 is the principal criterion used to evaluate the structural model [2 revealed that the variance in the endogenous construct (green purchase intention \u2013 GPI) is explained by the five exogenous constructs . The R2 value of the endogenous variable, GPI (66.9%) in the current study is presented in Table 2 should be greater than 0.26 (26%) for variance explained. This means that the descriptive power for GPI is acceptable among university students. The results derived from the SmartPLS bootstrapping, the structural model and its t-statistics are shown in Fig.\u00a0Ral model . The R2 F2. With effect sizes, the values 0.00 \u2013 0.15 mean small size, 0.15 \u2013 0.35 mean medium and over 0.35 mean large effect [F2 values show that all the endogenous variables GPA (0.004), GPP (0.058), GPQ (0.140), GPV (0.102) and GPAW (0.073) which had small effect sizes.The effect size was measured using e effect , 65. ThiThe predictor variables\u2019 variance inflation factor (VIF) ranged between 1.703 and 3.002, and this fell within the acceptable range where VIF values less than 5.0 mean no multicollinearity problem as acclaimed by . Therefo>\u2009GPI, H5, \u03b2\u2009=\u20090.293; p\u2009<\u20090.001). Following closely is the path green perceived price to green purchase intention . The rest of the path relationships are as follows: green perceived value to green purchase intentions , green product awareness to green purchase intentions  and, finally, green perceived availability to green purchase intentions . It is acknowledged that when the t-statistics is less than 1.96 and the p-value is greater than 0.05, then that particular hypothesis is not statistically significant. Looking at Table The results in Table The results point out that 66.9% of the variance in the dependent variable (green purchase intention) is explained by the five predictors . This implies that this study explains the green purchase intention of university students in an emerging country far better . While sResults of this study indicate that green perceived quality (\u03b2\u2009=\u20090.293) has the utmost significant positive impact on university students\u2019 green purchase intentions. In short, GPQ has the strongest effect on green purchase intentions. This means that students are eager to purchase green products when they identify that the quality is extraordinary if equated with conventional products. This result is in agreement with , 40, 50.Besides, the second strongest predictor of university students\u2019 green purchase intentions is green perceived price (\u03b2\u2009=\u20090.276). This finding shows that the relationship between GPP and GPI is positively significant. The university students\u2019 keenness to buy green products is certainly triggered by their lucid evaluation of the price. Thus, to purchase green products, students will have to assess the price to see how beneficial it will be or whether it is value for money. This again indicates that when prices of green products are similar with conventional products, respondents will definitely have high GPI and for that matter switch to green products. However, low prices of green products will definitely command high occurrence of purchase and that students will switch to green products when the price is the same as their preferred brands. Therefore, price is one of the key features when it comes to the promotion of green products purchases and this confirms hypothesis 2. This result supports earlier study that has been identified by , 24.>\u2009GPI, 0.268) and green product awareness to green purchase intentions (H1: GPAW-\u2009>\u2009GPI (\u03b2\u2009=\u20090.203). The paths of both had sturdier relationships which explained university students\u2019 intention to purchase green products. Thus, students\u2019 are anticipated to purchase products when the value is high, while students\u2019 positive green perceived value of a product could cause higher increase in its purchase [In addition, there were positive path relationships between green perceived value to green purchase intentions , whereas, at the same time, GPA was not positively linked to students\u2019 green purchase intentions. The implication is that availability is not a critical influencing factor when it comes to university students\u2019 green purchase intentions and subsequent behavior. In fact, this is very surprising, but what could have accounted for this? Although this result is supported by [Conversely, the result showed that there was no statistically significant relationship between green perceived availability and green purchase intentions  to identify whether awareness, price, availability, value and quality influence university students\u2019 intention to purchase green products and (2) to investigate how awareness, price, availability, value and quality predict university students\u2019 intention to purchase green products.This study\u2019s results suggested that green perceived quality has the most significant positive influence on university students green purchase intentions, whereas green perceived price is the second best predictor of university students green purchase intentions. However, the result revealed a negative relationship between perceived availability and green purchase intentions and that there was no statistically significant relationship between green perceived availability and green purchase intentions.This research has made contributions to the extant literature on green consumerism by focusing on the green products awareness in emerging SSA markets such as Ghana. Again, as a contribution to knowledge, it is worth mentioning that the present moment of COVID-19 pandemic across the globe makes this study important as businesses and governments\u2019 seek to encourage consumers/citizenry to purchase green products for environmental sustainability.The study\u2019s findings show that green product awareness effect on green purchase intention of university students is greatly driven by awareness, perceived price, perceived value, perceived quality and perceived availability. As regards the extent of the impact, it was found that GPQ and GPP correlated highly with GPI. Secondly, this study implies that it is among the few researches that consider how the awareness of green products by university students is impacted by GPQ and GPP in an emerging SSA market\u2019s perspectives. Thus, the results show that GPQ significantly predicts GPI of university students and this indeed contributes to the literature.The study\u2019s findings revealed that GPQ and GPP have strong positive relationships with GPI of university students. As a result, policy makers, marketers, green manufacturers and governments could influence GPQ by launching marketing campaigns/promotions that will help to distribute knowledge on the benefits of green products. Those campaigns may improve consumers\u2019 knowledge and understanding on green products and encourage positive approaches regarding its purchasing. It is necessary for consumers to be educated on the benefits that will accrue if they invest in green products. In so doing, they will be tempted to pay more for green products. Again, to resolve the issue of pricing, consumers\u2019 could be offered discounts so as to motivate them to purchase green products.This study was conducted among undergraduate students, in a single university with the setting in one city, and therefore, the results cannot be generalized to all students. It is hoped that in future, more cities, more universities and all age categories of the population would be included. Also, this study did not stipulate particular kinds of green products, so, in future research, could cover specific green products. Finally, the research only considered students green purchasing intentions, so future research should focus on assessing real behavior of students who purchase green products frequently."}
+{"text": "This study systematically analyzes the factors that affect consumers\u2019 green purchase intention. Through a comprehensive literature review, the influencing factors of consumers\u2019 green purchase intention are organized into three categories: cognitive factors, consumer individual characteristics, and social factors. Next, a meta-analysis of 54 empirical papers was conducted using Comprehensive Meta-Analysis 3.0 software to quantitatively assess these relationships. The results revealed that green perceived value, attitude, and green trust have a significant positive influence on green purchase intention. Perceived behavior control, perceived consumer effectiveness, and subjective norm also has a strong positive impact on green purchase intention. Collectivism has a positive effect on green purchase intention. Green perceived risk has a significant negative impact on green purchase intention. The study\u2019s findings provide references for enterprises engaged in green product diffusion and organizations responsible for environmental protection. Rapid economic development and technological progress bring even more convenience to people\u2019s lives but also pose many challenges to the environment, such as air pollution, climate change, and global warming. These problems directly affect the sustainability of economic development, the environment, and society. It has also attracted the attention of all parties to the environment. In the past few decades, environmentally conscious consumers have achieved positive and significant growth in environmental protection activities, attitudes and knowledge. People are paying more and more attention to the environment, which directly affects the changes in personal lifestyles and values. In the case of realizing the importance of the environment, many consumers also realize that their purchasing behavior will have an impact on the ecological environment. Consumers began to change their lifestyles and business activities, and gradually tended to increase consumption of green products . Green pConsumers\u2019 demand for green products has also prompted companies to pay attention to the market for green products. As sustainable development becomes a trend, the development of green products has become a broad field of social progress and commercial development, involving consumers and enterprises. As an important part of economic development, companies themselves have begun to pay attention to environmental issues. Because the development of green business helps reduce the cost of excessive useless waste, provide employees with a safe and healthy working environment, and ensure the sustainable and efficient operation of the enterprise. Therefore, companies have also begun to seek the coordinated development of environmental protection and economic growth, and strive to achieve a green economy. In order to gain a larger market for green products, companies have developed various green products to meet the needs of consumers . The chaAlthough consumers have more demand for green products and enterprises have a strong willingness to develop green markets, the degree of market development for green products is still insufficient. In one prior study, about 30% of consumers expressed concern about the environment and tried to translate this into their purchase behavior . HoweverConsumers\u2019 attention to the environment and green products will affect their purchase decisions . To promThe theory of consumer behavior is the main theoretical method of marketing . AccordiThe rest of the study is arranged as follows. Section \u201cTheoretical Framework\u201d proposes hypothetical development based on the analysis of the existing literature; Section \u201cMaterials\u201d prepares data for meta-analysis through data collection, data coding and data analysis; Section \u201cResults\u201d analyzes the specific process and final results of the meta-analysis; Section \u201cDiscussion\u201d discusses the results of the meta-analysis; Section \u201cImplications\u201d discusses the implications of the research results; Section \u201cConclusion\u201d draws conclusions and limitations of the research.The theory of consumer behavior is one of the main theoretical methods of marketing. According to the theory of consumer behavior , there aThe theory of planned behavior TPB;  is one oTherefore, based on the theory of consumer behavior, combined with the expansion of TPB model and ABC theory, this study puts forward a theoretical framework to influence consumers\u2019 green purchase intention. The first category is cognitive factors, including green perceived value, green perceived quality, green perceived risk, perceived behavioral control, and perceived consumer effectiveness, environmental knowledge. The second category is consumer individual characteristics, including environmental concern, green trust, and attitude. The third category is social factors, including subjective norm and collectivism.Purchasing intention is usually defined as a prerequisite for stimulating and pushing consumers to actually purchase products and services. Many studies examine consumers\u2019 intentions to test their actual behavior. The TPB model provides a valuable framework for studying consumers\u2019 green purchase intentions. The new variables that comprehensively affect these behavioral intentions will enhance the explanatory power of the TPB model. The literature on green purchase intention has been studied through cognitive antecedents of behavior. Cognitive factors in this study refer to consumers\u2019 perception of green products, which is likely to have an important impact on green purchase intention.Green perceived value refers to consumers\u2019 overall appraisal of what they give for and receives from a product or service, based on their environmental desires, sustainability expectations, and green needs . More geGreen perceived quality is consumers\u2019 judgment of the environmental excellence of a brand . PerceivPerceived risk is the subjective prediction of loss, and consumers typically aim to minimize perceived risk. Perceived behavioral control refers to an individual\u2019s judgment of their ability to perform a specific behavior . As an iPerceived consumer effectiveness is the degree to which consumers think their individual actions contribute to solving problems\u2014a subjective perception of the role of one\u2019s own efforts . PerceivEnvironmental knowledge is knowledge of the environment, key relationships that lead to environmental impacts, and environmental responsibility of the individual that leads to sustainable development . In the H1:Green perceived value has a positive effect on green purchase intention.H2:Green perceived quality has a positive effect on green purchase intention.H3:Green perceived risk has a negative effect on green purchase intention.H4:Perceived behavioral control has a positive effect on green purchase intention.H5:Perceived consumer effectiveness has a positive effect on green purchase intention.H6:Environmental knowledge has a positive effect on green purchase intention.Because consumers are heterogeneous, there are also differences in their purchase intentions for green products. Although researchers have improved the explanatory power of TPB by adding personality constructs , there aEnvironmental concern is the degree of concern over environmental problems, and indicative of efforts to solve these problems . EnvironTrust is considered a common mechanism for reducing perceived transaction risk by increasing expectations of positive outcomes and certainty of how trustees will behave. As one of the three core concepts in the theory of planned behavior, attitude refers to the positive or negative evaluation or appraisal of a particular object . AttitudH7:Environmental concern has a positive effect on green purchase intention.H8:Green trust has a positive effect on green purchase intention.H9:Positive attitude has a positive effect on green purchase intention.Consumer purchase intention for green products is affected by not only individual factors but also the social environment and other people. Social factors affect individual behavior decisions in many ways, such as social pressure from other people and collectivist ideas. This research mainly studies the impact on green purchase intention from two aspects: subjective norms and collectivism.Subjective norm refers to the social pressure perceived by individuals on whether to carry out or refrain from a certain behavior . In deciCollectivism is an important value that affects people\u2019s decision-making and consumption behavior . It holdH10:Subjective norm has a positive effect on green purchasing intention.H11:Collectivism has a positive effect on green purchasing intention.Based on the above hypotheses, the proposed research framework is depicted in This study comprehensively searched the relevant literature. Works were mainly retrieved from the following databases: Springer, EBSCO, Emerald, Elsevier, CNKI, Google Scholar, and Web of Science. In terms of time frame, the search included studies published or made available online from January 2010 to October 2020. Searches used a combination of the following keywords: \u201cgreen products,\u201d \u201cgreen purchase intention,\u201d \u201cgreen purchase behavior,\u201d \u201cgreen perceived value,\u201d \u201cgreen perceived quality,\u201d \u201cgreen perceived risk,\u201d \u201cperceived consumer effectiveness,\u201d \u201cperceived behavioral control,\u201d \u201cenvironmental knowledge,\u201d \u201cenvironmental concerns,\u201d \u201cgreen trust,\u201d \u201cattitude,\u201d \u201csubjective norm,\u201d and \u201ccollectivism.\u201dt-values and F-values; (3) independent samples\u2014 if multiple studies used the same sample for empirical analysis, only one was selected for the meta-analysis; (4) clear sample size; and (5) testing at least two variables or relationships of an original or modified model. According to the criteria, we search and screen studies. First, we found 1,938 studies in the database. After eliminating duplicates, 1,379 studies were identified. After scanning the title or abstract, 680 remained. Then, to ensure the accuracy of the research results, the full text of each study was screened according to the inclusion criteria, which left 74 studies. Finally, after excluding studies that lacked important evidence or did not otherwise meet the requirements of our meta-analysis, 54 studies were retained. These studies are all journal articles. The screening process is shown in Studies were screened using the following criteria: (1) quantitative research related to green purchase intention, excluding theoretical research, reviews, etc.; (2) containing correlation coefficients or other values convertible to them, such as t-values and F-values, that can be converted into them. Correlation coefficients are used as the evaluation index in this meta-analysis. After each study was coded, a certain number of studies were randomly selected and coded by another researcher to ensure the accuracy of coding. Any inconsistencies in coding content were resolved through revisiting the original text and discussing until consensus was reached. The 54 studies included in the meta-analysis are listed in As the data used in this research were derived from other studies differing greatly in terms of sample selection and data processing, it was necessary to code the data uniformly. To ensure the reliability of coding, the work was performed by two graduate students. The coding mainly includes two types of content: description items and the effective values of statistical items. The main description items are author, sample size, country or area, respondents, and year of publication. The main effect values of statistical items are correlation coefficients or other statistics, such as The core objective of the meta-analysis is to calculate the effect values for comparative analysis. Effect values reflect the intensity and direction of the relations among variables. If, the effect value and variable relation are both positive, then the higher the effect value, the stronger and more stable the variable relation. This study adopted Hunter and Schmidt\u2019s meta-analytic approach by using correlation coefficients to measure effect values . Becauser as the input effect size, adjust the effect value according to the following formula:First, this study used the correlation coefficient r\u2032 is the correlation coefficient of each study after reliability correction, and r represents the correlation coefficient of each independent study, that is, the effect size; \u03b1xx represents the reliability coefficient of the independent variable, and \u03b1yy represents the reliability coefficient of the dependent variable. This step is to correct the attenuation deviation of correlation coefficient due to reliability defect of the scale. For the missing reliability values of some variables in few studies, we used the weighted average reliability of other similar research samples instead.Where Z conversion of the effect size r\u2032. The calculation formulas are as follows:Second, obtaining the initial effect value using Fisher\u2019s r\u2032 is the correlation coefficient of each study after reliability correction, and ni is the sample size corresponding to the influence value of i.Where This study used Comprehensive Meta-Analysis 3.0 software. Meta-analysis is a statistical method for comprehensively and systematically analyzing many research results . It has N  as the threshold for publication bias: if the fail-safe N is greater than 5k + 10, there is no publication bias. As N is 10363 , which is well above the critical value of 280. As 10,363 unpublished studies would be needed to reverse this conclusion, there is no obvious publication bias problem in the analyzed studies.Publication bias is generally measured by the fail-safe N , which iZ effect value, and the longitudinal axis is the standard error of the Fisher\u2019s Z effect value. It can be seen from the figure that the funnel plot is usually funnel-shaped and symmetrically distributed, and the scattered points are mainly concentrated at the top of the funnel and near the average effect value, which indicates that the possibility of publishing bias in this study is very low.Q-value and I2 value. The Q-value was significant (p < 0.05), which indicates that there is heterogeneity between studies. I2 values of 25, 50, and 75%, respectively, correspond to low, middle, and high heterogeneity. If the distribution of the effect value is heterogeneous, a random-effects model is used; otherwise, a fixed-effects model is used. A fixed-effects model uses the differences within the study to calculate weight, assuming that all studies included in the meta-analysis use the same population and produce only one real effect value estimate. That is, the model considers that all possible factors affecting the effect value are the same, with any differences between studies coming only from negligible sampling errors. By contrast, a random-effects model uses intra- and inter-study differences to calculate weights, assuming that there are differences in the true effect values included in the meta-analysis and that the effect values depend on the subject characteristics, experimental methods, etc. This model also considers that the effect value calculation is influenced by between-study differences and within-study sampling errors. A random-effects model can also estimate the average value of the effective distribution and prevent underestimation of the weight of small-sample studies or overestimation of the weight of large-sample studies. Based on the results of the heterogeneity test, which are shown in To evaluate the significant level of difference between different results, we need to conduct a heterogeneity test. The statistical results of heterogeneity are generally based on Sensitivity analysis can also be understood as robustness analysis, which is an important method to evaluate the robustness and reliability of the combined results of meta-analysis. It is a common sensitivity analysis method to eliminate each included study one by one and then merge the effect quantity, change the exclusion criteria or eliminate a certain kind of literature before merging the effect quantity. Sensitivity analysis is to exclude meta analysis after low-quality research, or to include meta analysis after exclusion study. For example, after excluding a low quality study, the combined effect quantity is re-estimated and compared with the meta-analysis results before exclusion, and the influence degree and robustness of the study on the combined effect quantity are discussed. If the result does not change significantly after the exclusion, it means that the sensitivity is low and the result is more robust and credible; on the contrary, if there is a big difference or even the opposite conclusion after the exclusion, it means that the sensitivity is high and the robustness of the result is low. Based on the above theory, this study uses CMA software to perform the \u201cone study removed\u201d operation to check whether the effect size has changed. According to the results, it is found that the total effect value is 0.460, and the effect value between each relationship is distributed between 0.457 and 0.465. The results have not changed greatly, indicating that the sensitivity is low, and the results of this study are more robust and credible.p < 0.001) and green purchase intention. Green perceived value (r = 0.596) had the largest influence on green purchase intention. The effect size of perceived behavioral control (r = 0.499), perceived consumer effectiveness (r = 0.493), and green perceived quality (r = 0.462) each have medium correlations with green purchase intention. Green perceived risk (r = \u22120.373) had a negative effect on green purchase intention. Therefore, H1, H2, H3, H4, and H5 are supported.r = 0.530) and attitude (r = 0.535) were strongly correlated with green purchase intention, while environmental concern (r = 0.461) and environmental knowledge (r = 0.335) had a moderate correlation with this outcome variable. Therefore, H6, H7, H8, and H9 are supported.Regarding the relationship between consumer individual characteristics and green purchase intention, green trust  was significantly positively correlated with green purchase intention, and collectivism  was positively correlated with green purchase intention. Therefore, H10 and H11 are also supported.For social factors, subjective norm (This study used a meta-analysis to explore the influencing factors of green purchase intention. The results from 54 studies were analyzed, including 11 variables related to green purchase intention. The meta-analysis results show that green perceived value, attitude, and green trust each have a great positive impact on green purchase intention. Consumers are driven by value, often weighing the benefits and utility they gain when buying products . When coPerceived behavioral control, perceived consumer effectiveness, and subjective norm each have a moderate correlation with green purchase intention. The effect of perceived behavioral control mainly reflects consumers\u2019 stronger willingness to buy when they are more confident in their purchasing ability, which has been identified in previous studies . TherefoGreen perceived quality, environmental concern and environmental knowledge also have some influence on consumers\u2019 green purchase intention. Consumers who pay more attention to the ecological environment and have relevant knowledge are more willing to buy green products . TherefoFinally, green perceived risk has a negative impact on green purchase intention. The greater the risk consumers perceive in green products, the more reserved their attitude toward them and the lower their green purchase intention. Therefore, it is necessary for enterprises to minimize consumers\u2019 perceived risk regarding green product purchases.Based on the results of the meta-analysis, this study can provide a reference for companies to formulate marketing policies. The results of this study show that consumers\u2019 green perceived value, attitude and green trust are important indicators that affect consumers\u2019 green purchase intention. If companies want to improve consumers\u2019 green purchase intention, they should embed green perceived value, attitude and green trust in their long-term strategic planning. As the most important factors to influence green purchase intention, enterprises must improve green perceived value in order to improve consumers\u2019 green purchase intention in the environmental era. Therefore, marketers should formulate marketing strategies to improve consumers\u2019 perception of green value. Regarding attitudes, enterprise can use promotion and other methods to attract consumers\u2019 attention to green products, provide consumers with more experience opportunities, establish a good image of the practicality of green products, and encourage more green attitudes. In addition, the government can also cultivate people\u2019s green attitudes through social media and spread the benefits of green products to the public through various online channels. Companies should strengthen the environmental functions of green products and the environmental image of brands, and provide green products that meet consumer expectations to enhance consumer trust. In addition, enterprises can cultivate experienced retailers as effective and reliable information channels between consumers and manufacturers, salesmen should convey the environmental attributes and environmental protection effects of green products to consumers, so as to enhance consumers\u2019 trust, and thus enhance their green purchase intention.Moreover, we should also pay attention to the impact of perceived behavioral control, perceived consumer effectiveness and subjective norm on green purchase intention. When individuals are more confident in their purchasing ability, they are more likely to purchase products. In this perspective, enterprise should provide consumers with reliable information on the benefits of green products. Effective information is important to their decision-making process, which ensures that they have confidence in their ability to purchase green products. To enhance perceived consumer effectiveness, enterprises should transmit specific ideas to consumers through green labels, encourage them to participate in environmental protection, and clearly inform consumers how they contribute to environmental protection by buying green products. Policymakers can strengthen social norms of energy-saving behavior. For example, they could organize scale green environmental protection activities or use social media to disseminate norms encouraging more green purchasing behavior.Besides, perceived risk also plays a role in explaining consumer behavior, because consumers are often motivated to reduce risk rather than maximize effects in the purchase process. Because green perceived risk reduces green purchase intention, marketers need to eliminate and reduce their perception of green risk in purchasing products.This study uses meta-analysis to analyze the factors that affect consumers\u2019 green purchase intention based on previous studies. On the basis of the theory of consumer behavior, this study puts forward that the factors affecting consumers\u2019 green purchase intention mainly include three categories: cognitive factors, consumer individual characteristics and social factors. Meanwhile, combine the expansion of TPB model and ABC theory, this study puts forward a theoretical framework to influence consumers\u2019 green purchase intention. In previous studies, there is a conflict between some research results and a lack of consensus on the analysis of the influencing factors of consumers\u2019 green purchase intention. This study uses the meta-analysis method to overcome the limitation of sampling error and sample size of a single study, and to solve the conflict results in the study of the influencing factors of consumers\u2019 green purchase intention. The results show that, green perceived value, attitude, and green trust were most strongly positively correlated with green purchase intention, while perceived behavioral control, perceived consumer effectiveness, subjective norm, green perceived quality, and environmental concern were each moderately correlated with green purchase intention. Compared with other factors, environmental knowledge and collectivism had a relatively weak effect on green purchase intention, while the green perceived risk was negatively correlated with this outcome variable. According to the results, the assumptions involved in the study have been verified. This study\u2019s findings enrich the theoretical basis for understanding consumers\u2019 purchase intention for green products, provides new methods and ideas for exploring the influencing factors of green product purchase intention. Meanwhile, the results of this study can provide reference for enterprise marketing and government environmental protection propaganda. Enterprise marketers can make better use of the results of this study to formulate marketing strategies; government departments can publicize the advantages of green products and guide consumers to carry out environmental protection.However, this paper has some shortcomings. First, due to incomplete information, the factors used for meta-analysis are limited to those for which sufficient data are available, some factors had to be excluded from the meta-analysis. Second, although heterogeneity in the sample was confirmed, we did not analyze the causes of heterogeneity. Third, there is no moderating effect test for the relationship in this study. In future studies, we should consider the variables with fewer data in this research, explore the influence of other factors on green purchase intention and the reasons for the heterogeneity.The original contributions generated for this study are included in the article/supplementary material, further inquiries can be directed to the corresponding author.WZ contributed to research design, reference searching, data coding, formal analysis, and manuscript writing. XL contributed to conceptualization, secondary data coding, and funding acquisition. MR contributed to data coding and manuscript revision. All authors contributed to the article and approved the submitted version.The 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."}
+{"text": "The market for green agricultural products has tremendous growth potential as the pressure on resources and the environment increases and the safety of agricultural products is garnering attention. The demand for green food (tea) is also rising as tea is among the top three beverages consumed worldwide. The study attempts to propose a model of the relationship between green food (tea) customers\u2019 product knowledge, perceived product quality, trust, purchase intention, and purchase behaviour. In addition, we will provide an analysis of the role played by age, education, income, gender, etc. The study included 700 questionnaires on green food (tea) consumers that were collected through the Credemo questionnaire platform, and data analysis was carried out using the SmartPLS software to assess the model of product knowledge on green food (tea) consumption behaviour. The findings demonstrate that, concerning differences in age, education, income, and gender; product knowledge\u2014including attribute knowledge and green knowledge\u2014has a positive impact on perceived product quality and trust; perceived product quality has a positive impact on trust; perceived product quality and trust have a positive impact on purchase intention; and purchase intention has a positive impact on purchase behavior. Global environmental problems  frequently occur , triggerConsumers who lack knowledge of environmental issues are less likely to act in an environmentally friendly manner because knowledge is the biggest predictor of environmentally friendly behavior ,12. ProdFirst, it can be because the studies mentioned above did not focus on the same subjects . Second, the above scholars do not define and classify green product knowledge in the same way, either as product knowledge, green firm knowledge, label knowledge and environmental knowledge, mostly as a higher-order unidimensional variable ,24,25,26There are two primary categories of a recent study on consumers\u2019 purchase intentions of green products. First, from the standpoint of internal variables, researchers have found that consumers\u2019 purchase intentions of green products are significantly influenced by attitudes, perceived value, perceived risk, perceived quality, perceived behavioural control, perceived effectiveness, and environmental concerns ,33,34,35The study makes an effort to elaborate on the following research-related issues. Firstly, constructing a model of how product knowledge (attribute knowledge and green knowledge) affects consumers\u2019 purchase intention and behaviour around the consumption of green food-tea. Secondly, expanding the areas in which consumer product knowledge theory can be applied. The study used green tea as its research object, and 700 green food consumer surveys were gathered to build a structural equation model with numerous clusters and to examine the model of consumer product knowledge on green tea purchase behaviour. This will serve as a guide for increasing green consumption and lowering the ecological and environmental pollution.The word \u201cattribute knowledge\u201d describes pertinent, precise knowledge about the characteristics, lingo, frameworks, standards, etc., of green foods . AccordiConsumers have more faith in independent third-party certification organizations than they have in the product\u2019s manufacturers . This isThe hypothesis that is generated from the analysis above is as follows.Hypothesis\u00a01\u00a0(H1).Consumer attribute knowledge significantly improves the perceived product quality of green tea.Compared to regular food, green food has qualities such as being safe, healthy, and environmentally friendly . AdditioThe hypothesis that is generated from the analysis above is as follows.Hypothesis\u00a02\u00a0(H2).Consumer attribute knowledge significantly improves trust in green tea.The term \u201cgreen knowledge\u201d refers to consumers\u2019 subjective understanding of the environmental and food safety benefits of \u201cgreen\u201d foods . It is aConsumers with green knowledge are often more concerned about environmental and ecological issues, and they frequently relate products with ethics ,49. ConsThe hypothesis that is generated from the analysis above is as follows.Hypothesis\u00a03\u00a0(H3).Consumer green knowledge significantly improves the perceived product quality of green tea.Related research has demonstrated that as the market for green products grows and green information spreads broadly, consumers begin to feel more trust in green items . ParticuThe hypothesis that is generated from the analysis above is as follows.Hypothesis\u00a04\u00a0(H4).Consumer green knowledge significantly increases trust in green tea.As consumers base their purchasing decisions on incomplete and asymmetrical information, the perceived product quality is a signal from the company to the consumer based on which the consumer develops a sense of trust . ConsumeThe hypothesis that is generated from the analysis above is as follows.Hypothesis\u00a05\u00a0(H5).Consumer-perceived product quality significantly increases trust in green tea.By affecting the perceived advantages, which are higher than the costs when the perceived quality is higher, the perceived quality of the product or service can improve the consumer\u2019s purchase intention . When maThe hypothesis that is generated from the analysis above is as follows.Hypothesis\u00a06\u00a0(H6).Consumer-perceived product quality significantly increases the purchase intention of green tea.Consumers\u2019 trust in food firms\u2019 products regarding food safety, health, and environmental protection is referred to as trust. Due to the unique characteristics of green foods and the information asymmetry in the market, consumers can only evaluate the quality of a product through personal experience and product expertise, making the intake of green food risky . When coAccording to trust theory, a high level of trust is necessary for the establishment of a long-term, stable relationship with consumers and the creation of value in their transactional relationship. This helps consumers form a good corporate image and increases their likelihood of making a purchase . RelatedThe hypothesis that is generated from the analysis above is as follows.Hypothesis\u00a07\u00a0(H7).Consumer trust significantly increases the purchase intention of green tea.According to the Theories of Planned Behaviour and Rational Behaviour, an individual\u2019s behaviour can be inferred from behaviour intention. Behaviour intention is a measure of behaviour . AccordiHypothesis\u00a08\u00a0(H8).Consumer purchase intention significantly increases the purchase behaviour of green tea.The SOR (Stimulus-Organism-Response) theoretical model has been used extensively in the field of consumer behaviour . SOR theThe sample size of the study was calculated using Gpower3.1 and with an effect size f = 0.02, significance level = 0.05, test validity power = 0.80, and tested predictors = 5, the sample size was 647, therefore the study set a sample size of 700, which is sufficient to justify the adequacy of the study sample .The study was based on a questionnaire collected by Credamo, an internationally recognized questionnaire collection platform, and an additional RMB 595 was spent to set the completion conditions to ensure the quality of the returned sample. Firstly, a credit score of 80 or above was set, as the higher the credit score of the sample, the higher the quality of the completed questionnaire. Secondly, we set the sample\u2019s historical adoption rate to be greater than or equal to 80 to avoid the low quality of the questionnaire due to the unfamiliarity of the process. Thirdly, a sliding puzzle was set for intelligent human verification before answering, which helped to improve data quality and security. Fourthly, only one participant within 5 km was set to avoid sample homogeneity due to the bunching of participants.The questionnaire was randomly distributed to all provinces in China in September 2022, and 700 questionnaires were returned, with a participant reward amount of RMB 3 per questionnaire. Two screening questions were set, the automatic screening question \u201c20 + 40 = ?\u201d and \u201cHow often did you buy green tea in one year (repeated question)\u201d. During data cleaning, samples with inconsistent responses were removed to obtain the final sample for this study to ensure the reliability of the sample.The overall questionnaire is divided into three main blocks. The first block is a brief graphic introduction to green foods. The second block contains questions related to the study variables , external factor loadings, FORNELL, and heterotrait-monotrait (HTMT) ratio. The external factor loadings were all more than 0.7, as shown in As can be seen in The findings of the model evaluation using a bootstrapping subsamples 5000 study are displayed in All constructs are higher than the crucial value of 0.25, as determined by capacity . All conociation . The Staociation .p-value is less than 0.05 [p < 0.000 and did not cross 0 at the 5\u201395% confidence interval), and H1 was tested. Attribute knowledge had a significant positive effect on trust , and H2 was tested. Green knowledge had a significant positive effect on perceived product quality , and H3 was tested. Green knowledge had a significant positive effect on trust , and H4 was tested. Perceived product quality had a significant positive effect on trust , and H5 was tested. Perceived product quality had a significant positive effect on purchase intention , and H6 was tested. Trust had a significant positive effect on purchase intention , and H7 was tested. Purchase intention had a significant positive effect on purchase behaviour , and H8 was tested.The study employed PLS-SEM to assess the model\u2019s hypotheses, and the data\u2019s findings are displayed in han 0.05 ,92, and The outcomes of the multi-group analysis are displayed in The results of the multi-group analysis, which split the respondents\u2019 ages into middle-aged and older (30 years and older) and young (30 years and below) groups, showed that the influence of attribute knowledge on trust was not significant for older individuals. Multiple analyses of gender revealed that the relationship between attribute knowledge and trust was not significant for male individuals. From the overall results, those with relevant knowledge background, youth, and females, have higher awareness of green food labels, and label awareness further influences consumers\u2019 label attitude evaluation (trust). This may be because groups with better knowledge of attributes are more willing to trust green food labels and make positive evaluations, building positive beliefs based on the knowledge gained .Previous studies have shown that knowledge is an important variable in predicting consumer pro-environmental behaviour . HoweverFirst, it deepens the research related to green product knowledge. Previous studies on green product knowledge have mainly been conducted from the perspectives of environmental knowledge, overall knowledge, subjective knowledge, brand knowledge, cost knowledge, and quality knowledge ,51,94,95Second, it is necessary to improve the mechanism of the influence of product knowledge on green tea consumption behaviour. There are previous studies related to product knowledge on purchase intention, purchase intention on purchase behaviour, trust on purchase intention, and knowledge on trust ,25,26,67Third, the effect of green knowledge on perceived product quality was greater than that of attribute knowledge. This finding is similar to the findings of previous studies on environmental knowledge and label knowledge on attitudes , but theFourth, perceived product quality has a significant positive effect on trust. This finding is similar to that of previous studies on millennial samples , but differs in three ways. First, this study sample is more generalizable to the entire social sample. Second, that study was on brands, while this study is on food products, which extends the scope of the study. Finally, there is a gap between information quality and perceived product quality; information quality refers to the validity, accuracy, and timeliness of information, while perceived product quality focuses on consumers\u2019 perceptions of the product .Fifth, perceived product quality has a significant positive effect on purchase intention. This finding is similar to the findings of previous studies on suboptimal agricultural products, but with the following differences. The study started the research on food products from a negative perspective, i.e., lower quality food , while this study started the research on food products from a positive perspective, i.e., green tea , which helped to broaden the scope of application of the study .Sixth, trust has a significant positive effect on purchase intention. This finding is similar to the findings of a previous study (green trust positively affects green purchase intention) for electronic products in Taiwan . This suSeventh, purchase intention has a significant positive effect on purchase behaviour. This finding is similar to previous studies on overall green products (green purchase intention positively affects green purchase behaviour) , but witFirst, the data in this study were collected at a single point in time, and there may be seasonal influences that do not accurately predict changes in green food consumer attitudes and behaviours throughout the year. Second, the data in this study were collected in China, and there may be influences from the degree of economic development, consumption habits, consumer attitudes, and national cultures of different countries. Without the influence of these boundary conditions, the findings may not accurately predict the behaviour of green food consumers in every country in the world.First, future research can be launched to collect data at different time points and conduct in-depth tracking studies. Data from multiple periods can be used to analyse the dynamics of consumer purchasing behaviour. It is also possible to use experimental methods for analysis, to control the main research variables, and to find the impact of variables such as time point and season. Second, the number of questionnaires distributed and the scope of distribution should be broadened so that the sample can cover all types of countries more comprehensively and make the sample more representative. Future research can distribute questionnaires to different countries, add boundary variables such as economic development level, consumption habits, culture, etc., involve more consumer groups, and conduct cross-regional comparative analysis.Based on the SOR model, this study investigates the consumption behaviour of green tea by using green knowledge and attribute knowledge as external stimulus variables, perceived product quality and trust as mechanism variables, and purchase intention and behaviour as response variables, and draws the following conclusions. Consumers\u2019 attribute knowledge and green knowledge have a significant positive effect on perceived product quality and trust, most likely because green knowledge, compared to attribute knowledge, is part of the quality perceptions of the product that consumers have already formed in their subjective consciousness about green tea, and thus green knowledge has a higher impact on the product quality of green tea . Green kFirstly, when designing the packaging of tea products, green tea information should be displayed to the greatest extent on the outer packaging so that consumers can quickly access green tea-related knowledge. For example, the green food certification can be designed in a prominent position on the outer packaging.Secondly, when designing green tea publicity manuals, one should add various standards related to green tea from production, origin, storage, transportation, use of chemical fertilizers and pesticides, etc., in the manuals, and compare them with ordinary tea to cultivate consumers\u2019 subjective green knowledge of green tea from objective attribute knowledge.Thirdly, one should enhance the advertising and publicity of green tea. Green tea advertisements can be placed on TV, Taobao, JD, PDD, Tik Tok, Quick hand, WeChat, QQ, Weibo, Red, Bilibili, buses, and taxis to increase product publicity and enhance consumers\u2019 product knowledge awareness, product quality perception, and product trust in green tea.Fourthly, enhancing product knowledge training for green tea sales staff by enterprises is conducive to introducing consumers to the attributes of green tea such as health and pollution-free, food safety, and environmental protection before sales, thereby enhancing consumers\u2019 product knowledge.Fifthly, through technical innovation and service enhancement, consumer trust is enhanced. On the one hand, through technological innovation, the quality of green tea  will be improved, and corporate self-inspection and market inspection of green tea product quality will be carried out to prevent poor-quality green tea from using green food certification to deceive consumers, thereby enhancing consumers\u2019 trust in the product. On the other hand, one should build green tea communities to provide consumers with perfect pre-sales and after-sales services, and actively communicate with consumers, thereby enhancing the consumers\u2019 perception of product quality and consumers\u2019 trust in the products.Sixth, one should target different consumer segments and conduct precision marketing. Target marketing should be aimed at consumers of different ages, education, income, and gender."}
+{"text": "Gastric cancer (GC) is one of the most fatal cancers worldwide and is generally only detected after it has progressed to an advanced stage. Since there is a lack of comprehensive data on RHOA protein expression of patients with GC, this study utilized a systematic review and meta-analysis to address the limitation. The objective of this meta-analysis was to link GC clinical features with RHOA protein high- vs. low-expressing patients with GC.2, I2; and Q values. The symmetry of funnel plots were inspected for publication bias.The PubMed and Web of Science were used for a systematic literature review of GC related to RHOA. The included studies were obtained from two literature databases from past to Aug 31, 2021, by searching keywords. This meta-analysis followed the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. The odds ratios (ORs) and 95% confidential intervals (CIs) for clinical features were estimated according to the high and low protein expression levels of RhoA. The mean effect sizes of ORs were obtained using the random-effects and fixed-effects models of meta-analysis. Heterogeneity of the studies was assesed by using statistics: \u03c4P\u2009=\u20090.02) and poorly differentiated status (P\u2009=\u20090.02). The association between RHOA positivity and Lauren subtypes was not statistically significant (P\u2009=\u20090.07).Finally, 10 studies including 1,389 patients with GC (735 RHOA-positive and 654 RHOA-negative) were eligible for our meta-analysis to estimate associations between the protein expression and clinical features . In our meta-analysis, RHOA positive expression was determined to have a statistically significant association with UICC stage progression (This meta-analysis suggested that RhoA protein expression in patients with GC was associated with clinical features: UICC stage progression and poorly differentiated status. Our findings are inconclusive but indicate that high RHOA protein expressing patients with GC could predict advanced UICC stages. A large prospective cohort study is required for validation in future. There were 1,089,103 new cases of gastric cancer (GC) worldwide in 2020, with 768,793 deaths .Ras homologous A (RHOA), a Rho family small GTPase, is involved in diverse oncogenic processes, including proliferation, migration, cell polarity, and invasion , as wellRHOA in diverse GC cell lines, cell growth was inhibited, and apoptosis increased [RHOA, tumor size was decreased [Previous studies for RHOA in GC have dealt with biological functions and molecular subtypes. A molecular subtype in GC was associated with RHOA genetic events  in the Cancer Genome Atlas project , 10. RHOncreased . In vivoecreased . DespiteRHOA has been inspected mainly by individual studies in terms of biological functions, and genetic characterization in GC . RHOA fuThe objective of this meta-analysis was to estimate associations between GC clinical features and RHOA protein high- vs. low-expressing GC patients. The clinical utility of RHOA protein expression in patients with GC have not been reported. Thus, this systematic review and meta-analysis summarizes previous publications. This approach inspects whether RHOA protein expression predicts clinical features including GC progression, invasion, and GC cell histology. This comprehensive analysis helps to demonstrate the possible clinical associations of RHOA protein expression and the mean effect sizes of the clinical features in patients with GC by synthesizing evidence from the publications.This study was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA). PubMed and Web of Science were searched to obtain literatures on RHOA expression in GC, in order to identify appropriate publications for a meta-analysis, through Aug 31, 2021, with the following terms: \u201cRHOA,\u201d \u201ccancer,\u201d and \u201cexpression.\u201d Subsequently, the titles and abstracts of the publications that contained term \u201cgastric\u201d were retrieved 2. Studies relating to pathological diagnosis of GC3. Studies of case\u2013control design in GC4. Studies with sufficient data to derive odds ratios (ORs) and 95% confidence intervals (CIs).5. Articles published between the past and Aug 31, 2021.1. Reviews2. Non-English publications3. Access denied studies4. Studies on cancers other than GC5. Studies lacking clinicopathological data for RHOA positive and negative protein expression6. clinicopathological studies for other genes.The included studies were retrospective case\u2013control studies that compared high- and low-RHOA protein expressing patients with GC. These studies had clinical information .To obtain relevant information for our meta-analysis, the two author reviewers (SN and YL) independently assessed the literature that satisfied the two previous criteria. Authors, publication year, study objects, RHOA positivity, technique, and clinical information were included Table .Table 1TThe author (SN) reviewed the whole search, and confirmed the data.Recommendations and quality of evidence of the included studies were evaluated according to the guidelines of Robinson et al.  Table 22.Table 2The R library \"meta\"  was utilThe pooled effect sizes of the ORs were estimated using either random-effects models or fixed-effects models . The pooled effect size of the OR is a critical tool in assessing the clinical relevance of RHOA protein high- vs. low-expressing patients with GC and refers to the collective effect size estimates of the studies.2 and Higgins\u2019 I2 including Cochran\u2019s Q-tests) and was obtained by the R library \"meta\" [2\u2009\u2264\u200950% or P\u2009\u2265\u20090.05 showed the absence of heterogeneity [The heterogeneity was assessed using statistics . In visual inspection of funnel plots, lack of skewness and asymmetry generally indicates an absence of publication bias.The search of PubMed and Web of Science generated 2,208 and 2,372 studies, respectively. After removing duplicates and carefully reviewing the titles and abstracts of the studies, 105 studies were found. Subsequently, the 105 studies were thoroughly reviewed, and 95 were eliminated due to a lack of data and an unclear number of patients. Resultingly, ten publications were selected for meta-analysis . Notably, the fixed effect model results require careful interpretation due to heterogeneity . The association between RHOA positivity and vascular invasion was not statistically significant , and between-study variance could not be ignored.The For sensitivity analysis of UICC stage progression Fig.\u00a0A, the ORThe visual inspection for the funnel plot of UICC stage progression Fig.\u00a0A showed Our research question was to inspect whether clinical features were linked with RHOA protein high- vs. low-expressing patients with GC. Our key finding in this meta-analysis was that RHOA protein-high expressing patients with GC were statistically significantly associated with UICC stage progression, and poorly differentiated status. Our findings suggest that RHOA protein expression is a GC biomarker candidate. Our findings also need to be validated by large prospective cohort studies to secure reproducibility in future.Regarding associations between protein expression and clinical features in GC, a few recent meta-analysis studies were available for Sp1, CD133 and heparanase \u201333. To tRHOA has emerged as a functionally important molecule in GC . RHOA knRHOA has been identified as a mediator of EMT , 35. EMTEMT is involved in cellular morphology changes, mainly by blocking differentiation-related genes . TherefoRhoA signaling is crucial for two clinical aspects , 38, 39:Our meta-analysis has strengths. We believe that the included studies for the systematic review and meta-analysis cover a comprehensive collection of RHOA protein expression measured by IHC in the field of GC. In addition, 1,389 patients with GC in the ten studies provides statistical strengths for robust meta-analyses.There are limitations in this study. Since our meta-analysis utilized published studies, publication bias is unavoidable, indicating statistical heterogeneity is inevitable . The divOur study suggests that high RHOA protein expression is associated with.UICC stage progression  and poorly differentiated status in patients with GC. However, our study suggests the need of prospective large-scale cohort studies for validation, which helps to prove feasibility of RHOA protein expression as a biomarker to predict GC progression."}
+{"text": "Oculomotor nerve palsy is a common and well-described disease with diverse etiologies. Clinicians should quickly and correctly diagnose and treat oculomotor nerve palsy according to its characteristics and the accompanying symptoms and signs. Intracranial aneurysm is an important and frequent cause of oculomotor nerve palsy. Considering the catastrophic consequences of rupture, the possibility of an urgent, life-threatening disease should always be considered.A 63-year-old Chinese woman presented with intermittent left ptosis and diplopia and painless incomplete oculomotor nerve palsy without pupil involvement. She manifested no mydriasis or extraocular muscle weakness, and the light reflex was normal. Other cranial nerves and somatosensory and somatomotor examinations were normal. The neostigmine experiment and electromyography were normal, so the diagnosis of myasthenia gravis was excluded. Brain magnetic resonance angiography showed a 4-mm aneurysm located at the cavernous segment of the left internal carotid artery. Unfortunately, the patient refused digital subtraction angiography and was discharged home without further treatment.Neuroimaging must be performed to exclude intracranial aneurysms in oculomotor nerve palsy regardless of whether the pupils are involved, as aneurysm rupture carries substantial morbidity and mortality. Oculomotor nerve palsy is a common and important neurological disease that classically manifests with ptosis, diplopia and extraocular muscle weakness. The presenting clinical symptoms and signs of oculomotor nerve palsy vary according to the etiologies and locations of the lesion. Oculomotor nerve palsy can result from intracranial aneurysm, trauma, diabetes mellitus, tumors, stroke, and infection. Due to the complexity of its etiologies, comprehensive neurological examinations, blood tests, neuroimaging, and lumbar puncture should be performed to help quickly and correctly diagnose oculomotor nerve palsy. It is also very important to distinguish isolated oculomotor nerve palsy  from those that are not isolated. An ischemic neuropathy and a compressive lesion are leading causes of isolated oculomotor nerve palsy . Traditi9/L, but thrombocytes and red blood cells were normal. Electrolytes, hepatic tests, renal tests, erythrocyte sedimentation rate, vitamin B12, and thyroid tests were normal. Systematic autoimmune autoantibody testing was unrevealing. Blood testing of treponemal results with rapid plasma reagin (RPR) was negative. The initial diagnosis of ocular myasthenia gravis was made on the basis of intermittent and progressive ptosis. Pyridostigmine bromide (720\u00a0mg/day) was started, but her symptoms did not resolve. The neostigmine experiment and electromyography were normal, so the diagnosis of myasthenia gravis was ultimately ruled out. Brain magnetic resonance imaging (MRI) was unremarkable, but magnetic resonance angiography (MRA) showed a 4-mm aneurysm located at the cavernous segment of the left internal carotid artery (Figs.\u00a0A 63-year-old Chinese woman presented with intermittent left ptosis and diplopia beginning half a year ago Fig.\u00a0. The ptoThe oculomotor nerve nucleus is located in the dorsal midbrain. The oculomotor nerve fascicle tracks through the red nucleus, exits ventrally, passes between the posterior communicating artery and the superior cerebellar artery, lies in a dural fold along the superolateral wall, and heads toward the superior orbital fissure to innervate the extraocular muscles, the iris sphincter muscle, and the elevator palpebrae superioris muscle . Any lesThe Rule of the Pupil states that, when an aneurysm compresses the oculomotor nerve, the iris sphincter will be impaired, and the affected pupil will be dilated or sluggishly reactive to light . Walsh aIntermittent ptosis and diplopia were the only neurological signs in our aneurysmal oculomotor nerve palsy. As Jaeger reported previously, mild ptosis with mild mydriasis was the minimum sign of aneurysm-related oculomotor nerve function . PreviouThe presentation of our case underlines the existence of pupil-sparing incomplete aneurysmal oculomotor palsy. Aneurysm-related oculomotor nerve palsy is a warning sign of potential aneurysm ruptures. Considering the serious consequences of rupture, regardless of whether the pupil is affected in oculomotor nerve palsy, neuroimaging should be performed as soon as possible to rule out intracranial aneurysm."}
+{"text": "Recently, a novel integrated dilator-needle system  was introduced to reduce the number of exchanges for a transseptal access. This system can be used in combination with large bore sheaths. In this pilot study, we evaluated the safety and efficacy of a zero-exchange approach with the AcQCross system in cryoballoon procedures.n\u2009=\u200920; control group: n\u2009=\u200920) who underwent a cryoballoon procedure for the treatment of atrial fibrillation. In the AcQCross and control group, patients underwent ablation with POLARx (Boston Scientific) and Arctic Front Advance Pro  in equal numbers (n\u2009=\u200910). In the AcQCross group, the AcQGuide Max sheath  was used in all POLARx cases.In this retrospective single-center study, we included 40 patients (AcQCross: P\u2009<\u20090.001) and time from puncture until balloon delivery  in comparison with the control group. The balloon in body time, fluoroscopy time, number of cryoapplications, and biophysical parameters were similar between groups. There was one temporary phrenic nerve injury in the AcQCross group. Importantly, no signs of air embolism were noted with the AcQGuide Max sheath.The baseline characteristics of the study population were comparable between groups. In the AcQCross group, there was a reduction in procedure time (49.7\u2009\u00b1\u20099.0\u00a0min vs. 59.6\u2009\u00b1\u20098.1\u00a0min, The use of the novel AcQCross system improves procedural efficacy in cryoballoon procedures by reducing the number of exchanges. Cryoballoon ablation (CBA) has become a well-established approach to achieve pulmonary vein isolation (PVI) for the treatment of atrial fibrillation (AF) . The majDuring cryoballoon procedures, the transseptal access is usually gained through a standard 8F transseptal sheath with the use of a transseptal needle. After placement of the guidewire in the pulmonary vein (PV), the transseptal sheath is replaced by a large bore sheath (15.0 to 15.9F) to accommodate the cryoballoon. Recently, a novel integrated dilator-needle system  became available (US FDA clearance in 2021 and CE mark in 2020). In the AcQCross system, the dilator and transseptal needle form a single component. The lumen of the tapered-tip shaft is fitted with a hollow stainless steel transseptal needle, and both the shaft and needle are connected to the same proximal handle. The AcQCross system allows a 0.032\u2033 guidewire to be loaded during the transseptal puncture (TSP) . This prThe aim of the present pilot study was to evaluate the safety and efficacy of a zero-exchange approach with the AcQCross system in cryoballoon procedures. We hypothesize that the use of AcQCross is safe and results in reduction in the procedure time by a more efficient delivery of the cryoballoon in the left atrium (LA).In this retrospective nonrandomized single-center study, we included 40 patients who underwent a first\u2010time CBA for the treatment of symptomatic paroxysmal or persistent AF. The AcQCross group consisted of the first 10 consecutive patients who underwent ablation with AcQCross and POLARx cryoablation catheter  and the first 10 consecutive patients who underwent ablation with AcQCross and Arctic Front Advance Pro cryoablation catheter . The historical control group consisted of the last 10 consecutive patients undergoing ablation with POLARx and the last 10 consecutive patients undergoing ablation with AFA-Pro before the introduction of AcQCross in our clinical practice.All patients received oral anticoagulation for at least 3\u00a0weeks before ablation. Direct-acting oral anticoagulants were withheld in the morning of the procedure. Vitamin K antagonists continued with a target INR between 2.0 and 2.5. To exclude left atrial thrombi, all patients underwent transesophageal echocardiogram just before the procedure.All procedures were performed under deep sedation by two experienced cryoballoon operators . Femoral vein punctures were performed under ultrasound guidance. After placement of 2 short introducer sheaths (8F and 10F) in the femoral vein, a bolus of intravenous heparin (5000\u00a0IE) was given. In the control group, a 0.032\u2033 guidewire was placed from the femoral access site to the superior vena cava (SVC) Fig.\u00a0. The shoIn the AcQCross group, a 0.032\u2033 guidewire was placed from the femoral access site to the SVC Fig.\u00a0. After rn\u2009=\u20094) and FlexCath Advance steerable sheaths (n\u2009=\u20096) were used. After optimal PV occlusion was achieved, as assessed by contrast injection, cryoablation was started. A TTI-guided ablation protocol was used. The freeze duration was 180\u00a0s if TTI was\u2009<\u200960\u00a0s; otherwise, a 240\u2010s freeze cycle was employed. No bonus freeze was employed routinely. PVI was confirmed by entrance/exit block at the end of the procedure. During cryoablation of the right\u2010sided PVs, high\u2010output right phrenic nerve stimulation was performed using a diagnostic catheter in the right subclavian vein or superior vena cava. Diaphragmatic excursion was assessed by palpation or, in case of the POLARx system, by using the diaphragmatic movement sensor (DMS). The DMS uses an accelerometer and provides a relative measure of the diaphragmatic excursion. Whenever the diaphragmatic excursions decreased or the DMS percentage drops below a cutoff (65%), cryoablation was immediately terminated. During cryoablation of the left\u2010sided PVs, a diagnostic catheter was placed in the right ventricle to provide ventricular pacing in case of a vagal response after cryoablation. The day following the procedure, the groin was inspected for groin hematoma, and a transthoracic echocardiogram was performed to rule out pericardial effusion.Patients underwent PVI using either the POLARx or AFA-Pro cryoablation catheter. Because there is no dedicated AcQCross system for the POLARSHEATH, we used the 15.2F AcQGuide Max sheath in all 10 patients in the AcQCross group who underwent ablation with a POLARx cryoballoon. We deemed the FlexCath Advance sheath too small for the POLARx cryoballoon. In patients undergoing ablation with AFA-Pro in the AcQCross group, both the AcQGuide Max , duration of CBA, and TTI (if measurable). For CBA applications\u2009>\u2009120\u00a0s, we collected the balloon nadir temperature and thawing time until 0\u00a0\u00b0C.The primary efficacy endpoints were procedure time , time from puncture until balloon delivery, balloon in body time, and fluoroscopy time. The primary safety endpoint was defined as a composite of stroke, transient ischemic attack (TIA), air embolism (including transient ST elevation), cardiac tamponade, and myocardial infarction.t-test and Mann\u2013Whitney U\u2010test. Differences between categorical variables were evaluated using the chi-square test or Fisher\u2019s exact test in case of low numbers per cell. Statistical analyses were performed using SPSS (version 28.0.1.0). P-values\u2009<\u20090.05 were considered statistically significant.Continuous data are presented as mean\u2009\u00b1\u2009standard deviation (SD) or median with 25th and 75th percentile, as appropriate. Categorical variables are presented by frequencies and percentages. Differences between continuous variables were tested with the Student\u2019s In total, 40 patients were included in this study. In both groups, there were 10 patients treated with POLARx and 10 patients with AFA-Pro. The baseline characteristics between the AcQCross group and control group were similar (Table P\u2009<\u20090.001) and time from puncture until balloon delivery  in comparison with the control group , there is the option to use RF on the needle by connecting the system to a standard electrosurgery pencil. The AcQCross is the only commercially available transseptal access system which is cleared for both mechanical and RF crossing. It is not clear whether RF on a hollow needle may cause electrocautery tissue coring . AlternaCurrently, there is a specific AcQCross system for the FlexCath Advance sheath but not for the POLARSHEATH. Interestingly, Medtronic has recently (April 2022) acquired the left-heart access portfolio, including the AcQCross Qx system, from Acutus Medical. We used the AcQGuide Max 2.0 sheath in all POLARx cases and in a few AFA-Pro cases. There are small differences in the specifications of the different sheaths Table . The innBesides the AcQCross system, the VersaCross RF system  was recently introduced aiming to reduce the number of exchanges during a TSP . This isAlthough we present an over-the-wire TSP technique, Str\u00f6ker et al. previously demonstrated the safety and efficacy of a direct approach with the FlexCath Advance sheath using an over-the-needle technique . By presThis was a retrospective nonrandomized study with its inherent limitations. However, all study endpoints are collected as part of standard clinical practice. Furthermore, all procedures were performed by experienced operators. We cannot comment on the presence of cerebral micro-embolization because we did not perform intraprocedural transcranial Doppler or post-ablation diffusion-weighted magnetic resonance imaging. It is expected that reduction in sheath exchanges is associated with a reduction of cerebral micro-embolization, but this should be further evaluated in future research. Finally, considering the nonrandomized study design, all results of this study should be interpreted with caution.In this pilot study, we demonstrated that the use of the novel AcQCross system improves procedural efficacy in cryoballoon procedures by reducing the time from puncture until balloon delivery. Importantly, there were no signs of coronary air emboli, TIA, or stroke. Larger international registries are necessary to confirm the safety and efficacy of this novel integrated needle-dilator system. We believe that the AcQCross system may be useful tool to optimize the procedural transseptal workflow during a CBA procedure."}
+{"text": "Prezado editor, Gostar\u00edamos, no entanto, de tecer algumas considera\u00e7\u00f5es relacionadas ao cuidado dirigido ao paciente com acidente vascular cerebral (AVC) agudo. O texto atribui corretamente a Hipertens\u00e3o Arterial como principal causa de AVC isqu\u00eamico (AVCi) e AVC hemorr\u00e1gico (AVCh). Apontaremos, a seguir, algumas considera\u00e7\u00f5es quanto ao manejo da press\u00e3o arterial (PA) nesses pacientes.Inicialmente, a Sociedade Brasileira de Doen\u00e7as Cerebrovasculares (SBDCV) parabeniza o Departamento de Hipertens\u00e3o Arterial da Sociedade Brasileira de Cardiologia, a Sociedade Brasileira de Hipertens\u00e3o e a Sociedade Brasileira de Nefrologia pela realiza\u00e7\u00e3o das Diretrizes Brasileiras de Hipertens\u00e3o Arterial 2020. A eleva\u00e7\u00e3o da PA na fase aguda, resposta fisiol\u00f3gica ao quadro de AVCh, correlaciona-se a um pior progn\u00f3stico e expans\u00e3o do hematoma, dado demonstrado no estudo INTERACT-1. Posteriormente, o estudo de fase 3 INTERACT-2 avaliou o controle intensivo de PA nestes pacientes, com meta Press\u00e3o Arterial Sist\u00f3lica (PAS) <140mmHgversuscontrole de diretrizes da \u00e9poca (meta de PAS <180mmHg). Analisando o desfecho prim\u00e1rio de morte ou depend\u00eancia funcional , n\u00e3o foi alcan\u00e7ada signific\u00e2ncia estat\u00edstica em rela\u00e7\u00e3o \u00e0 redu\u00e7\u00e3o de PA na fase aguda para esse desfecho . O desfecho secund\u00e1rio analisado, padr\u00e3o de distribui\u00e7\u00e3oshiftna mRS, tamb\u00e9m n\u00e3o teve resultados estatisticamente significativos. Entretanto, quando avaliado em an\u00e1lise ordinal, houve menor incapacidade (mRS 0-2) no grupo com tratamento intensivo da PA, comodds ratio0.87 , al\u00e9m de melhor qualidade f\u00edsica e mental, medidos pela escala EQ-5D, no grupo tratado com o controle intensivo da PA. Ao contr\u00e1rio do que a atual Diretriz Brasileira de Hipertens\u00e3o Arterial recomenda, e baseada nos resultados dessa an\u00e1lise ordinal, a recomenda\u00e7\u00e3o atual daAmerican HearteAmerican Stroke Association, endossada pela SBDCV, \u00e9 a redu\u00e7\u00e3o aguda da PAS em pacientes com AVCh que se apresentem com PAS elevada entre 150-220mmHg e que n\u00e3o tenham contraindica\u00e7\u00f5es ao controle intensivo da PA, com alvo PAS <140mmHg, medida que pode ser efetiva na melhora do desfecho cl\u00ednico funcional. Ainda n\u00e3o h\u00e1 dados suficientes para sustentar de maneira sistem\u00e1tica a seguran\u00e7a e a efetividade do manejo agudo de PA em pacientes com PAS >220mmHg, por\u00e9m considerase razo\u00e1vel a redu\u00e7\u00e3o agressiva da PA nesse perfil de pacientes, com infus\u00e3o de medicamentos endovenosos com titula\u00e7\u00e3o de dose e controle rigoroso da PA na fase aguda. Com base nos dados do INTERACT-2 e na recomenda\u00e7\u00e3o das sociedades supracitadas, gostar\u00edamos de sugerir a corre\u00e7\u00e3o dos itens 10.6.1 e 13.7.2 na diretriz em quest\u00e3o, quando se aponta aus\u00eancia de benef\u00edcio em redu\u00e7\u00e3o de incapacidade grave com o controle intensivo da PA. Refor\u00e7amos ainda que, ao contr\u00e1rio do exposto no texto, j\u00e1 foi tamb\u00e9m demonstrado em estudos cl\u00ednicos que a redu\u00e7\u00e3o proposta da PA \u00e9 segura. A SBDCV n\u00e3o recomenda o alvo proposto de PAS <180mmHg para o AVCh apontado na diretriz.Em rela\u00e7\u00e3o ao controle da PA do paciente com AVCh, foi mencionado na diretriz, no item 10.6.1, que \u201cestudos robustos sugerem que reduzir a PA (dentro de 6h) para valores <140/90mmHg n\u00e3o diminui eventos prim\u00e1rios importantes, inclusive mortalidade\u201d, com refer\u00eancia ao estudo INTERACT-2.Em rela\u00e7\u00e3o ao manejo de PA no AVCi agudo, t\u00f3pico 10.6.2, refor\u00e7amos que a redu\u00e7\u00e3o da PA em pacientes candidatos a tromb\u00f3lise dever\u00e1 ser feita quando os valores estiverem >185/110mmHg na primeira hora. Ap\u00f3s o t\u00e9rmino da tromb\u00f3lise, de fato, o valor de PA recomendado \u00e9 <180/105mmHg nas primeiras 24 horas, como apontado na diretriz de Hipertens\u00e3o.Em rela\u00e7\u00e3o ao t\u00f3pico 13.7.1, a PA recomendada para indica\u00e7\u00e3o de tratamento trombol\u00edtico \u00e9 <185/110mmHg, devendo ser iniciado medicamento anti-hipertensivo endovenoso imediatamente, para tentativa de corre\u00e7\u00e3o, caso a medida inicial de PA esteja acima desse patamar. A contraindica\u00e7\u00e3o \u00e0 tromb\u00f3lise ocorre somente se houver PA refratariamente elevada em tr\u00eas medidas consecutivas, com intervalo de 5 minutos entre elas, a despeito de tratamento otimizado. Ao contr\u00e1rio, o AIT \u00e9 considerado um evento equivalente ao AVCi agudo, e deve ser manejado com os mesmos par\u00e2metros do AVCi n\u00e3o trombolisado ou n\u00e3o tratado com trombectomia mec\u00e2nica, ou seja, tolerabilidade de PA at\u00e9 220/120mmHg, usualmente com a suspens\u00e3o de drogas antihipertensivas por via oral na fase hiperaguda do atendimento, e desde que n\u00e3o existam outras condi\u00e7\u00f5es cardiovasculares impeditivas para permitir estes n\u00edveis press\u00f3ricos . Assim, cabe tamb\u00e9m uma revis\u00e3o do t\u00f3pico e quadro sobre as recomenda\u00e7\u00f5es de redu\u00e7\u00e3o imediata da PA no AIT, e de n\u00e3o redu\u00e7\u00e3o da PA em AVCi de uma forma geral (quadro 10.2).Tamb\u00e9m desconhecemos a refer\u00eancia que sugere redu\u00e7\u00e3o imediata da press\u00e3o arterial em pacientes com ataque isqu\u00eamico transit\u00f3rio, conforme sugerido no quadro 10.2. Por este motivo, nos \u00faltimos anos, a grande maioria dos especialistas em Neurologia Vascular e organiza\u00e7\u00f5es de pacientes, em conjunto com a SBDCV e o Departamento Cient\u00edfico de Doen\u00e7as Cerebrovasculares da Academia Brasileira de Neurologia, t\u00eam recomendado e disseminado, tanto no meio acad\u00eamico quanto em materiais e campanhas educativas junto \u00e0 popula\u00e7\u00e3o, comunicados, entrevistas na m\u00eddia e redes sociais, a unifica\u00e7\u00e3o do termo ideal na nossa l\u00edngua como sendo a sigla \u201cAVC\u201d, visando uma maior educa\u00e7\u00e3o da popula\u00e7\u00e3o quanto \u00e0 doen\u00e7a, e evitando-se o uso de outros termos, fator que pode confundir e dificultar o seu reconhecimento r\u00e1pido, primordial ao adequado tratamento e melhor progn\u00f3stico. Assim, sugerimos, em documento futuro da Diretriz de Hipertens\u00e3o, rever a utiliza\u00e7\u00e3o do termo \u201cAVE\u201d, preferindo-se denominar a doen\u00e7a como temos denominado e advogado nos \u00faltimos anos: Acidente Vascular Cerebral \u2013 AVC.\u00c9 sabido que a denomina\u00e7\u00e3o do AVC no nosso pa\u00eds \u00e9 bastante diversa, at\u00e9 mesmo entre m\u00e9dicos, a depender do estado ou regi\u00e3o do pa\u00eds , fato demonstrado inclusive em estudo brasileiro sobre este tema.Por fim, agradecemos a oportunidade desta manifesta\u00e7\u00e3o, e em nome da Sociedade Brasileira de Doen\u00e7as Cerebrovasculares e do Departamento Cient\u00edfico de Doen\u00e7as Cerebrovasculares da Academia Brasileira de Neurologia, nos colocamos \u00e0 disposi\u00e7\u00e3o para futuras colabora\u00e7\u00f5es e discuss\u00f5es de t\u00f3picos que envolvam o manejo de pacientes com AVC. Carta resposta \u00e0 Sociedade Brasileira de Doen\u00e7as Cerebrovasculares e ao Departamento Cient\u00edfico de Doen\u00e7as Cerebrovasculares da Academia Brasileira de Neurologia.e pelos coment\u00e1rios que permeiam o conte\u00fado exposto no cap\u00edtulo 10, mais especificamente ao t\u00f3pico 10.6 que trata da tem\u00e1tica referente \u00e0 hipertens\u00e3o arterial (HA) e acidente vascular encef\u00e1lico (AVE) hemorr\u00e1gico e isqu\u00eamico.Inicialmente manifestamos nosso respeito e agradecimentos pela leitura atenta \u00e0s Diretrizes Brasileiras de Hipertens\u00e3o Arterial 2020 (DBHA2020),contou com a participa\u00e7\u00e3o intelectual coletiva da Sociedade Brasileira de Cardiologia por meio do seu Departamento de Hipertens\u00e3o Arterial, da Sociedade Brasileira de Hipertens\u00e3o e da Sociedade Brasileira de Nefrologia, representadas por 97 especialistas nomeados por crit\u00e9rios cient\u00edficos. Durante todo o ano de 2020, esses profissionais trabalharam com o objetivo comum de construir esse documento, elaborado a partir de reuni\u00f5es semanais do comit\u00ea diretivo, duas reuni\u00f5es com os 18 coordenadores de cap\u00edtulo, e duas reuni\u00f5es plen\u00e1rias com os integrantes da DBHA2020. O texto final representa a opini\u00e3o da maioria desse colegiado.Importante salientar que a DBHA2020Passamos agora a discutir sobre os pontos questionados pelos subscritores da carta ao editor e, para isso, gostar\u00edamos de tecer as seguintes reflex\u00f5es:A quest\u00e3o da meta de tratamento da HA, quando iniciar e quais medicamentos utilizar em eventos cerebrovasculares \u00e9 certamente mat\u00e9ria de grande complexidade e um desafio, mesmo para especialistas.Em janeiro de 2021, uma revis\u00e3o narrativa compilou os principais estudos em AVE isqu\u00eamicos e hemorr\u00e1gicos, apontando para um conservadorismo em rela\u00e7\u00e3o \u00e0 fase aguda desses eventos, e limita\u00e7\u00f5es, instranspon\u00edveis at\u00e9 o presente momento, mesmo quando se utilizam estrat\u00e9gias de revis\u00f5es sistem\u00e1ticas e metan\u00e1lises.que apresenta a seguinte reda\u00e7\u00e3o sobre o tratamento da HA na fase aguda do AVE hemorr\u00e1gico: \u201cSe houver aumento da press\u00e3o arterial, podem ocorrer maior probabilidade de expans\u00e3o do hematoma, aumento do risco de morte e pior progn\u00f3stico. Estudos robustos sugerem que reduzir a press\u00e3o arterial (dentro de 6 h) para valores <140/90 mmHg n\u00e3o diminuiu eventos prim\u00e1rios importantes, inclusive mortalidade. Assim, n\u00e3o deve ser recomendada imediata redu\u00e7\u00e3o da press\u00e3o arterial nos casos de AVE hemorr\u00e1gico, a menos que o valor da press\u00e3o arterial sist\u00f3lica esteja > 220 mmHg\u201d.Refor\u00e7amos que o texto afirma n\u00e3o haver evid\u00eancia de redu\u00e7\u00e3o de eventos prim\u00e1rios, o que est\u00e1 em acordo com a carta encaminhada ao editor pela Sociedade Brasileira de Doen\u00e7as Cerebrovasculares e pelo Departamento Cient\u00edfico de Doen\u00e7as Cerebrovasculares da Academia Brasileira de Neurologia onde est\u00e1 escrito: \u201co estudo de fase 3 INTERACT-2 avaliou o controle intensivo de press\u00e3o arterial nestes pacientes, com meta press\u00e3o arterial sist\u00f3lica <140mmHg versus controle de diretrizes da \u00e9poca . Analisando o desfecho prim\u00e1rio de morte ou depend\u00eancia funcional , n\u00e3o foi alcan\u00e7ada signific\u00e2ncia estat\u00edstica em rela\u00e7\u00e3o \u00e0 redu\u00e7\u00e3o de press\u00e3o arterial na fase aguda para esse desfecho.\u201dSobre o Item 10.6.1 da DBHA2020,A nosso ver, assumir recomenda\u00e7\u00e3o de diretrizes com base em resultados delineados como objetivos secund\u00e1rios, quando o objetivo prim\u00e1rio n\u00e3o foi alcan\u00e7ado, n\u00e3o \u00e9 a orienta\u00e7\u00e3o mais adequada. A proposi\u00e7\u00e3o no Quadro 10.2 para a redu\u00e7\u00e3o da press\u00e3o arterial na fase aguda (dentro de 6 horas) ap\u00f3s um acidente vascular encef\u00e1lico hemorr\u00e1gico atendeu \u00e0 decis\u00e3o da coordena\u00e7\u00e3o geral, coordena\u00e7\u00e3o do cap\u00edtulo e da maioria em reuni\u00e3o plen\u00e1ria como a mais alinhada com as evid\u00eancias cient\u00edficas atuais.Guideline da European Stroke Organization(ESO),e daAmerican Heart Association e American Stroke Association.Quanto \u00e0 orienta\u00e7\u00e3o de conduta na fase aguda do AVE isqu\u00eamico (item 10.6.2), a recomenda\u00e7\u00e3o da DBHA2020 \u00e9 para a redu\u00e7\u00e3o da press\u00e3o arterial sist\u00f3lica a valores menores que 180 mmHg, e da diast\u00f3lica a valores inferiores a 105 mmHg apenas nos pacientes candidatos \u00e0 tromb\u00f3lise, sem evid\u00eancia de benef\u00edcio cl\u00ednico nos demais pacientes (Quadro 10.2). Pela leitura atenta que fizemos da Carta ao Editor, entendemos haver concord\u00e2ncia com essa orienta\u00e7\u00e3o que, coincidentemente, \u00e9 a mesma do rec\u00e9m-publicadoEm rela\u00e7\u00e3o ao coment\u00e1rio sobre a conduta na fase aguda ap\u00f3s acidente isqu\u00eamico transit\u00f3rio, estamos alinhados com as recomenda\u00e7\u00f5es de outras sociedades cient\u00edficas como, por exemplo, a \u00faltima diretriz europeia de hipertens\u00e3o arterial publicada em 2018 que cita na p\u00e1gina 3086 a mesma orienta\u00e7\u00e3o.guidelineda ESO,cujos 11 autores s\u00e3o da \u00e1rea de neurologia, a primeira frase expressa exatamente a possibilidade construtiva de diverg\u00eancias:\u201cThe optimal blood pressure (BP) management in acute ischaemic stroke (AIS) and acute intracerebral haemorrhage (ICH) remains controversial.\u201dConclui o documento apontando para a necessidade de estudos randomizados e controlados que possam corroborar metas, tempo e estrat\u00e9gias para diminui\u00e7\u00e3o da press\u00e3o arterial na fase aguda em diferentes subgrupos de pacientes com AVE.No\u2013 AVE \u2013, essa foi a opini\u00e3o da maioria dos integrantes como sendo o mais adequado a ser empregado no documento atual. Entendemos tratar-se de uma quest\u00e3o sem\u00e2ntica e n\u00e3o anat\u00f4mica propriamente dita e que a doen\u00e7a cerebrovascular pode envolver qualquer estrutura encef\u00e1lica. \u00c9 curioso que o descritor acidente vascular cerebral tenha 17 (dezessete) alternativas de sinon\u00edmia reconhecidas na Biblioteca Virtual de Sa\u00fade (BVS), entre elas, acidente vascular encef\u00e1lico e sua correspondente abreviatura, AVE.No entanto, essa sugest\u00e3o ser\u00e1 considerada na edi\u00e7\u00e3o de uma nova atualiza\u00e7\u00e3o das diretrizes, tendo em vista os argumentos apresentados e a familiaridade existente na classe m\u00e9dica em geral com o termo AVC.Finalmente, sobre a op\u00e7\u00e3o de nomenclatura utilizada nas DBH2020Uma vez mais, agradecemos a oportunidade de debate t\u00e9cnico e intelectual, e esperamos ter justificado os questionamentos encaminhados. Dear EditorOur aim is to raise some issues related to stroke care and BP management in acute setting, an important topic of discussion and controversy. The guideline correctly attributes arterial hypertension as the main cause of ischemic stroke and intracranial hemorrhage. We would like to point out some issues regarding managing blood pressure (BP) in these patients.Initially, the Brazilian Society of Cerebrovascular Diseases (SBDCV) congratulates the Department of Arterial Hypertension of the Brazilian Societies of Cardiology, Hypertension, and Nephrology for the Brazilian Guidelines on Arterial Hypertension 2020 publication.The physiological response of increased BP levels in ICH is correlated with worse prognosis and hematoma expansion, as demonstrated in the INTERACT-1 study.Subsequently, the INTERACT-2 trial compared intensive BP control [target systolic blood pressure (SBP) <140 mmHg] versus the guidelines-recommended levels (SBP <180 mmHg) in acute setting, and the primary outcome of death or functional dependence  were similar .Further, there was no significant difference in the shift distribution pattern in mRS. However, the ordinal analysis revealed a lower disability (mRS 0-2) with intensive BP treatment, with an odds ratio of 0.87 , and additional better physical and mental quality, measured by EQ-5D scale.Contrary to the 2020 Brazilian Arterial Hypertension Guidelines recommendation, and based on the results of this ordinal analysis, the current recommendation of the American Heart and American Stroke Association,endorsed by the SBDCV, is to achieve an acute reduction of SBP in patients with ICH who present with high SBP (150-220 mmHg) without contraindications for intensive BP control. The SBP target is <140 mmHg, which can improve functional clinical outcomes. There is no sufficient data to systematically support the safety and effectiveness of the acute management of BP in patients with SBP >220 mmHg; however, a more aggressive reduction of BP in this profile of patients is reasonable, using intravenous drugs, dose titration and strict BP control in the acute phase.Based on this trial and the recommendation of the societies mentioned above, we would like to suggest the correction of items 10.6.1 and 13.7 of the brazilian hypertension guideline, that states no benefit in reducing severe disability with intensive BP control.We also emphasize that the proposed reduction in BP is safe.The SBDCV does not recommend the proposed target of SBP <180 mmHg for acute ICH management.Regarding BP control in patients with intracranial hemorrhage (ICH), the guidelines mentioned that \u201crobust studies suggest that reducing BP (within 6h) to values <140/90 mmHg does not decrease important primary events, including mortality\u201d (item 10.6.1), according to INTERACT-2 study.Regarding the management of BP in acute ischemic stroke (IS), topic 10.6.2, we emphasize that BP reduction in patients who are candidates for thrombolysis should be performed when the values are >185/110 mm Hg in the first hour. After the end of thrombolysis, the recommended BP value is <180/105 mmHg in the first 24 h, as indicated in the hypertension guideline.Regarding topic 13.7.1, the recommended BP for the indication of thrombolytic treatment is <185/110 mmHg, and intravenous antihypertensive medication should be started immediately above this level. A contraindication to thrombolysis occurs only if refractory elevated BP occurs in three consecutive measurements, with an interval of 5 min, despite optimized treatment.In contrast, TIA is considered an equivalent to acute IS, and must be managed with the same parameters of a non-thrombolysed IS, or a IS not submitted to thrombectomy, that is, BP tolerability up to 220/120mmHg and suspension of oral antihypertensive drugs in the hyperacute phase of care, unless there are other impeding cardiovascular conditions to allow these blood pressure levels .Thus, it is also worth reviewing the topic and table of recommendations for immediate BP reduction in TIA, as well as no reduction of BP in all types of IS (Table 10.2).We are also unaware of the reference that suggests immediate BP reduction in patients with a transient ischemic attack (TIA), as suggested in Table 10.2.For this reason, recently, the portuguese term \u201cAVC\u201d have been widely recommended by specialists in the field, patient organizations, together with the SBDCV and the Scientific Department of Cerebrovascular Diseases of the Brazilian Academy of Neurology, as well as in academic research, campaigns, educational activities, press releases and interviews, with the purpose of better educate the population regarding the disease, and to avoid using other terms that may confuse and hinder the rapid recognition, essential for immediate stroke treatment and better prognosis. Thus, for future hypertension guidelines, we suggest the use of this recommended standardized portuguese terminology: \u201cAVC - Acidente Vascular Cerebral\u201d, instead of other terms, like \u201cAVE\u201d.The denomination of stroke in our country is quite diverse, depending on the state or region of the country , a fact that has been demonstrated in a Brazilian study.Finally, we are grateful for the opportunity to present our observations. On behalf of the Brazilian Society of Cerebrovascular Diseases and the Scientific Department of Cerebrovascular Diseases of the Brazilian Academy of Neurology, our goal as a stroke society is to help and contribute with other partners in future discussions of topics involving managing patients with stroke. Response to the Brazilian Society for Cerebrovascular Diseases and the Scientific Department on Cerebrovascular Diseases of the Brazilian Academy of Neurology.and for the comments on Chapter 10, more specifically on the topic 10.6 that addresses arterial hypertension (AH), and hemorrhagic and ischemic stroke.First, we would like to express our respect and gratitude to the reviewers for the careful reading of the 2020 Brazilian Guidelines on Arterial Hypertension (DBHA2020)was developed by the collective effort of the Brazilian Society of Cardiology through the Department of Arterial Hypertension, the Brazilian Society of Hypertension, and the Brazilian Society of Nephrology, represented by 97 specialists chosen by scientific criteria. During the entire year of 2020, these experts worked together to construct this document, which was achieved by weekly meetings of the directive committee, two meetings with 18 chapter coordinators, and two plenary sessions of all members involved in the elaboration of the DBHA2020. The final text represents the opinion of the majority of this work group.It is worth pointing out that the DBHA2020We will now discuss the questions raised by the authors of the Letter to the Editor, by first making the following considerations:The question about the treatment target for HA, the time when treatment should be initiated, and the drugs of choice in cerebrovascular events is certainly a complex and challenging issue.In January 2021, a narrative review summarized the main studies on ischemic and hemorrhagic stroke and pointed to a conservative approach in the acute phase, in addition to associated limitations that cannot be overcome even when strategies proposed by systematic reviews and meta-analyses are used.which describes recommendations on the treatment of AH in the acute phase of hemorrhagic stroke: \u201cIn case of an increase in blood pressure, the odds of hematoma expansion, death and worse prognosis may increase. Robust studies have suggested that the reduction of blood pressure (within six hours) to levels < 140/90mmHg does not reduce important primary events, including mortality. Therefore, the immediate reduction of blood pressure in case of hemorrhagic stroke is not recommended, unless systolic blood pressure is > 220 mmHg \u201d. We reinforce that the text states that there is no evidence of reduction of primary events, which is in accordance with the Letter to the Editor submitted by the Brazilian Society of Cerebrovascular Diseases and by the Scientific Department of Cerebrovascular Diseases of the Brazilian Academy of Neurology, which affirms: \u201cthe phase 3 INTERACT-2 study evaluated the strict control of blood pressure in these patients, with a target systolic blood pressure below 140 mmHgversusthe target recommended by guidelines (<180mmHg). Considering the primary outcome of death or functional dependence , no statistically significance was found in this outcome by reducing blood pressure levels in the acute phase\u201d.Regarding the item 10.6.1 of the DBHA2020,In our opinion, it is not appropriate to adopt guidelines\u2019 recommendations based on results obtained from secondary objectives, when the primary objective was not achieved. In Chart 10.2, the suggestion of reducing blood pressure (within six hours) in the acute phase after a hemorrhagic stroke was in line with the decision of the general coordination, chapter coordination and the plenary session, that this approach was in accordance with current scientific evidence.and of the American Heart Association and American Stroke Association.As for the acute phase of ischemic stroke (item 10.6.2), the recommendation of the DBHA2020 is to reduce systolic blood pressure to values below 180 mmHg and diastolic blood pressure to values below 105 mmHg only for patients candidate for thrombolysis, with no evidence of clinical benefit for other patients. (Chart 10.2). Based on the close reading of the Letter to the Editor, we understand that there is a consensus on this recommendation, which coincidently is the same of the recently published European Stroke Organization (ESO), guidelinesIn response to the comment on the management in the acute phase following a transient ischemic attack, we are in line with recommendations of other scientific societies, like the last European guidelines on arterial hypertension published in 2018 which recommends the same approach (see page 3086).whose 11 authors work in the field of Neurology, the first sentence expresses the constructive possibility of divergence: \u201cThe optimal blood pressure (BP) management in acute ischemic stroke (AIS) and acute intracerebral hemorrhage (ICH) remains controversial\u201d. The document concludes that further randomized controlled studies are needed to support treatment targets, time and strategies to reduce blood pressure levels in the acute phase in different subgroups of patients with stroke.In the ESO guidelines,\u2013 stroke \u2013 it was considered by most specialists as the most appropriate to be used in the current document. We understand that this is a semantic rather than an anatomic issue, and that cerebrovascular disease may involve any encephalic structure. Curiously, the descriptor brain vascular accident has 28 alternative synonyms, including its abbreviation, BVA, recognized by the virtual health library.However, this suggestion should be further discussed in a new update of the guideline, because of the arguments presented, and the familiarity with the term stroke by physicians in general.Finally, concerning the nomenclature used in the DBH2020Once again, we are grateful for the opportunity to have this technical and intellectual debate, and we hope that we have clarified the questions raised."}
+{"text": "O tratamento adequado e a obten\u00e7\u00e3o das metas na hipertens\u00e3o arterial s\u00e3o importantes na redu\u00e7\u00e3o dos desfechos cardiovasculares. Descrever os bloqueadores do receptor de angiotensina (BRA) em monoterapia ou combina\u00e7\u00e3o dupla e a taxa de controle da hipertens\u00e3o arterial.t pareado, qui-quadrado e Fisher, al\u00e9m de sobreposi\u00e7\u00e3o dos intervalos de confian\u00e7a de 95% com n\u00edvel de signific\u00e2ncia de 5% . Estudo transversal que avaliou pacientes em uso de BRA entre 2017 e 2020. Foram exclu\u00eddos aqueles em uso de tr\u00eas ou mais anti-hipertensivos. As vari\u00e1veis analisadas foram: sexo, idade, \u00edndice de massa corporal, medidas v\u00e1lidas da medida residencial da press\u00e3o arterial (MRPA); press\u00e3o arterial sist\u00f3lica (PAS) e diast\u00f3lica (PAD) obtidas pela MRPA e de forma casual; variabilidade press\u00f3rica; classe dos anti-hipertensivos e dos BRAs. Foram utilizados testes de  Foram selecionados 17.013 pacientes; destes, 12.813 preencheram os crit\u00e9rios, dos quais 62,1% eram do sexo feminino. O n\u00famero m\u00e9dio de medidas v\u00e1lidas foi de 23,3 , com m\u00e9dias para a PAS de 126,8\u00b115,8 mmHg e 133,5\u00b120,1 mmHg  e para a PAD de 79,1\u00b19,7 mmHg e 83,6\u00b111,9 mmHg  pela MRPA e medida casual, respectivamente. Losartana foi o BRA mais utilizado e o que apresentou comportamentos mais elevados da press\u00e3o arterial. As combina\u00e7\u00f5es de BRA com diur\u00e9ticos ou com antagonistas de canal de c\u00e1lcio tiveram menores valores de press\u00e3o arterial. Losartana foi utilizada em mais da metade dos pacientes, apesar de ser a menos eficiente na redu\u00e7\u00e3o e no controle da press\u00e3o arterial. Al\u00e9m disso, sabe-se que pequenas redu\u00e7\u00f5es da PA, mesmo nas fases iniciais da HA, s\u00e3o capazes de promover redu\u00e7\u00e3o nos principais desfechos cardiovasculares.5O tratamento e controle adequado da hipertens\u00e3o arterial (HA) ainda hoje \u00e9 um dos grandes desafios no tratamento dessa doen\u00e7a, que \u00e9 a principal causa de morte em todo o mundo. A ado\u00e7\u00e3o de estrat\u00e9gias de tratamento alinhadas com as evid\u00eancias cient\u00edficas mais atuais \u00e9 um dos caminhos para otimizar esses resultados.6Por outro lado, apesar de todas essas evid\u00eancias, encontramos na cesta b\u00e1sica de medicamentos ofertados pelo Sistema \u00danico de Sa\u00fade (SUS) f\u00e1rmacos de meia-vida curta, em monoterapia e com a necessidade de v\u00e1rias tomadas ao dia; caracter\u00edsticas que podem impactar negativamente na ades\u00e3o e dificultar o controle adequado da PA. Destaca-se que a realidade do SUS reflete o contexto das estrat\u00e9gias medicamentosas adotadas em nosso pa\u00eds para 75% dos pacientes hipertensos.7Artigo publicado em 2021 que avaliou uma base de dados de 22.446 indiv\u00edduos submetidos a medida da PA no consult\u00f3rio e no domic\u00edlio, dos quais 11.337 eram hipertensos tratados por cardiologistas com f\u00e1rmacos anti-hipertensivos, identificou que, em 74,6% dos casos, o bloqueio do sistema renina-angiotensina-aldosterona foi a estrat\u00e9gia adotada, sendo que o uso dos bloqueadores dos receptores de angiotensina (BRAs) ocorreu em 58,7%, seja em monoterapia ou em combina\u00e7\u00e3o.Foram objetivos do presente estudo: (i) verificar a distribui\u00e7\u00e3o da prescri\u00e7\u00e3o dos BRAs em monoterapia e combina\u00e7\u00e3o total e por sexo, regi\u00e3o geogr\u00e1fica e presen\u00e7a de diabetes; (ii) comparar a taxa de controle da PA segundo a medida casual e a medida residencial da PA (MRPA) para todas as estrat\u00e9gias de tratamento medicamentoso com BRA; (iii) comparar o controle da PA entre a MRPA e a medida casual; e (iv) comparar as m\u00e9dias de press\u00e3o arterial sist\u00f3lica (PAS), press\u00e3o arterial diast\u00f3lica (PAD), press\u00e3o de pulso (PP) e variabilidade da PA obtidas com utiliza\u00e7\u00e3o de BRAs em monoterapia ou em combina\u00e7\u00e3o dupla, considerando a classe como um todo e os v\u00e1rios tipos de medicamentos que a comp\u00f5em.Este estudo foi aprovado pelo Comit\u00ea de \u00c9tica em Pesquisa Humana do Hospital das Cl\u00ednicas da Universidade Federal de Goi\u00e1s sob CAE n\u00famero 99691018.7.0000.5078 e avaliou pacientes que realizaram exames na plataforma TeleMRPA (www.telemrpa.com) de maio de 2017 at\u00e9 outubro de 2020.software e sim de uma plataforma acess\u00edvel em qualquer terminal de computador, tablet ou smartphone , a inser\u00e7\u00e3o dos dados relativos \u00e0 medida da PA pode ser feita de forma remota e simplificada.8A plataforma foi desenvolvida como ferramenta de laudo a dist\u00e2ncia por telemonitoramento, com caracter\u00edsticas que permitem a an\u00e1lise e o filtro do banco de dados de acordo com as perguntas cient\u00edficas que se pretende investigar. O algoritmo matem\u00e1tico utilizado possibilita a an\u00e1lise com prote\u00e7\u00e3o dos dados pessoais do paciente, assim como das cl\u00ednicas ou unidades de sa\u00fade, seja para a interpreta\u00e7\u00e3o do exame, seja para a constru\u00e7\u00e3o de projetos de pesquisa. Por n\u00e3o se tratar de um A base de dados analisada se restringiu aos pacientes que estavam em uso de BRAs. Foram inclu\u00eddos pacientes com idade igual ou superior a 18 anos e em uso de monoterapia ou combina\u00e7\u00e3o dupla. Foram exclu\u00eddos pacientes em uso de combina\u00e7\u00e3o de tr\u00eas ou mais anti-hipertensivos ou em combina\u00e7\u00e3o com inibidores da enzima conversora de angiotensina ou combinados com anti-hipertensivos de uso pouco frequente nas combina\u00e7\u00f5es duplas    . Em relForam utilizados os seguintes dados da plataforma TeleMRPA: sexo (masculino/feminino); idade ; \u00edndice de massa corporal (IMC); n\u00famero de medidas v\u00e1lidas da MRPA; PAS e PAD obtidas pela MRPA e de forma casual; variabilidade press\u00f3rica pela MRPA obtida a partir do desvio padr\u00e3o das 24 medidas domiciliares durante o protocolo; classe dos medicamentos utilizados; e os f\u00e1rmacos da classe dos BRAs.Tamb\u00e9m foi avaliada a distribui\u00e7\u00e3o da amostra por regi\u00f5es geogr\u00e1ficas do Brasil, assim como a preval\u00eancia de indiv\u00edduos em uso de medicamentos para o tratamento de diabetes melito (antidiab\u00e9ticos orais e/ou insulina).9 Para a medida da MRPA, o aparelho foi disponibilizado para o paciente, que foi orientado sobre o manuseio e a t\u00e9cnica adequados para a medida da PA no dia da entrega do aparelho.1 Ainda nesse primeiro dia, foram realizadas duas medidas no ambiente da cl\u00ednica/consult\u00f3rio e, nos 4 dias subsequentes, o paciente (e/ou cuidador/acompanhante) realizou as medidas em seu domic\u00edlio, conforme o protocolo. Considerou-se como medida casual a m\u00e9dia das duas medidas do primeiro dia, e como medida domiciliar a m\u00e9dia das 24 medidas do segundo ao quinto dia.10Para o c\u00e1lculo do IMC, foram utilizados o peso e a altura aferidos e a f\u00f3rmula de Quetelet.Foram utilizados aparelhos autom\u00e1ticos validados das marcas Omron, Geratherm e Microlife.1Os dados foram exportados da plataforma TeleMRPA para o Excel. Todas as classes de medicamentos descritas na plataforma foram revisadas e codificadas por duas equipes de trabalho; em seguida, os bancos de dados foram cruzados para identifica\u00e7\u00e3o de dados discrepantes, que, quando presentes, foram revisados com toda a equipe e a coordena\u00e7\u00e3o. Foram considerados com PA controlada os indiv\u00edduos com a PA casual menor que 140 mmHg e 90 mmHg e, pela MRPA, com valores menores que 130 mmHg e 80 mmHg para a PAS e PAD, respectivamente.software Stata, vers\u00e3o 14.0. As vari\u00e1veis quantitativas foram apresentadas com m\u00e9dia e desvio padr\u00e3o, e as qualitativas com frequ\u00eancia absoluta e relativa. Para a verifica\u00e7\u00e3o da normalidade dos dados, utilizou-se o teste de Kolmogorov-Smirnov.A an\u00e1lise estat\u00edstica foi realizada utilizando o Nas compara\u00e7\u00f5es das m\u00e9dias de PAS, PAD e PP obtidas na MRPA e na medi\u00e7\u00e3o casual, utilizou-se o teste t de Student pareado, e os testes qui-quadrado ou de Fisher foram utilizados para comparar as taxas de controle obtidas por meio da medi\u00e7\u00e3o casual com aquelas identificadas pela MRPA, e tamb\u00e9m para comparar as taxas de controle e de n\u00e3o controle da PA segundo a utiliza\u00e7\u00e3o ou n\u00e3o de cada estrat\u00e9gia medicamentosa, considerando a medida casual e a MRPA.A sobreposi\u00e7\u00e3o dos intervalos de confian\u00e7a de 95% foi utilizada para comparar as diferen\u00e7as entre as m\u00e9dias de PAS, PAD, PP e a variabilidade da PA obtidas com a utiliza\u00e7\u00e3o de BRAs em monoterapia ou em combina\u00e7\u00e3o dupla, considerando a classe como um todo e os v\u00e1rios tipos de medicamentos que a comp\u00f5em. Foi adotado um n\u00edvel de signific\u00e2ncia de 5% .Foram avaliados 12.813 pacientes, a maioria do sexo feminino, e quase a metade da regi\u00e3o Nordeste. A preval\u00eancia de diabetes foi de 6,2%   .A estrat\u00e9gia de tratamento encontrada na amostra se dividiu em 48,5% dos pacientes em uso de monoterapia e 51,2% com combina\u00e7\u00f5es duplas. Os f\u00e1rmacos que compuseram a classe dos BRAs apresentaram a seguinte distribui\u00e7\u00e3o: 57,2%, losartana; 18,8%, olmesartana; 15,0%, valsartana; 4,8%, telmisartana; 3,8%, candesartana; e 0,4%, irbesartana.O n\u00famero m\u00e9dio de medidas v\u00e1lidas da MRPA foi de 23,3 . As diferen\u00e7as nos valores m\u00e9dios entre PA casual e MRPA para a PAS e PAD foram de 6,7 mmHg  e 4,5 mmHg , respectivamente. Essas diferen\u00e7as caracterizam o efeito do avental branco e se mant\u00eam em todas as estrat\u00e9gias de tratamento. Esse comportamento se repete com todos os BRAs, seja em monoterapia ou combina\u00e7\u00e3o. Tamb\u00e9m avaliamos e comparamos o percentual de controle pela medida casual e pela MRPA em monoterapia e por estrat\u00e9gia de combina\u00e7\u00e3o   .Na 1 a taxa de controle da PA na amostra total foi melhor quando obtida pela medida casual.Quando avaliamos o controle da PA de acordo com as metas menores que 140 mmHg e 90 mmHg para a medida casual e 130 mmHg e 80 mmHg para a MRPA, conforme diretrizes vigentes,O controle da PA, considerando a MRPA, foi menor naqueles que utilizavam BRA em monoterapia e BRA com BB. Quando analisamos os tipos de BRA utilizados em monoterapia ou em combina\u00e7\u00e3o, o controle da PA foi menor considerando a MRPA naqueles que utilizavam losartana e maior nos que estavam em uso de BRA de meia-vida longa. Essa mesma an\u00e1lise para a medida casual em rela\u00e7\u00e3o ao controle da PA repete essa tend\u00eancia.As taxas de controle dos diversos tipos de BRA combinados com ACC, BB ou DIU, tanto pela MRPA quanto pela medida casual, foram menores nas combina\u00e7\u00f5es com a losartana e maiores com BRAs de meia-vida longa.A m\u00e9dia da PAS pela MRPA com utiliza\u00e7\u00e3o de BRA + ACC e BRA + DIU foi menor do que aquela com BRA em monoterapia. Quando verificado o tipo de BRA utilizado em monoterapia, os valores de PA s\u00e3o progressivamente maiores com olmesartana, candesartana, telmisartana, valsartana e losartana   . Em relAs m\u00e9dias de PAD foram maiores com a utiliza\u00e7\u00e3o de BRA como monoterapia quando comparadas a qualquer combina\u00e7\u00e3o dupla. Quanto ao tipo de BRA, em monoterapia os maiores valores m\u00e9dios de PAD pela MRPA foram com losartana   . QuandoA PP foi maior na utiliza\u00e7\u00e3o de BRA + BB quando comparada \u00e0s outras combina\u00e7\u00f5es e \u00e0 BRA em monoterapia. A losartana em monoterapia ou em combina\u00e7\u00e3o dupla apresentou maior m\u00e9dia de PP que a candesartana e a telmisartana.A variabilidade da PA foi maior com uso de BRA + ACC comparado com as combina\u00e7\u00f5es com DIU e BB e monoterapia. A variabilidade foi menor com a telmisartana, em monoterapia ou combina\u00e7\u00e3o, quando comparado a valsartana. A losartana com ACC apresentou menor m\u00e9dia de variabilidade quando comparada a outras combina\u00e7\u00f5es. A candesartana com BB apresentou maior variabilidade que a candesartana com ACC. N\u00e3o houve diferen\u00e7a da variabilidade entre as diversas combina\u00e7\u00f5es com valsartana, olmesartana e telmisartana.7 Faz sentido, portanto, avaliar aspectos relacionados ao comportamento da PA, tanto no consult\u00f3rio quanto no domic\u00edlio do paciente, com os diversos f\u00e1rmacos que comp\u00f5em a classe dos BRAs.O presente estudo apresenta uma evolu\u00e7\u00e3o da an\u00e1lise publicada em 2020 que demonstrou, em hipertensos tratados com monoterapia ou combina\u00e7\u00e3o dupla, m\u00e9dias de PAS e PAD significativamente mais baixas pela MRPA em compara\u00e7\u00e3o \u00e0 medida casual e encontrou, nos BRAs, a op\u00e7\u00e3o mais utilizada no tratamento.13A amostra estudada foi constitu\u00edda por uma popula\u00e7\u00e3o de idade m\u00e9dia pr\u00f3xima aos 60 anos e com IMC aumentado; al\u00e9m disso, h\u00e1 predom\u00ednio do sexo feminino. Com rela\u00e7\u00e3o \u00e0 distribui\u00e7\u00e3o da amostra pelas regi\u00f5es geogr\u00e1ficas, h\u00e1 predom\u00ednio das regi\u00f5es Nordeste e Sudeste. \u00c9 importante considerar que os fatores idade e excesso de peso aqui encontrados podem causar maior dificuldade na obten\u00e7\u00e3o das metas preconizadas para o tratamento da HA.16 Essa mudan\u00e7a justifica a diferen\u00e7a encontrada nas taxas de controle pela medida casual e MRPA encontradas nesta an\u00e1lise em rela\u00e7\u00e3o ao artigo publicado previamente.7Cabe ressaltar que ocorreu no \u00faltimo ano, fruto de evid\u00eancias publicadas com base de dados nacional sobre a MRPA, ajuste no valor de refer\u00eancia para a normalidade de 135x85 mmHg para 130x80 mmHg.3 As combina\u00e7\u00f5es duplas com DIUs e ACCs foram preferidas, o que est\u00e1 bem alinhado com as recomenda\u00e7\u00f5es atuais.19Em rela\u00e7\u00e3o \u00e0 estrat\u00e9gia de tratamento adotada nesta amostra, os BRAs foram usados em monoterapia em 48,5% dos casos; combinados com DIUs em 23,4%; com ACCs em 16,8%; e com BBs em 11,2%. Interessante notar que, a despeito de recomenda\u00e7\u00f5es un\u00e2nimes das diretrizes de hipertens\u00e3o para a combina\u00e7\u00e3o dos f\u00e1rmacos na maioria dos cen\u00e1rios da doen\u00e7a hipertensiva, a monoterapia ainda permanece muito frequente.22Outro aspecto relevante para a escolha de f\u00e1rmacos no tratamento da HA \u00e9 que tenham preferencialmente meia-vida longa e que permitam uma \u00fanica tomada ao dia, pois essas caracter\u00edsticas interferem diretamente na ades\u00e3o ao tratamento e no controle adequado da PA. Para os f\u00e1rmacos de meia-vida curta, h\u00e1 que se respeitar as caracter\u00edsticas farmacodin\u00e2micas e prescrever a tomada duas ou mais vezes ao dia para que o n\u00edvel plasm\u00e1tico e a efic\u00e1cia na redu\u00e7\u00e3o dos n\u00edveis tensionais sejam mantidos.23\u00c9 interessante relembrar que existem diferen\u00e7as importantes, do ponto de vista farmacol\u00f3gico, entre esses f\u00e1rmacos, pois esses aspectos podem estar relacionados \u00e0s diferen\u00e7as que encontramos na an\u00e1lise de comportamento da PA, visto que os BRAs t\u00eam meias-vidas diferentes: losartana 2 h, valsartana 6 h, candesartana 9 h, olmesartana 12h e telmisartana 24 h.Quando avaliamos a taxa de controle da PA pela medida casual e pela MRPA, encontramos 56,3% e 44,5% dentro das metas, respectivamente, e, ao analisarmos o comportamento da PA com os diferentes BRAs e tamb\u00e9m com as diferentes combina\u00e7\u00f5es duplas, encontramos diferen\u00e7as no percentual de pacientes controlados.Para uma an\u00e1lise mais refinada desse comportamento, realizamos (pela MRPA) uma avalia\u00e7\u00e3o das m\u00e9dias e do intervalo de confian\u00e7a de PAS e PAD, assim como da variabilidade press\u00f3rica. Em rela\u00e7\u00e3o \u00e0 estrat\u00e9gia de combina\u00e7\u00e3o dos BRAs com outras classes de anti-hipertensivos, a combina\u00e7\u00e3o com BB apresentou valores mais altos para a m\u00e9dia e variabilidade da PAS em compara\u00e7\u00e3o \u00e0 combina\u00e7\u00e3o com DIU ou ACC. Entre os diferentes tipos de BRA em monoterapia, a losartana apresenta os valores m\u00e9dios de PAS e PAD mais elevados em rela\u00e7\u00e3o aos BRA de meia-vida mais longa.S\u00e3o limita\u00e7\u00f5es desta an\u00e1lise o fato de estarmos lidando com um estudo observacional, sem o detalhamento das doses empregadas em cada f\u00e1rmaco, e que tamb\u00e9m n\u00e3o \u00e9 representativo da popula\u00e7\u00e3o brasileira. Por outro lado, trata-se de uma grande base de dados, e o estudo reflete as estrat\u00e9gias relacionadas ao uso dos BRA em pacientes hipertensos e permite obter par\u00e2metros importantes em rela\u00e7\u00e3o ao comportamento da PA com os diferentes f\u00e1rmacos em monoterapia e em combina\u00e7\u00e3o.28 e, ainda mais importante, refletem a necessidade de revisarmos a estrat\u00e9gia dos f\u00e1rmacos anti-hipertensivos disponibilizados na cesta b\u00e1sica do SUS6 e prescritos para o tratamento da doen\u00e7a hipertensiva, pois, sabidamente, pequenas diferen\u00e7as na redu\u00e7\u00e3o da PA em hipertensos t\u00eam repercuss\u00f5es importantes na morbimortalidade cardiovascular.Esses achados s\u00e3o compat\u00edveis com estudos randomizados previamente publicados que avaliaram a pot\u00eancia anti-hipertensiva dos diferentes BRAsEm hipertensos tratados com BRAs, a estrat\u00e9gia de monoterapia ainda \u00e9 frequente e, quando combinados, a op\u00e7\u00e3o pelos DIUs e pelos ACCs \u00e9 a preferida.Dentre os BRAs, a losartana em monoterapia e combina\u00e7\u00e3o dupla ainda \u00e9 utilizada em mais da metade dos pacientes, apesar de ser a menos eficiente na redu\u00e7\u00e3o e no controle da PA.Existem claras diferen\u00e7as na meia-vida entre os f\u00e1rmacos que comp\u00f5em a classe dos BRAs, percebidas no comportamento da PA avaliado tanto pela medida casual quanto pela MRPA. Essas diferen\u00e7as podem refletir na efetividade do controle press\u00f3rico.Adriana Siqueira Serpa de Menezes, SAVE, Recife, PE. Andr\u00e9a Ara\u00fajo Brand\u00e3o, Universidade do Estado do Rio de Janeiro, RJ. Anibal Prata Barbosa, Prog de Hip. Arterial Secretaria de Sa\u00fade de Duque de Caxias, RJ. Antonio Almeida Braga, PROCAPE, UPE, Recife, PE. Antonio Eduardo de Melo Filho, Cl\u00ednica de Sa\u00fade Dr Antonio Eduardo de Melo, Triunfo, PE. \u00c1tila de Oliveira Melo, Liga de Hipertens\u00e3o Arterial UFG, Goi\u00e2nia, GO. Andr\u00e9 K Vidigal de Vasconcellos, Instituto de Cardiologia do Agreste, Caruaru, PE. Audes D. M. Feitosa, Unidade de Hipertens\u00e3o e Cardiologia Preventiva, PROCAPE/UPE, Recife, PE. Breno Gontijo de Camargos, AngioCor, Taguatinga, DF. Bruno Alencar Fonseca, Cl\u00ednica Blues, Belo Horizonte, MG. Bruno Daniel Ferrari, Funda\u00e7\u00e3o Educacional do Munic\u00edpio de Assis, FEMA, SP. Bruno Jos\u00e9 Peixoto Coutinho, Cardiologia Hospital Oswaldo Cruz, Universidade de Pernambuco, PE. Carlo Bonasso, Cl\u00ednica M\u00e9dica Carlo Bonasso SS Ltda, S\u00e3o Paulo, SP. Carlos Jos\u00e9 Mota de Lima, Centro Cardiol\u00f3gico S\u00e3o Camilo, CE. Carlos Filinto de Almeida, Instituto do Cora\u00e7\u00e3o de Mato Grosso do Sul, Campo Grande, MS. Claudinelli Alvarenga Aguilar, Cl\u00ednica do Esporte, Goi\u00e2nia, GO. C\u00e9sar Ricardo Soares Medeiros, Cl\u00ednica de Cardiologia Dr C\u00e9sar Medeiros, Ribeir\u00e3o Preto, SP. Cristiano Pederneiras Jaeger, Instituto de Medicina Vascular - Coracentro, Porto Alegre/RS. Daniel Lages Dias, Novacordis, Paul\u00ednia, SP. Diogo da Silva Amorim, Liga de Hipertens\u00e3o Arterial UFG, Goi\u00e2nia, GO. Ednaldo M. Fontes Segundo, Cardiologista pela SBC, Instituto Paulo Gomes (IPG) em Est\u00e2ncia, SE. Eduardo C. D. Barbosa, Dept Hipertens\u00e3o e Cardiometabolismo Hospital S\u00e3o Francisco, Santa Casa Porto Alegre, RS. Eduardo \u00c9rico Zen, Hospital Cardiol\u00f3gico Costantini, Curitiba, PR. Elder Gil A. Cruz, Cl\u00ednica do Cora\u00e7\u00e3o Dr. Elder Gil, Salgueiro, PE. Esther G. Di\u00f4go de Lima de B. Carvalho, Cl\u00ednica S\u00e3o Lucas, Guarabira, PB. F\u00e1bio Argenta, Mediodonto, Cuiab\u00e1, MT. Fabiano de Souza Ramos, MEDCOR Cardiologia, Nova Igua\u00e7u- RJ. Fl\u00e1via Karina Silva e Oliveira, Centro de Cardiologia, S\u00e3o Jos\u00e9 dos Campos, SP. Fl\u00e1vio H. A. P. V\u00e9ras, Cl\u00ednica do Cora\u00e7\u00e3o, Mossor\u00f3, RN. Francisco Deoclecio Pinheiro, Cl\u00ednica de Especialidades M\u00e9dicas de Itapipoca, Itapipoca, CE. Frank Land L. de Carvalho, Cardiovasf, Petrolina, PE. Germano Granja, Cl\u00ednica do Cora\u00e7\u00e3o, Ouricuri, PE. Giovanni Saraiva, Imedi e Icordis, Recife, PE. Gleidson Junio Oliveira de Souza, Liga de Hipertens\u00e3o Arterial UFG, Goi\u00e2nia, GO. Gustavo Barros - MCOR / Recife-PE. Gustavo Guimar\u00e3es Moreira de Castro, ITACORDIS e Universidade Igua\u00e7u \u2013 UNIG, Itabora\u00ed e Nova Igua\u00e7u, RJ. Jadil Francisco Fusturath J\u00fanior, Cardio Service, Porto Velho, RO. Jos\u00e9 Wladimir Tambelli Pires, Cl\u00ednica de Cardiologia, Itapetininga/SP. Jo\u00e3o Evaristo de Oliveira Dantas, Cardiomed/Multimed, S\u00e3o Lu\u00eds, MA. Jo\u00e3o F\u00e9lix de Morais Filho, Cl\u00ednica Angioc\u00e1rdio, Natal / RN. Jo\u00e3o Francisco Martins Pacheco, Endocardio, Bel\u00e9m, PA. Jonathan Scapin Zagatti, Cardio Ritmo Diagn\u00f3sticos, Jales, SP. Jos\u00e9 Joaquim Raposo, Servi\u00e7o de Cardiologia da Santa Casa de Limeira, SP. Jos\u00e9 Roberto Moya, Biocardios, Cuiab\u00e1, MT. Josaf\u00e1 de Oliveira Costa, Cl\u00ednica Vitta, Igarassu, PE. Josiedson Pontes de Farias, Cardio Diagn\u00f3sticos, Caruaru, PE. Juan Carlos Yugar Toledo. Endocor, Rio Preto, SP. Lilian Mesquita, Ergo Med Setor De Cardiologia/Geriatria, RJ. Lola Helbingen Santos, Cardiodiagn\u00f3sticos, Goiania, GO. Luam Vieira de Almeida Di\u00f3genes, Procardiaco, Teresina, PI. Luiz Kencis J\u00fanior, Lapacor, S\u00e3o Paulo, SP. Marcelo J\u00falio de Oliveira, Cl\u00ednica Cardiograficos, Ribeir\u00e3o Preto, SP. Marco Ant\u00f4nio de M Alves, Escada Clinical Center, Escada, PE. Marco A. M. Gomes, Centro de Pesquisas Cl\u00ednicas do Cesmac/Hospital do Cora\u00e7\u00e3o de Alagoas. Marcos Alberto Pires Meira J\u00fanior, Clincar Jo\u00e3o Pessoa PB. Maria Christina Cavalcanti Ballut, MEDCENTRO, Manaus, AM. Marcus Vin\u00edcius de Oliveira, Cardiodiagn\u00f3sticos, Goi\u00e2nia-GO. Maria Beatriz M. B. L. Rodrigues, Cardiovida, Porto Velho, RO. Mayara Cedrim Santos, Instituto UNICAP de Pesquisa Cl\u00ednica, Recife, PE. Naiara Pedrassi Engracia Garcia Caluz, Centro de Medicina Avan\u00e7ada Dr Luiz Kencis, S\u00e3o Paulo, SP. Nelson Dinamarco, Ambulat\u00f3rio de Hipertens\u00e3o Arterial, Colegiado de Medicina, Universidade Estadual Santa Cruz \u2013 UESC. Nildo Magalh\u00e3es, SOBAM Jundia\u00ed, SP. Paulo Roberto Pereira de Sant\u2019Ana, MEDCOR Cardiologia, Nova Igua\u00e7u- RJ. Rafael Nogueira de Macedo, Centro Cardiol\u00f3gico S\u00e3o Camilo, CE. Paulo S\u00e9rgio Lopes Soares, Universidade De Vassouras - Hospital Universit\u00e1rio de Vassouras, RJ. Ricardo Mesquita de Freitas, Cardiocenter, Barreiras, BA. Roberto de A. Dultra, Clinicor, Itabuna, Bahia, BA. Roberto Dischinger Miranda, Servi\u00e7o de Cardiologia, Disciplina de Geriatria e Gerontologia, Escola Paulista de Medicina, Universidade Federal de S\u00e3o Paulo. Rodrigo Cunha de Sousa, Centro Integrado de Medicina Invasiva \u2013 CIMI, Uberaba, MG. Rog\u00e9rio Krakauer, Santa Casa de SP. Rogerio Ruiz, HD HomeDoctor, SP. Ruy Morando, Cincor - Centro Integrado do Cora\u00e7\u00e3o, Americana, SP. S\u00e9rgio Augusto Vieira Sim\u00f5e, Consult\u00f3rio M\u00e9dico Integrado, T\u00e1ssia T\u00e2mara Silva Feitosa, Ok Doutor, Recife, PE. Tobias Barreto, SE. Sheyla Cristina Tonheiro Ferro da Silva, CLINSAUDE e CEMISE, Aracaju, SE. Vanderlei Magalh\u00e3es da Silveira, Cardiologista, Faculdade de Medicina, Universidade de Passo Fundo. Vanildo Guimar\u00e3es, Diagn\u00f3stico Card\u00edaco, Recife, PE. Vilma Helena Burlamaqui, Consult\u00f3rio de Cardiologia, Niter\u00f3i, RJ. Vitor Bruno Teixeira de Holanda, Climile, Ananindeua, PA. Walmir de Vasconcelos Ratier Thomaz, Rio de Janeiro, RJ. Weimar Sebba Barroso, Liga de Hipertens\u00e3o Arterial UFG, Goi\u00e2nia, GO. Wenderson Tavares dos Santos, Hospital Biocor, Belo Horizonte, MG 3 Drugs that effectively reduce blood pressure (BP) also protect against the main outcomes of hypertensive disease, and the best results can be expected of drugs with a long half-life  that do not negatively interfere in metabolic parameters. It is also known that small BP reductions, even in the early stages of arterial hypertension, can lead to reductions in the main cardiovascular outcomes.5Adequate treatment and control is one of the great challenges in arterial hypertension, which is the leading cause of death worldwide. Aligning treatment strategies with the most current scientific is one way to optimize these results.6On the other hand, despite such evidence, the Brazilian Unified Health System provides medications with a short half-life that are used in monotherapy and require several doses a day. Such characteristics can negatively impact adherence and hinder adequate BP control. It should be emphasized that the Brazilian Unified Health System reflects the drug strategy used for 75% of the hypertensive patients in our country.7A 2021 study evaluated a database of 22,446 individuals who underwent home and office BP measurement, 11,337 of whom were being treated for hypertension by cardiologists with antihypertensive drugs. In 74.6% of the cases, renin-angiotensin-aldosterone system blockade was used, including angiotensin receptor blockers (ARBs) in 58.7%, either in monotherapy or combination therapy.The objectives of the present study were: (i) to verify the distribution of ARB prescription in monotherapy and combined therapy according to sex, geographic region, and diabetes status; (ii) to compare BP control according to casual and home BP monitoring measurement (HBPM) for all ARB treatment strategies; (iii) to compare BP control in casual and HBPM measurements; and (iv) to compare mean systolic blood pressure (SBP), diastolic blood pressure (DBP), pulse pressure (PP), and BP variability obtained through ARBs in monotherapy or double combination therapy, considering the class as a whole and individual types.This study was approved by the Human Research Ethics Committee of the Hospital das Cl\u00ednicas of the Federal University of Goi\u00e1s (opinion 99691018.7.0000.5078) and evaluated patients who were examined on the TeleHBPM platform (www.telemrpa.com) between May 2017 and October 2020.8The platform was developed as a remote reporting tool for telemonitoring, including features that allow the database to be analyzed and filtered according to research questions. The mathematical algorithm allows analysis while protecting the personal data of patients and health facilities, whether interpreting exams or developing research projects. Since it is not software, but a platform accessible on any device via an Internet connection, BP measurements can be uploaded quickly and remotely.The database search was limited to patients who used ARBs. Patients aged at least aged 18 years on monotherapy or double combination therapy were included. Patients on a combination of three or more antihypertensives, antihypertensives in combination with angiotensin-converting enzyme inhibitors, or antihypertensives in double combination therapy with infrequently used antihypertensives  were excluded   . We alsThe following data were collected from the TeleHBPM platform: sex, age (in years), body mass index, number of valid HBPM measurements, casual and HBPM SBP and DBP measurements, blood pressure variability based on HBPM measurements obtained through the standard deviation of the 24 household measurements taken during the protocol, drug class used, and type of ARB. The regional distribution of the sample was also evaluated, as was the prevalence of individuals who used medications to treat diabetes mellitus .9 HBPM was performed with the provided device; patients were instructed about proper handling and BP measurement on the day the device was delivered.1 On that day, two measurements were taken in a clinical/office environment and, over the next 4 days, the patient (and/or caregiver/companion) performed the measurements at home according to protocol. The mean of the two measurements taken on the first day was considered the casual measurement, and the mean of the 24 measurements taken from the second to the fifth day was considered the HBPM measurement.10 Only validated automatic devices  were used.The Quetelet formula was used to calculate body mass index based on weight and height data.1The data were exported from the TeleHBPM platform to Microsoft Excel. All drug classes described on the platform were reviewed and coded by two work teams. The databases were then cross-referenced to identify discrepant data, which, when present, were reviewed by the entire team. Individuals whose SBP/DBP values were <140/90 mmHg in casual measurement and <130/80 mmHg in HBPM, respectively, were considered to have controlled BP.Statistical analysis was performed in Stata 14.0. Quantitative variables were expressed as mean and standard deviation, and qualitative variables were expressed as absolute and relative frequencies. The Kolmogorov-Smirnov test was used to verify the normality of the data.t -test. The chi-square test or Fisher\u2019s exact test was used to compare BP control rates according to the casual and HBPM measurements, as well as to compare the rates of BP control for each drug strategy.The mean SBP, DBP and PP values obtained in casual and HBPM measurements were compared using a paired Student\u2019s Overlapping 95% confidence intervals were used to compare the differences in mean SBP, DBP, PP and BP variability obtained with ARB monotherapy or double combination therapy, considering the class as a whole and individual types. P-values <0.05 were considered significant.A total of 12,813 patients were evaluated, the majority of whom were female. The Northeast was the most prominently represented region, with approximately half of the patients. The prevalence of diabetes was 6.2%   .Double combination therapy was slightly more prevalent than monotherapy (51.2% vs. 48.5%). The following types of ARBs were used: losartan (57.2%), olmesartan (18.8%), valsartan (15.0%), telmisartan (4.8%), candesartan (3.8%), and irbesartan (0.4%).The mean number of valid HBPM measurements was 23.3. The differences in mean casual and HBPM values for SBP and DBP were 6.7 mmHg (p < 0.001) and 4.5 mmHg (p < 0.001), respectively. These differences characterize the white-coat effect and were maintained across all treatment strategies. This behavior was repeated in all ARBs, whether in monotherapy or combination therapy. We also compared the rate of BP control by casual and HBPM measurements in monotherapy and combination therapy   .1 overall BP control was better in casual measurement. In HBPM, BP control was lower in ARB monotherapy and in ARBs combined with beta-blockers. Among the ARB types used in monotherapy or combination therapy, BP control was lower with losartan and higher with long half-life ARBs. This trend was repeated in the casual measurements.According to the goals of <140/90 mmHg  and <130/80 mmHg (HBPM) recommended by current guidelines,The control rates of different ARBs in combination with CCA, BB, or diuretics were lower in combinations with losartan and higher in ARBs with a long half-life in both HBPM and casual measurements. In HBPM, the mean SBP for ARB + CCA and ARB + diuretics was lower than that of ARB monotherapy. In monotherapy, the BP values were progressively higher for olmesartan, candesartan, telmisartan, valsartan and losartan   . In comPP was higher with ARB + BB than any other combination or ARB monotherapy. Losartan in monotherapy or in double combination therapy resulted in a higher mean PP than candesartan or telmisartan.BP variability was greater with ARB + CCA than in combinations with diuretics or BB or in monotherapy. Whether in monotherapy or combination therapy, BP variability was lower with telmisartan than valsartan. Losartan + CCA had lower mean variability than other combinations. Candesartan + BB showed greater variability than candesartan + CCA. There was no difference in BP variability between combinations with valsartan, olmesartan and telmisartan7 Thus, it makes sense to assess BP behavior in response to various ARB types in both clinical and home settings.The present study, a further development of an analysis published in 2020, found that, in hypertensive patients treated with monotherapy or double combination therapy, different possible combinations of ARB types resulted in significantly lower mean SBP and DBP in HBPM than in casual measurements, as well as that ARBs were the most common treatment option.13Our sample population had a mean age of approximately 60 years and a high body mass index. The patients were also predominantly women, and most resided in the Northeast and Southeast regions. It is important to consider that advanced age and excess weight can impede achieving recommended arterial hypertension treatment goals.16 This change explains the difference in BP control rates found in casual and HBPM measurements in this analysis compared to our previous article.7It should also be noted that in the last year, as a result of HBPM evidence published in the national database, the reference values for normality were lowered from 135/85 mmHg to 130/80 mmHg.3 Dual combination therapy with diuretics and CCAs was preferred, which is in line with current recommendations.19Regarding the treatment strategies used in this sample, 48.5% received ARB monotherapy, 23.4% received ARBs combined with diuretics, 16.8% received ARBs combined with CCAs, and 11.2% received ARBs combined with BBs. Interestingly, although hypertension guidelines unanimously recommend drug combinations for most cases of hypertension, monotherapy was still quite frequent.22Another relevant aspect in selecting arterial hypertension drugs is a long half-life, which allows a single daily dose; these characteristics directly interfere with treatment adherence and adequate BP control. Drugs with a short half-life must be taken twice or more daily to maintain their plasma level and efficacy in reducing BP levels.23It is interesting to note that, from a pharmacological point of view, there are important differences between these drugs, and the different half-lives of ARBs  may be related to the differences we found in BP behavior.When evaluating the BP control rate by casual and HBPM measurements, we found that 56.3% and 44.5% of the patients, respectively, were within the goals. We found different percentages of patients with controlled BP among the different ARB types and combinations.For a more refined analysis of this behavior, we determined the mean HBPM measurements and confidence intervals of SBP, DBP, and pressure variability. Combinations with BBs resulted in higher mean SBP values and variability than combinations with diuretics or CCAs. In monotherapy, losartan had the highest mean SBP and DBP values of the longer half-life ARBs.This observational study was limited by the fact that it did not assess the dosage of each drug, and the sample was not representative of the Brazilian population. On the other hand, it analyzed data from a large database that reflected ARB usage strategies in hypertensive patients, allowing important parameters to be determined regarding BP behavior with different drugs in monotherapy and combination therapy.28 and, more importantly, they reflect the need to review the Brazilian Unified Health System\u2019s strategy for antihypertensive drugs,6 since it is known that small BP reductions in hypertensive patients have important repercussions on cardiovascular morbidity and mortality.These findings are consistent with those of previously published randomized studies that evaluated the antihypertensive efficacy of different ARBsIn hypertensive patients treated with ARBs, monotherapy is still frequent. In combined therapy, diuretics and CCAs are preferred. Among ARBs, losartan is still used in more than half of patients, whether in monotherapy or double combination therapy, despite being the least efficient medication for reducing and controlling BP. There are clear differences in the half-life of ARBs, which was seen in BP behavior through both casual and HBPM measurements. These differences may reflect the effectiveness of blood pressure control.Adriana Siqueira Serpa de Menezes, SAVE, Recife, PE. Andr\u00e9a Ara\u00fajo Brand\u00e3o, Universidade do Estado do Rio de Janeiro, RJ. Anibal Prata Barbosa, Prog de Hip. Arterial Secretaria de Sa\u00fade de Duque de Caxias, RJ. Antonio Almeida Braga, PROCAPE, UPE, Recife, PE. Antonio Eduardo de Melo Filho, Cl\u00ednica de Sa\u00fade Dr Antonio Eduardo de Melo, Triunfo, PE. \u00c1tila de Oliveira Melo, Liga de Hipertens\u00e3o Arterial UFG, Goi\u00e2nia, GO. Andr\u00e9 K Vidigal de Vasconcellos, Instituto de Cardiologia do Agreste, Caruaru, PE. Audes D. M. Feitosa, Unidade de Hipertens\u00e3o e Cardiologia Preventiva, PROCAPE/UPE, Recife, PE. Breno Gontijo de Camargos, AngioCor, Taguatinga, DF. Bruno Alencar Fonseca, Cl\u00ednica Blues, Belo Horizonte, MG. Bruno Daniel Ferrari, Funda\u00e7\u00e3o Educacional do Munic\u00edpio de Assis, FEMA, SP. Bruno Jos\u00e9 Peixoto Coutinho, Cardiologia Hospital Oswaldo Cruz, Universidade de Pernambuco, PE. Carlo Bonasso, Cl\u00ednica M\u00e9dica Carlo Bonasso SS Ltda, S\u00e3o Paulo, SP. Carlos Jos\u00e9 Mota de Lima, Centro Cardiol\u00f3gico S\u00e3o Camilo, CE. Carlos Filinto de Almeida, Instituto do Cora\u00e7\u00e3o de Mato Grosso do Sul, Campo Grande, MS. Claudinelli Alvarenga Aguilar, Cl\u00ednica do Esporte, Goi\u00e2nia, GO. C\u00e9sar Ricardo Soares Medeiros, Cl\u00ednica de Cardiologia Dr C\u00e9sar Medeiros, Ribeir\u00e3o Preto, SP. Cristiano Pederneiras Jaeger, Instituto de Medicina Vascular - Coracentro, Porto Alegre/RS. Daniel Lages Dias, Novacordis, Paul\u00ednia, SP. Diogo da Silva Amorim, Liga de Hipertens\u00e3o Arterial UFG, Goi\u00e2nia, GO. Ednaldo M. Fontes Segundo, Cardiologista pela SBC, Instituto Paulo Gomes (IPG) em Est\u00e2ncia, SE. Eduardo C. D. Barbosa, Dept Hipertens\u00e3o e Cardiometabolismo Hospital S\u00e3o Francisco, Santa Casa Porto Alegre, RS. Eduardo \u00c9rico Zen, Hospital Cardiol\u00f3gico Costantini, Curitiba, PR. Elder Gil A. Cruz, Cl\u00ednica do Cora\u00e7\u00e3o Dr. Elder Gil, Salgueiro, PE. Esther G. Di\u00f4go de Lima de B. Carvalho, Cl\u00ednica S\u00e3o Lucas, Guarabira, PB. F\u00e1bio Argenta, Mediodonto, Cuiab\u00e1, MT. Fabiano de Souza Ramos, MEDCOR Cardiologia, Nova Igua\u00e7u- RJ. Fl\u00e1via Karina Silva e Oliveira, Centro de Cardiologia, S\u00e3o Jos\u00e9 dos Campos, SP. Fl\u00e1vio H. A. P. V\u00e9ras, Cl\u00ednica do Cora\u00e7\u00e3o, Mossor\u00f3, RN. Francisco Deoclecio Pinheiro, Cl\u00ednica de Especialidades M\u00e9dicas de Itapipoca, Itapipoca, CE. Frank Land L. de Carvalho, Cardiovasf, Petrolina, PE. Germano Granja, Cl\u00ednica do Cora\u00e7\u00e3o, Ouricuri, PE. Giovanni Saraiva, Imedi e Icordis, Recife, PE. Gleidson Junio Oliveira de Souza, Liga de Hipertens\u00e3o Arterial UFG, Goi\u00e2nia, GO. Gustavo Barros - MCOR / Recife-PE. Gustavo Guimar\u00e3es Moreira de Castro, ITACORDIS e Universidade Igua\u00e7u \u2013 UNIG, Itabora\u00ed e Nova Igua\u00e7u, RJ. Jadil Francisco Fusturath J\u00fanior, Cardio Service, Porto Velho, RO. Jos\u00e9 Wladimir Tambelli Pires, Cl\u00ednica de Cardiologia, Itapetininga/SP. Jo\u00e3o Evaristo de Oliveira Dantas, Cardiomed/Multimed, S\u00e3o Lu\u00eds, MA. Jo\u00e3o F\u00e9lix de Morais Filho, Cl\u00ednica Angioc\u00e1rdio, Natal / RN. Jo\u00e3o Francisco Martins Pacheco, Endocardio, Bel\u00e9m, PA. Jonathan Scapin Zagatti, Cardio Ritmo Diagn\u00f3sticos, Jales, SP. Jos\u00e9 Joaquim Raposo, Servi\u00e7o de Cardiologia da Santa Casa de Limeira, SP. Jos\u00e9 Roberto Moya, Biocardios, Cuiab\u00e1, MT. Josaf\u00e1 de Oliveira Costa, Cl\u00ednica Vitta, Igarassu, PE. Josiedson Pontes de Farias, Cardio Diagn\u00f3sticos, Caruaru, PE. Juan Carlos Yugar Toledo. Endocor, Rio Preto, SP. Lilian Mesquita, Ergo Med Setor De Cardiologia/Geriatria, RJ. Lola Helbingen Santos, Cardiodiagn\u00f3sticos, Goiania, GO. Luam Vieira de Almeida Di\u00f3genes, Procardiaco, Teresina, PI. Luiz Kencis J\u00fanior, Lapacor, S\u00e3o Paulo, SP. Marcelo J\u00falio de Oliveira, Cl\u00ednica Cardiograficos, Ribeir\u00e3o Preto, SP. Marco Ant\u00f4nio de M Alves, Escada Clinical Center, Escada, PE. Marco A. M. Gomes, Centro de Pesquisas Cl\u00ednicas do Cesmac/Hospital do Cora\u00e7\u00e3o de Alagoas. Marcos Alberto Pires Meira J\u00fanior, Clincar Jo\u00e3o Pessoa PB. Maria Christina Cavalcanti Ballut, MEDCENTRO, Manaus, AM. Marcus Vin\u00edcius de Oliveira, Cardiodiagn\u00f3sticos, Goi\u00e2nia-GO. Maria Beatriz M. B. L. Rodrigues, Cardiovida, Porto Velho, RO. Mayara Cedrim Santos, Instituto UNICAP de Pesquisa Cl\u00ednica, Recife, PE. Naiara Pedrassi Engracia Garcia Caluz, Centro de Medicina Avan\u00e7ada Dr Luiz Kencis, S\u00e3o Paulo, SP. Nelson Dinamarco, Ambulat\u00f3rio de Hipertens\u00e3o Arterial, Colegiado de Medicina, Universidade Estadual Santa Cruz \u2013 UESC. Nildo Magalh\u00e3es, SOBAM Jundia\u00ed, SP. Paulo Roberto Pereira de Sant\u2019Ana, MEDCOR Cardiologia, Nova Igua\u00e7u- RJ. Rafael Nogueira de Macedo, Centro Cardiol\u00f3gico S\u00e3o Camilo, CE. Paulo S\u00e9rgio Lopes Soares, Universidade De Vassouras - Hospital Universit\u00e1rio de Vassouras, RJ. Ricardo Mesquita de Freitas, Cardiocenter, Barreiras, BA. Roberto de A. Dultra, Clinicor, Itabuna, Bahia, BA. Roberto Dischinger Miranda, Servi\u00e7o de Cardiologia, Disciplina de Geriatria e Gerontologia, Escola Paulista de Medicina, Universidade Federal de S\u00e3o Paulo. Rodrigo Cunha de Sousa, Centro Integrado de Medicina Invasiva \u2013 CIMI, Uberaba, MG. Rog\u00e9rio Krakauer, Santa Casa de SP. Rogerio Ruiz, HD HomeDoctor, SP. Ruy Morando, Cincor - Centro Integrado do Cora\u00e7\u00e3o, Americana, SP. S\u00e9rgio Augusto Vieira Sim\u00f5e, Consult\u00f3rio M\u00e9dico Integrado, T\u00e1ssia T\u00e2mara Silva Feitosa, Ok Doutor, Recife, PE. Tobias Barreto, SE. Sheyla Cristina Tonheiro Ferro da Silva, CLINSAUDE e CEMISE, Aracaju, SE. Vanderlei Magalh\u00e3es da Silveira, Cardiologista, Faculdade de Medicina, Universidade de Passo Fundo. Vanildo Guimar\u00e3es, Diagn\u00f3stico Card\u00edaco, Recife, PE. Vilma Helena Burlamaqui, Consult\u00f3rio de Cardiologia, Niter\u00f3i, RJ. Vitor Bruno Teixeira de Holanda, Climile, Ananindeua, PA. Walmir de Vasconcelos Ratier Thomaz, Rio de Janeiro, RJ. Weimar Sebba Barroso, Liga de Hipertens\u00e3o Arterial UFG, Goi\u00e2nia, GO. Wenderson Tavares dos Santos, Hospital Biocor, Belo Horizonte, MG"}
+{"text": "Oxidative phosphorylation is an essential feature of Animalian life. Multiple adaptations have developed to protect against hypoxia, including hypoxia-inducible-factors (HIFs). The major role of HIFs may be in protecting against oxidative stress, not the preservation of high-energy phosphates. The precise mechanism(s) of HIF protection is not completely understood.To better understand the role of hypoxia-inducible-factor-1, we exposed heart/myocardium cells (H9c2) to both normoxia and hypoxia, as well as cobalt chloride (prolyl hydroxylase inhibitor), echniomycin (HIF inhibitor), A2P (anti-oxidant), and small interfering RNA to beclin-1. We measured cell viability, intracellular calcium and adenosine triphosphate, NADP/NADPH ratios, total intracellular reactive oxidative species levels, and markers of oxidative and antioxidant levels measured.Hypoxia (1%) leads to increased intracellular Ca2+ levels, and this response was inhibited by A2P and echinomycin (ECM). Exposure of H9c2 cells to hypoxia also led to an increase in both mRNA and protein expression for Cav 1.2 and Cav 1.3. Exposure of H9c2 cells to hypoxia led to a decrease in intracellular ATP levels and a sharp reduction in total ROS, SOD, and CAT levels. The impact of hypoxia on ROS was reversed with HIF-1 inhibition through ECM. Exposure of H9c2 cells to hypoxia led to an increase in Hif1a, VEGF and EPO protein expression, as well as a decrease in mitochondrial DNA. Both A2P and ECM attenuated this response to varying degrees.Hypoxia leads to increased intracellular Ca2+, and inhibition of HIF-1 attenuates the increase in intracellular Ca2+ that occurs with hypoxia. HIF-1 expression leads to decreased adenosine triphosphate levels, but the role of HIF-1 on the production of reactive oxidative species remains uncertain. Anti-oxidants decrease HIF-1 expression in the setting of hypoxia and attenuate the increase in Ca2+ that occurs during hypoxia (with no effect during normoxia). Beclin-1 appears to drive autophagy in the setting of hypoxia (through ATG5) but not in normoxia. Additionally, Beclin-1 is a powerful driver of reactive oxidative species production and plays a role in ATP production. HIF-1 inhibition does not affect autophagy in the setting of hypoxia, suggesting that there are other drivers of autophagy that impact beclin-1. Animalian world with which we concern ourselves, the most efficient source of energy is mitochondrial oxidative phosphorylation  but also through NF-\u03ba\u03b2, the unfolded protein response, and mammalian target of rapamycin (mTOR) kinase \u20138. DurinZhang et al. significantly increased our understanding of HIF-1\u03b1. Using mouse embryonic fibroblasts as a model, they found that knockout cell (KO) lines without HIF-1\u03b1 consumed more oxygen and had higher ATP levels when compared to wild-type (WT) cells during both normoxia and hypoxia \u201314. A la2+]i levels? Is the role of HIF-1 in attenuating oxidative stress universal across all cell types? How do anti-oxidant defenses (exogenous or endogenous) impact HIF-1\u03b1 expression? Is HIF-1\u03b1 the only pathway that drives autophagy via the BNIP3-Beclin-1-Atg5 pathway? The purpose of this study was to answer these questions. Our primary hypothesis was that in the setting of hypoxia, HIF-1\u03b1 inhibition would lead to increased intracellular calcium-mediated by reduced expression of L-type voltage-dependent Ca channels.While Zhang et al. have made major contributions to our understanding of adaptive responses to hypoxia , important questions remain. Does HIF-1\u03b1 exert an influence on [CaRattus norvegicus) heart/myocardium cells  were cultured and maintained following standard provider protocol. H9c2 cells (1 \u00d7 106 cells/ml) were cultured in complete cell culture media [ATCC-formulated Dulbecco\u2019s Modified Eagle\u2019s Medium  supplemented with 10% fetal bovine serum ] and were incubated in a humidified atmosphere with 5% CO2 at 37\u00b0C, and allowed to grow up to 70\u201380% confluence before exposure to hypoxia or normoxia. When present, 200 \u03bcM . In the hypoxic experimental group, H9c2 cells were cultured under Hypoxia (1%O2) in a complete culture medium supplemented with ECM or A2P and were maintained in a modular incubator chamber flushed with a gas mixture containing 1%O2, 5% CO2, and 94% N2 at 37\u00b0C for 24 h. For controls, H9c2 cells were cultured in a complete culture medium under normoxia conditions. After incubation, cells were harvested immediately for analysis or preserved at \u201380\u00b0C freezer for future studies. Cell culture environment, reagent concentrations and specificity of all experimental groups are listed in To mimic hypoxic conditions, H9c2 cells were cultured at a density of 1 \u00d7 106 cells/ml) were seeded in a 6-well plate containing a complete culture medium and cultured to 70\u201380% confluence before transfection. For knockdown of Beclin-1/Becn1 , 10 \u03bcl of transfection reagent  was diluted in 250 \u03bcl OptiMEM serum reduced medium, to this 25 nM of Beclin-1 siRNA was added and incubated at room temperature for 30 min to form transfection reagent: siRNA complex development. The transfection reagent: siRNA complex was then added to 70\u201380% confluence H9c2 cells (replaced with culture medium supplemented with 1%FBS) and incubated at 37\u00b0C with 5% CO2 in an incubator for 1 h. After 1 h incubation, culture medium FBS was increased to 10% (complete culture medium), and these pre- Beclin-1- siRNA transfected cells were used for mimicking hypoxic conditions. siRNA and transfection reagents concentrations were used according to the manufacturer\u2019s instructions.H9c2 cells (1 \u00d7 102 and Hypoxia) exposed H9c2 cells viability was measured using the CellTiter-Glo\u00ae Luminescent cell viability assay kit . Briefly, H9c2 cells were plated in 6-well plates and grown up to 70\u201380% confluence, then the plates were exposed to hypoxia or normoxia conditions, or Cobalt chloride as described in the methodology section. After incubation, H9c2 cells were incubated at room temperature for 15 min, and then, 100 \u03bcL of CellTiter-Glo reagent were added to cells and then placed on an orbital shaker for 2 min to induce cell lysis. Lysed cells were incubated at room temperature for 10 min to stabilize the luminescent signal and then using Synergy\u2122 Multi-Mode Microplate Reader  luminescence was recorded. All the samples were read in duplicate. To represent the relative density of adhering cells (live cells) in culture dishes, we performed the crystal violet staining on CoCl2 and Hypoxia exposed H9c2 cells. In brief, after cells were exposed to hypoxic conditions, culture media was removed and the cells were washed with PBS  and fixed for 20 min at RT with staining solution . Fixed cells were rinsed with water and air-dried and images were obtained using a Nikon TS100 digital camera microscope.Experimental groups  levels in CoCl2 and Hypoxia exposed H9c2 cells were assessed using the Fluo-4 NW calcium assay kit , by following the manufacturer\u2019s protocol. Briefly, after cells were exposed to CoCl2 or hypoxia conditions, culture media was removed to avoid the source of baseline fluorescence then quickly 100-\u03bcl of Fluo-4 NW assay reagent was added and incubated for 30 min at 37\u00b0C, then incubated for an additional 30 min in the dark at room temperature. The relative fluorescence units (RFU) emitted by the Fluo-4-NW dye were quantified using a Synergy\u2122 Multi-Mode Microplate Reader , excitation at 494 nm, and emission at 516 nm.Intracellular energy (ATP) levels were measured using the Bioluminescent Assay kit  according to the manufacturer\u2019s protocol and as previously described . The addChanges in the NADP/NADPH ratio were evaluated using an NADP/NADPH -Glo assay kit . Briefly after exposing cells to CoCl2 or hypoxia conditions, culture media was removed then quickly 50-\u03bcl of PBS and 50-\u03bcl of NADP/NADPH -Glo assay reagents were added, gently mixing the plate, and incubated for 30 min in dark at room temperature. After 30 min, NADP/NADPH ratios were (luminescence) measured using Synergy\u2122 Multi-Mode Microplate Reader . All the samples were read in duplicate.Osuru et al. H9C2 cells Lipid peroxidation level was determined using the thiobarbituric acid (TBA) method with Lipid Peroxidation [malondialdehyde (MDA)] assay kit . H9C2 cell\u2019s antioxidant levels were measured by quantifying the total SOD (Superoxide dismutase) levels using the SOD colorimetric activity kit  and by estimating the catalase (CAT) protein levels using western blotting method (for antibody concentration and methodology please see the western blotting section).The myocardium (H9C2 cells) total ROS, oxidative stress, and antioxidant levels toward hypoxia, HIF-1\u03b1 inhibition, and under different treatments were measured using commercially available assay kits by following the manufacturer\u2019s protocol. Intracellular total ROS levels were measured using a 2\u2032-7\u2032-dichlorofluorescein diacetate-based total ROS detection kit . Oxidative stress levels were assayed using Protein Carbonyl assay using the bioluminescent assay kit  by following the manufacturer\u2019s protocol and as previously described by \u00ae hypoxia/oxidative stress detection kit . H9c2 cells were cultured and maintained as described above in 8- chambered cell culture slides  and allowed to grow up to 70\u201380% confluence, then the cells were exposed to CoCl2 or hypoxia conditions. After cells were exposed to CoCl2 or hypoxia, culture media was removed and cells were washed gently twice with PBS then ROS-ID\u00ae hypoxia/oxidative stress detection solution was added and then cells were re- incubated under normal tissue culture conditions for 30 min. The detection solution was carefully removed, cells were washed twice with PBS, nuclei were stained with Hoechst 33342, then the slides were covered with a coverslip and a fluorescence microscope  was used to capture the images using Green Filter for Oxidative stress and Texas Red Filter for Hypoxia detection. Data were normalized with nuclei stain (Hoechst 33342) fluorescence intensity and expressed as changes in fluorescence intensity.Cellular hypoxia/oxidative stress levels were evaluated using a ROS-ID\u00ae Green supermix  and specific gene primers  and the extracted RNA was quantified using a spectrophotometer- NanoDrop . Total RNA (2\u20135 \u03bcg) was reverse transcribed to cDNA using the iScript-Adv cDNA Synthesis Kit according to the manufacturer\u2019s recommendation . The cDNA (50\u2013100 ng) was used for real-time PCR analysis in a final volume of 20 \u03bcl containing, iTaq universal SYBR primers . qPCR wa primers . Each reThe sources of antibodies were as follows: from Cell Signaling Technology Inc: Hypoxia-inducible factor 1-alpha , Hypoxia-inducible factor 2-alpha , Hypoxia-inducible factor 1-beta . Autophagy Antibody Sampler Kit #4445 , Autophagy Related-5 , Autophagy Related-12 , Atg16L1,#8089,1:1000), heme-oxygenase-1  and nitric oxide synthase . From Novus Biologicals: B-cell lymphoma 2  and BCL2 Interacting Protein 3 Antibody . From GeneTex Inc: BCL Interacting Protein 3L  Calcium Voltage-Gated Channel Subunit Alpha1 S , Calcium Voltage-Gated Channel Subunit Alpha1 C , Calcium Voltage-Gated Channel Subunit Alpha1 D , Superoxide Dismutase 1 antibody  and Catalase antibody . From Proteintech Group Inc: Mechanistic Target of Rapamycin Kinase , AKT Serine/Threonine Kinase , AMP-activated protein kinase , Vascular endothelial growth factor-a , Erythropoietin  and loading control \u03b2-actin antibody .C and were separated using 4\u201320% Tris-Glycine polyacrylamide gradient gels (Bio-Rad). Proteins were transferred to PVDF membranes (Millipore) and residual protein sites were blocked with SuperBlock phosphate-buffered saline (PBS) Blocking Buffer (Thermo-Fisher #37515) and then incubated overnight with primary antibodies at the recommended concentrations at 4\u00b0C. Membranes were washed and incubated with horseradish peroxidase (HRP) conjugated secondary antibodies anti-mouse or anti-rabbit  antibodies at room temperature. Membranes were washed then the antigen-antibody complexes were visualized using Super Signal West Femto enhanced chemiluminescence substrate (Thermo Scientific). Images were captured using GBOX , and the intensity of the bands corresponding to the protein expression level was quantified with the computerized image analysis software . Target protein expression levels were normalized to the housekeeping protein levels (\u03b2-actin) and changes in protein expression levels were presented as ratios.Selected protein expression levels were estimated using Western blot analysis by following previous reports with slight modification . H9c2 ce2+levels in H9c2 cells in addition to mRNA and protein analysis, we also used the immunofluorescence staining technique for Cav1.1 (1:200), Cav1.2 (1:200), and Cav1.3 (1:200) proteins, by following previous reports with slight modification  to detect autophagy in CoCl2 or hypoxia treated H9c2 cells. Briefly, H9c2 cells were cultured in 8-chambered cell culture slides and exposed to CoCl2 or hypoxia conditions. After cells were exposed to CoCl2 or hypoxia, cells were incubated with fluorescent Cyto-ID dye at 37\u00b0C for 30 min followed by nuclei stain-Hoechst 33342 for 5 min. After incubation, representative images were acquired using a fluorescence microscope .Mt_Nd1 and Mt_Nd6) and a nuclear gene Tubulin (Tuba1) (\u00ae Green supermix (BioRad) were used for real-time PCR analysis. Gene expression (Fold changes) was calculated using the 2-\u0394\u0394Ct method  levels of the H9C2 cells were quantified by following previously reported methods with slight modification . H9C2 ce (Tuba1) . 100 ng t method , each re*, +, # and \u2260 denotes significant, \u201cns\u201d represents not significant by two-way ANOVA- multiple comparisons test. P-values are nsp > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001 and ****p < 0.0001 compared to Control; +p < 0.05, ++p < 0.01, + + +p < 0.001 and + + + +p < 0.0001 compared to Cocl2; #p < 0.05, ##p < 0.01, ###p < 0.001 and ####p < 0.0001 compared to Hypoxia; \u2260p < 0.05, \u2260\u2260p < 0.01, \u2260\u2260\u2260p < 0.001 and \u2260\u2260\u2260\u2260p < 0.0001 compared to si-becn1 (21%O2) Vs, si-becn1 (1%O2).All the data groups mRNA, protein expression, Mitochondrial DNA levels, ATP, ROS, NADP/NADPH ratios, and cell viability assays, control Vs experimental groups , and siRNA groups , were compared using the two-way ANOVA (or Mixed model) multiple comparisons compare groups column means (main column effect) with 0.05 . All calculations were performed using GraphPad Prism 8 . Unless otherwise noted, each experiment was performed at least two to three times . All the data are shown in mean \u00b1 S.D, Symbols 2+ levels  and hypoxia (1%O2) did not change the Cav1.1 protein expression significantly  as well as reactive species  and EPO (Erythropoietin) protein expression levels were also significantly increased in Hypoxia and in Cocl2 exposed cells and these overexpressed protein levels were ameliorated with A2P (anti-oxidant) and ECM (Hif1 inhibition) treatment. Beclin-1 inhibition cells grown under normoxia and in hypoxia conditions also expressed the decreased VEGF and EPO protein expression levels compared to Hypoxia and Cocl2 exposed cells .Exposure of H9c2 cells to hypoxia led to an increase in both ATG5 mRNA  and protBecn1 or beclin-1 mRNA and protein expression with exposure of H9c2 cells to hypoxia  exposed cells. In addition, Becn1 protein expression levels also decreased in Hypoxia and in Hypoxia (1%O2) + ECM groups. Beclin-1 inhibition also confirms the decreased Becn1 protein under normoxia (21%O2) exposure and in Hypoxia  levels  levels . Hypoxia) levels \u20134. Hypox) levels , and inc) levels . NADP/NA) levels . HIF1\u03b1 a) levels  and Hypo) levels  are decr) levels  and decr) levels  and mito) levels . The maj2+]i which is not unexpected based on previously published data showing increased [Ca2+]i in both endothelial (2+ channel) which is normally known to be increased during hypoxia  leads to a significant increase in [Ca2+]i  is unclear. Zhang et al. exposed mouse embryonic fibroblast (MEFs) to hypoxia (1%O2) and found that hypoxic MEFs deficient in HIF-1\u03b1 exhibited higher levels of O2 consumption, ATP, and ROS than wild-type (WT) cells exposed to 20% O2 ] to hypoxia leads to decreased ROS  and deserve further investigation in other cell lines. We found that exposure of WT cells  increasl et al. . We alsoWe also observed that the specific Oxidative stress marker; total protein carbonyl content and Lipid peroxidation levels were markedly increased both in Hypoxia and in Cocl2 exposed cells compared to controls. Antioxidant (A2P) and HIF-1a inhibition (ECM) cells showed low Lipid peroxidation levels compared to Hypoxia and in Cocl2 exposed cells. Total Protein carbonyl (PC) content was also reduced with A2P treatment in Cocl2 exposed cells (Cocl2 + A2P), but these PC levels were not significantly changed in hypoxia + A2P and hypoxia + ECM exposed cells. The decreased anti-oxidant markers SOD and Catalase levels were significantly increased in A2P treated myocardium cells compared to Hypoxia and Cocl2 exposed cells. While, Hif1 inhibition (1%O2 + ECM) increased the cells total SOD levels compared to Hypoxia cells; however, Catalase levels were not significantly increased to control levels with HIF-1a inhibition (1%O2 + ECM). This difference can likely be attributed to differences in technique (knockout versus inhibition with ECM).increase in HIF-1\u03b1 mRNA and protein expression during normoxic conditions  decreases . This makes sense and is consistent with other published findings . Howevernditions . In H9c2nditions , which m2 and prolyl hydroxylase (PHD) inhibition in melanoma cells  but significantly reduced [Ca2+]i in the setting of hypoxia, which might be protective. Given the role of [Ca2+]i in cell death as well as the possible role of moderate increases in [Ca2+]i on preconditioning, further investigation seems warranted. We are not aware of any reports of anti-oxidants impacting [Ca2+]i in the setting of hypoxia, although there is evidence that antioxidants improve skeletal function during exposure to hypoxia , not using a separate control siRNA is possibly a limitation. The data are shown in N = two experiment only, this is also a limitation because of lack of enough samples].This study has several limitations. First, it was a study of cell lines, which means that the observed results are not directly applicable to intact organisms. Second, we used only one cell type (H9c2), which we selected because of the importance of the myocardium in long-term clinical outcomes in humans exposed to stress. Third, we only tested one anti-oxidant agent and one HIF-1\u03b1 inhibitor. Though, we used pre-designed specificity-enhanced SignalSilenceOur results pose several questions that need answers through further work - how are antioxidants  able to exert an impact on HIF-1\u03b1 protein expression without affecting detectable ROS levels? How ATG5 is upregulated despite the fact that beclin-1 is not upregulated in H9c2 cells exposed to hypoxia? How does beclin-1 mediate ROS production independent of autophagy? Involvement of PINK1/Parkin pathway in hypoxia-induced autophagy, studies with specific autophagy inhibitors , in H9c2 cells. In addition, are moderate intracellular calcium increases during hypoxia exposure truly mediated by HIF-1\u03b1 and upregulation of Cav 1.2 and if so, does this protect against cell death? What is the role of SERCA, PLN, Na + /Ca2 + exchangers, and plasma membrane Ca2 + ATPase on H9c2 cell\u2019s intracellular Ca2 + levels in Hypoxia?++, increases ATG5 and autophagy through a beclin-1 independent mechanism, and reduces ATP and ROS production in H2c9 cells.Though additional confirmatory studies are needed, in the setting of hypoxia, HIF-1\u03b1 upregulates both Cav 1.2 and 1.3 expression that results in an increase in intracellular CaThe original contributions presented in this study are included in the article/HO: study design, the performance of the study, data analysis and interpretation, and manuscript writing and preparation. ML: data analysis and interpretation and manuscript writing. RT: design the study, data analysis and interpretation, and manuscript writing and preparation. All authors contributed to the article and approved the submitted version.The 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": "Ilex aquifolium L. is a paramount species which is exceptionally suitable for studying phenotypic variability and plasticity through the assessment of morphological, physiological, biochemical and genomic features with respect to acclimation and/or adaptation efficiency. The current study is focused on four insular populations of Ilex aquifolium from Eastern Europe , and presents an initial evaluation of phenotypic variability in order to conclude our research on phylogenetic relationships and phytochemical profiles, including several descriptive and quantitative morphological traits. Taken together, the data from different methods in this paper indicate that the Bulgarian and Romanian populations can be distinguished from each other and from Serbian and Hungarian populations, while the latter show a higher level of resemblance with regards to their quantitative morphological traits. It is likely that these morphological traits are determined through some quantitative trait loci implicated in stress responses generated by light, temperature, soil water, soil fertility and salinity conditions that will need to be analysed in terms of their physiological, genomic and metabolomics traits in future studies.Through its natural or cultivated insular population distribution,  Ilex aquifolium L. is an evergreen, dioecious and heterophyllous species with temperate region morphology characteristics that develops well on drained soils ; coumarin\u201412-hydroxyjasmonic acid-12-O-glucoside like and tuberonic acid glucoside; 12-Oxo phytodienoic acid; 12-hydroxyjasmonic acid-12-O-glucoside like tuberonic acid glucoside; trans-melilotoside (trans-glucosyl-2-hydroxycinnamate); adenine and pyridoxine.Interestingly, when the phytochemical profile of leaves was assessed, once again, the BG population featured significant qualitative differences compared to the HU, RO and SR samples .It has been shown that the synthesis of phytochemicals during the secondary metabolism of plants is geo-climate-dependent and could be correlated with a wide range of environmental factors such as light, temperature, soil water, soil fertility and salinity . ModifyiAmaranthus L. [Ilex paraguariensis [Mostly similar flavonoid profiles were identified in all four locations, although their number appeared significantly lower in the BG leaves . It has nthus L. . It is tariensis .2 concentration [There were higher degrees of similarity between the HU and SR phytochemical profiles regarding the presence of amino acids, carboxylic acid, coumarin and terpenoids, and differences in the presence of flavonoids, polyphenols and saponins. The polyphenolic and flavonoid profiles compound numbers were greater in comparison with those for terpenoids, which were decreased in all four locations. These results could be a consequence of an increased COntration ,75. TherTaken together, the molecular validation and phytochemical profile analyses suggest that the BG population is relatively distinguishable from the others, while cultivated HU individuals showed a similar phytochemical profile to the native RO and SR populations.In order to evaluate the distance isolation (IBD) and the environment isolation (IBE), the Mantel test was used. The results did not reveal the existence of a significant correlations between phytochemicals/MTs and climate/geographical data. Similar studies  have shoDescriptive morphological traits and MT set BG/RO apart from HU/SRQuercus ssp.) have been identified: one located east of the Carpathians and another in the south [Ilex aquifolium (with exception of HU population) are relatively distanced from each other, both latitudinal and longitudinally, with north\u2013south distances of 150\u2013300 km. Similar studies on the variation of MTs at a species level, in different populations from the Balkan Peninsula, were performed on the service tree (Sorbus domestica) L., a rare and endangered species [The gradual decline of water, along with cooling over the last 15 million years, has led to a change in the European plant profile. Species from previously widespread wet temperate tertiary forests have found refuge in the Balkan Peninsula as well as in Turkey ,79. Studhe south ,81. It hhe south . The geo species . The resJovibarba heuffelii (Schott) by \u00c1. L\u00f6ve and D. L\u00f6ve showed that some environmental and bioclimatic factors  can influence the morphology of a species, thereby guiding interpopulation differentiation [Studies on ntiation .Recent experiments have shown that the cuticle thickness, stomatal density, shape and density of trichomes are involved in adaptive mechanisms of the leaf to reduce water loss, playing an important role in protecting the cuticles by integrating flavonoid compounds as a response to the light and UV intensity ,85.Ilex aquifolium. Morphological resemblances were observed between the HU and SR populations by the presence of a prominent follicular cuticle  Heynh., [A box plot analysis revealed the continuous distribution of the assessed MT-specific values in an interval range related to the median value of each trait and location . The hig Heynh., . STF had Heynh., ,90.Meanwhile, for the RO population, STL and STF were more determinative, although DTS could also be invoked, but to a lesser extent. Therefore, it seems likely that both STL and STF could be the major environmentally influenced quantitative traits in the RO population.Interestingly, the BG population-specific cluster comprising the variability of all MT seemed not to feature any MTs exerting an increasing or reducing effect on variability. Such an observation would indicate that the BG population is fairly stable, at least in the case of the analyzed MTs, which also means that the climatic conditions do not fluctuate excessively. The stability of the BG population is further substantiated by the scatterplot data, indicating six, the largest number of MT correlations among all the studied populations.Ilex aquifolium species. At each location  following the manufacturer\u2019s protocol. The DNA concentration was quantified by a spectrophotometric method using a NanoDrop-2000 . The primers and methodology for PCR amplification conditions were as follows: rbcLa fw 5\u2032-ATGTCACCACAAACAGAGACTAAAGC-3\u2032 and rbcLa rev 5\u2032-GTAAAATCAAGTCCACCRCG-3\u2032, mastermix DreamTaqPCR (Thermo) with a PCR reaction volume of 25 \u03bcL/sample; PCR conditions: 98 \u00b0C 0.45\u2033, 98 \u00b0C 0.15\u2033, 59 \u00b0C 0.30\u2033, 72 \u00b0C 0.40\u2033, 35 cycles . A modifIlex aquifolium fresh leaves were frozen in liquid nitrogen and ground to obtain a fine powder. The Nihal [he Nihal  extractihe Nihal . The cap2\u2014SF and stem trichome frequency on approximatively 3 mm2 each\u2014STF, as shown in For our morphological analyses, a Quanta 250 FEI scanning electron microscope (SEM) was used. The samples were coated with gold using an AutoAgar sputter-coater. Three gold deposit layers (4 nm thick) in cycles of 10 s were applied to every sample surface. The samples were examined under vacuum using a secondary-electron detector. Measurements were carried out for the following MTs: leaf stomata distance\u2014SD; leaf D-type stomata distance\u2014DTS; leaf stomata length\u2014SL; leaf stomata width\u2014SW; stem trichome length\u2014STL; leaf stomata frequency on approximately 1.5 mmWith the exception of STF, a trait assessed on stem cuticle surface, all the other MTs were studied on the lower/abaxial surface of prickly leaves situated on one-year-old shoots of female individuals. From the mid-zone of every shoot, a leaf was selected and further processed for microscopic examinations.2 and STF per 3 mm2) per individual. Mean values were obtained from 100 measurements, except for DTS, where the averages were calculated from 10 measurements/individual, because the frequency of appearance of these structures was comparatively low.From each location , a 10 individuals were studied, making 40 individuals in total. For each individual, seven morphological traits (MTs) were assessed, from which five  were represented as mean values per character per individual and two (SF and STF) were expressed as absolute values obtained by counts per unit of surface , the assumptions for running ANOVA were not met.We chose to assess the same set of seven quantitative MTs for every studied population, and a great deal of primary data were recorded and statistically analyzed. Right after computing the basic descriptive statistical parameters see , we chosStatistical analyses were performed on the 280 datapoints outlined above, for the 10 individuals for each of the four locations, characterizing seven MTs in each individual. Microsoft Excel 2019 and the R language,  version 4.1.2 (1 November 2021)  were usep-value cut-off of 0.05 was used to assess significance and the Benjamini-Hochberg FDR approach [p-value adjustment for multiple comparisons in the Dunn\u2019s test.The Kruskal-Wallis test was performed using the built-in R function, kruskal.test, while the follow-up Dunn\u2019s test for pairwise comparisons was run using the dunn_test function from the R rstatix package v.0.7.0 . For botapproach  was applFinally, a Mantel test based on Pearson\u2019s product-moment correlation was performed to see if there was a significant correlation between the two distance matrices used for generating dendrograms corresponding to the phytochemical data and MTs, respectively. Dendrograms were generated for both the MT data and the biochemical data, by first computing the Euclidean distance matrix using the dist function in R, and then performing hierarchical clustering using the R function hclust and plotting the results using the standard plot function in R. The Mantel test was then performed on the two distance matrices using the mantel function from the R vegan package . GeoTIFFIlex aquifolium species. Despite the habitual diversity  and the different geographical locations of the four populations, a molecular analysis proved their affiliation. A phytochemical profile analysis of the leaves distinguished the BG population from the those of RO, HU and SR, while a SEM analysis of descriptive MTs related to leaf and stem indicated three subgroups of morphological diversity: HU-SR, BG and RO. Basic statistical parameters followed by Kruskal-Wallis and Dunn\u2019s tests for pairwise comparisons showed similarity between the populations in HU and SR in most of the analyzed MTs, while in the populations in RO and BG, the values were different compared with the HU-SR subgroup and also between each other (RO-BG). The PCA study showed the four location-specific populations where SR and HU were almost overlapping, while RO and BG were represented as distinct populations in terms of the first two identified PCs.Our comparative multidisciplinary study clearly demonstrates that the four eastern European insular populations belong to the Ilex aquifolium featured differences and similarities whose genetic and environmental dependence require further analysis .Looking at the MT-associated phenotypic variability, it seems obvious that the four isolated populations of Accordingly, the presented data represent an early phase of a comprehensive study intended to run over a longer period of time which will monitor the evolution of phenotypic plasticity, and in particular, seasonal responses to water and light."}
+{"text": "Socially withdrawn children tend to perform poorer academically than their peers. What remains unknown, is the temporal ordering of the two phenomena. Also, substantial gender differences exist in both social withdrawal and academic achievement; thus, it is conceivable that the strength of the relation between them is gendered as well.To investigate cross\u2010sectional correlates and test directional effects of social withdrawal and academic achievement from primary to upper secondary school, and to examine potential gendered effects.n\u00a0=\u00a0845), by means of random intercept cross\u2010lagged panel modelling.Prospective associations were analysed from age 6 to age 14 using biannual teacher ratings of children's social withdrawal and academic achievement in a representative community sample (In boys, increased academic achievement at ages 8 and 12 forecasted decreased social withdrawal 2\u00a0years later, whereas increased social withdrawal at age 10 predicted reduced academic achievement at age 12. No such effects were seen in girls.Social withdrawal and academic achievement are bidirectionally related among boys, but not girls. Results are discussed in light of need\u2010to\u2010belong theory, and practical implications for schools and teachers are illuminated. At T1, 81% of the children were accompanied by their mothers, more than 99% of the children were of Western ethnic origin , and 86% of these children had parents who lived together. Further reading of the procedure, recruitment and sample has been presented elsewhere .Teacher data were collected through questionnaires sent to schools and colleges. Teachers provided information on social withdrawal and academic achievement at all measurement points. Response rates among teachers were as follows: age 6\u00a0=\u00a094.1%, age 8\u00a0=\u00a092.2%, age 10\u00a0=\u00a085.8%, age 12\u00a0=\u00a082.3% and age 14\u00a0=\u00a080.7%. This project was approved by the Regional Committee for Research Ethics, Mid\u2010Norway  to 5 (to a large degree). The total of these responses was computed as the sum score for each child  , 2 (somewhat below grade), 3 (at grade level), 4 (somewhat above grade) or 5 (far above grade), with a maximum score of 15 and a minimum score of 3 (see Table The Teacher Report Form (TRF) from the Achenbach System of Empirically Based Assessment (ASEBA)  and a latent within\u2010person component, which assesses changes in one's own mean level (e.g. AAt3) as a function of changes in one's own levels at a previous time point . Missing values were treated using the full information maximum\u2010likelihood estimator (FIML). Multi\u2010group analyses by sex were evaluated using the Satorra\u2013Bentler Chi\u2010square test  in Mplus 8.1, are presented in Table Weighted descriptive analyses, including mean\u2010level differences between measurement times and bivariate correlations from M2 (21)\u00a0=\u00a035.759, p\u00a0=\u00a0.02, RMSEA\u00a0=\u00a0.029, SRMR\u00a0=\u00a0.034, CFI\u00a0=\u00a0.992, TLI\u00a0=\u00a0.982). At the between\u2010person level, children with higher AA manifested less SW . At the within\u2010person level, no significant prospective cross\u2010lagged associations were detected between AA and SW.A RI\u2010CLPM fitted the data well . However, these effects were not significantly different from the respective non\u2010significant paths at other ages . In contrast, the effects of boys\u2019 AA on later SW varied across age groups . Boys\u2019 AA at age 8 predicted decreased SW from age 8 to age 10 , and a similar effect was observed for higher AA at age 12, predicting attenuated SW from age 12 to age 14 . Inconsistent with these results, boys\u2019 AA at age 6 predicted higher SW at age 8 . None of these effects was significant in girls , AA 6 \u2192 SW 8  and AA 12 \u2192 SW 14 .Multi\u2010group analyses for each gender revealed that among boys at the within\u2010person level, more SW at age 10 predicted decreased AA from age 10 to age 12  to the ninth grade (age 14). Bidirectional effects of this interplay were proposed: (1) increased social withdrawal predicts decreased academic performance and (2) improved academic performance promotes lessened social withdrawal .First, and cross\u2010sectionally, we found that social withdrawal and academic achievement were negatively related at all five measurement points. Of note, such a relationship has been found in several cross\u2010sectional studies  was high across the respective school years, there is a risk that a differentiated pattern of relationships is masked when using computed sum scores. However, although domain\u2010specific hypotheses were not investigated in this study, they may be warranted in future research. Third, this study only assessed the conflicted shyness dimension of the Child Social Preference Scale (Coplan et al., As shown in this study, social withdrawal is entangled with academic achievement both cross\u2010sectionally and prospectively. Our findings show that academic performance promotes social functioning, and vice versa, but primarily among boys, which is in accordance with previous research showing that boys are more vulnerable to social withdrawal than girls (Doyle et al, All authors declare no conflict of interest.Frode Stenseng: Conceptualization; Data curation; Formal analysis; Funding acquisition; Investigation; Methodology; Project administration; Visualization; Writing \u2013 original draft; Writing \u2013 review & editing. Lars Wichstr\u00f8m: Funding acquisition; Investigation; Methodology; Project administration; Writing \u2013 original draft; Writing \u2013 review & editing. Eivind Tingstad: Conceptualization; Project administration; Writing \u2013 original draft; Writing \u2013 review & editing. Vera Skalicka: Conceptualization; Data curation; Formal analysis; Methodology; Writing \u2013 original draft; Writing \u2013 review & editing."}
+{"text": "What are the main findings?The proportion of nonadherence to medication in asthma patients was very high (22.3%), and patients\u2019 behavior was the main reason (44.1%).Clinical pharmacist-led intervention in enhancing medication adherence in asthma patients, increasing treatment efficacy, relieving symptom severity, and reducing the medication burden from the disease.What is the implication of the main finding?Intervention in medication adherence applied strictly following the education and counseling method would be beneficial for every healthcare practitioner to enhance the treatment outcome.Intervention in medication adherence should emphasize young patients and those with limited mobility.p = 0.001). Patient behavior and knowledge were enhanced in the intervention group (p < 0.05). Asthma symptoms were relieved in the intervention group (p = 0.014). Pharmacist-led interventions on adherence rate were higher with OR: 3.550, 95% CI: 1.378\u20139.143, p = 0.009. Conclusions: pharmaceutical intervention could improve medication adherence, treatment efficacy, and the outcome should not be taken for granted; further research should be carried out in this regard.Background: Medication adherence in asthmatic patients enhances the effectiveness of treatments, but some studies in low and middle-income countries still show some limitations. Our study aimed to determine if pharmacist-led interventions could increase medication adherence, improve treatment effectiveness, and relieve symptom severity in outpatients with asthma. Methods: We conducted a randomized, controlled trial on 247 asthmatic outpatients (aged \u2265 16) with a 1:1 ratio randomization at the hospitalization time and repeated after 1-month discharge. The primary outcome was to detect the difference in medication adherence between groups. Adherence was assessed by the general medication adherence scale (GMAS). Data collected by questionnaire was coded and entered into SPSS_20 for statistical analysis; Results: 247 patients  were enrolled . After intervention, the adherence rate was higher among the intervention group than the control group (94.3% vs. 82.8%,  Asthma is a chronic respiratory disease characterized by coughing, wheezing, shortness of breath, and chest tightness. Asthma affects 339 million people around the world . AccordiDrug treatments have demonstrated efficacy in reducing clinical morbidity and mortality. Although access to medication is necessary, it is not sufficient to successfully control the disease . TherefoWe conducted a randomized, controlled trial on 247 outpatients diagnosed with asthma, meeting the inclusion criteria at the Respiratory Clinic, Ba Ria Hospital, from 18 October 2021 to 10 January 2022.Including criteria: outpatients with a diagnosis of asthma, patients aged 16 years or older, patients with old prescriptions at follow-up, and patients with complete administrative and paraclinical information.Exclusion criteria: outpatients who did not agree to participate in the study, could not communicate in Vietnamese, and participated in a study of adherence within 1 year (up to the invitation timeline).1 = 0.491) and 64.5% in the intervention group (p2 = 0.645) [n = 215 with alpha = 0.05, beta = 0.1 (z(\u03b1/2): 1.96 and z\u03b2: 1.282).The sample size was estimated based on the study by Bruce G. Bender et al., 2010. The adherence prevalence in asthma patients after 10 weeks of intervention was 49.1% in the control group (p= 0.645) . We calcData were collected from medical records and patient interviews. All patients who participated in the study had signed informed consent forms. All patients were interviewed twice; the second time was 1 month apart from the first, collecting information about changes after the intervention. Counseling for the intervention group was conducted in the initial interview and 1 month after. The interview contents include (1) Patient information collection form; (2) Knowledge of diseases and drugs; (3) Compliance assessment using the general medication adherence scale (GMAS) ; (4) Asshttps://www.random.org/integer-sets/, accessed on 15 October 2021) decided on 247 random numbers, with the first 124 random numbers being the control group and the remaining 123 being the intervention group. By rearranging each group in ascending order we obtained 2 tables with random numbers. Patients at the enrolled time were numbered cumulatively during the study period. Based on the table we determined whether the patient belonged the intervention group or the control group. Our study designed an open trial with open information for pharmacists and patients.We used a random sampling method by selecting all patients who met the criteria during the study period. The website (Information bias may occur due to: (1) The patients not correctly remembering or understanding the question; (2) The interviewer not fully expressing the question\u2019s meaning during the interview. Remedial measures include: (1) Clearly explaining the research purpose and calling for the support of the patients; (2) Questioning more clearly; (3) Guiding the patient to answer specific and detailed right at the beginning of the interview; (4) Directly explaining when requested; (5) Combining viewing medical records and prescriptions while interviewing patients or family members to understand the patient\u2019s follow-up status and medications.The primary outcome was to assess the pharmacist intervention effectiveness on medication adherence. Patients in the control group received routine care at the Hospital, including counseling and education from doctors, nurses, and pharmacists dispensing drugs. Patients in the intervention group received routine care, counseling, and education from the research team. The education and counseling content for patients of the research team consisted of 4 main parts: (1) Knowledge of diseases and drugs: providing information on the pathophysiology of asthma , exacerbation prevention, differentiating between controller and reliever (reliever), recognition and prevention of drug side effects, correcting inhalation technique errors, and correct storage of inhaler devices; (2) Advise based on the patient\u2019s non-adherence reasons (according to the GMAS), emphasizing the role of adherence to inhaled corticosteroids (ICS) in reducing the risk of hospitalizations and death from asthma, especially for patients with mild asthma (steps 1 and 2), reminders to reduce forgotten medication intake, supplement knowledge, improve inhaler skills, increase adherence motivation. By joining the asthma club, patients could discuss the diseases, the benefits of drug cessation in symptom relief, disease progression, limiting exposure to individual risk factors, vaccination, diet, exercise, asthma prevention during exercise, and measures to prevent COVID-19 infection with doctors or pharmacists (directly or mobile); (3) Pharmacists also advise the patient\u2019s doctors, discuss with the doctor about any encountered side effects, and choose the inhaler and spacer suitable for the patient; (4) Issuing documents for patients, the set of documents was built based on guidelines of The Global initiative against asthma 2020 (GINA) . The intSecondary outcomes were (1) the medication adherence rate; (2) determining the associated risk factors. Adherence was determined based on Atta Abbas Naqvi\u2019s general medication adherence scale (GMAS), which is a Likert scale consisting of 11 questions across 4 levels from 0 to 3 points. Medication adherence was divided into 5 levels, including high adherence (30\u201333 points), good adherence (27\u201329 points), partial adherence (17\u201326 points), low adherence (11\u201316 points), and poor adherence (0\u201310 points). Adherence rates were divided into 2 groups: non-adherence (GMAS scores < 30 points) and adherence (GMAS scores from 30 to 33 points) . The GMAp-value \u2264 0.05.The data were statistically analyzed using SPSS 20.0 software. Descriptive statistics were used for qualitative variables. Ratios, mean, standard deviation (SD), median, and interquartile range (IQR) were used for quantitative variables. The association was assessed by univariate and multivariable logistic regression analysis, OR, 95% confidence interval (CI). We compared ratios using the chi-squared test. The results were statistically significant when the The protocol was approved by the Ethics Committee in Biomedical Research of Can Tho University of Medicine and Pharmacy (No. 02/PCT-H\u0110\u0110\u0110 dated 15 March 2021). All study participants, including patients or the family members of patients, voluntarily signed informed consent to participate with a clear understanding of data collection and use purposes.From 18 October 2021 to 10 January 2022, 247 asthma patients met the criteria out of the 590 that were examined. A total of 343 patients were excluded due to not meeting the criteria or not agreeing to sign the informed consent. A total of 247 asthma patients were randomly divided into 2 groups: intervention (123 patients) and control (124 patients). After 1 month of interviews, the control group had 2 patients drop out (0.8%). Thus, the remaining 245 asthmatics were included in the analysis of drug adherence rates 1 month after counseling .p > 0.05).p > 0.05). After the intervention, the adherence rate in the intervention group was 94.3%, which is significantly higher than the control group (82.8%) (p = 0.005). Patient behavior was the most important reason for non-adherence (31.8%), and this prevalence was significantly lower in the intervention group . Comorbidities and medication burden were also significantly lower in the intervention group (p = 0.004).p = 0.014). There were no patients uncontrolled in the intervention group. Intervention improved patient knowledge due to distinguishing medication and correct inhalation technique, which is a significant difference between the two groups.p = 0.009. The COVID-19 impact reduced medication adherence with OR: 12.2, 95% CI: 3.02\u201350.00, p < 0.001.p < 0.001)  . The con = 0.009 . Non-adh = 0.009 . The pre = 0.009 .n > 215; we had 247 patients, estimated 20% error). Our study showed a significant difference between the intervention group and the control group, clearly showing the role of clinical pharmacists in improving patient adherence, and thereby improving the treatment outcome, relieving symptoms, and reducing the disease burden. In addition, our study assessed several risk factors related to patient adherence. This could be a basis for policy reforms to improve adherence in patients.Our study had a straightforward sample collection process with inclusion and exclusion criteria. All study participants volunteered and benefited from the study; asthmatic clubs were established to improve the adherence rate reduction over the period. The study design closely followed the checklists of CONSORT 2010. Research methods were clearly described and reproducible. The sample size was calculated clearly; therefore, data were specific to the Vietnamese population  [p < 0.05) (p < 0.001) [p < 0.05) [p < 0.001) and much greater than the control group (p < 0.001) [p < 0.001), possibly because the patients in the intervention group had significantly better compliance and correct inhalation technique than the control group (p < 0.001) [After the intervention, the adherence rate of asthma patients in the intervention group was significantly higher than the control group , which i< 0.001) . In the  < 0.05) , which i< 0.001) . The int< 0.001) . There w < 0.05) . The stu< 0.001) . The stu< 0.001) . This ra< 0.001) .p = 0.009 (p > 0.05) (p = 0.001)  . A highe= 0.001) , which iOur study significantly shows the role of clinical pharmacist-led intervention in enhancing medication adherence in asthma patients, increasing treatment efficacy, relieving symptom severity, and reducing the medication burden from the disease. Risk factors related to medication adherence, such as pharmacist-led intervention, in the age group >60 had an increased adherence rate. In addition, the COVID-19 pandemic had a major impact and significantly reduced the adherence. These data could be the basis for policy reforms to improve the adherence rate. In addition, medication adherence intervention should emphasize young patients and those with limited mobility . Therefore, the intervention will be beneficial for every healthcare practitioner to apply if strictly following the education and counseling content."}
+{"text": "The valve was actuated by PBS solution, effectively preventing evaporation of the solution from the micro- and nanochambers and allowing the assay to be performed over a long period of time. The hydrolysis rates of \u03b2-D-galactosidase , kcat, were decreased according to the decrease of the chamber size, and the overall tendency seems to be symmetrically related to the specific surface area of the chambers even under the prevented condition of non-specific adsorption. The spatial localization of the protons in the chambers, which might could affect the dissociation state of the proteins, was also investigated to explain the decrease in the hydrolysis rate. The developed chamber system developed here may be useful for artificially reproducing the confined intracellular environment and molecular crowding conditions.Arrays of small reaction containers, ranging from 624 femtoliters (10 Therefore, a three-dimensional confinement such as liposomes is essential for accurate measurements in enzymology3. If we focus on an outer boundary of a small reaction space containing a single enzyme molecule, it can be classified into three systems: liquid\u2013liquid, liquid\u2013solid, and liquid\u2013air systems. The liquid\u2013air system is well known as an airborne particle but is not suitable for this purpose due to its fast and random movement and fast and easy evaporation. Therefore, liquid\u2013liquid and liquid\u2013solid systems have been mainly proposed as a reaction container and used for single molecule enzymology.Methodological developments in microscopy and related techniques have provided new insights into enzymology at the level of a single enzyme molecule to understand what causes static heterogeneity and the dynamic fluctuation arise from and how they affect the activity. For this implementation, different methodological approaches have been proposed to isolate a single enzyme molecule with enough substrates to observe the activity. A simple approach has been carried out using highly diluted enzyme solution in capillary electrophoresis (CE)4. The activities of individual \u03b2-D-galactosidase  encapsulated in 14\u201315\u00a0\u00b5m diameter droplets with a fluorogenic substrate were measured. The emulsion-defined femtoliter droplet format, equivalent to 10 cubic microns, gave rise to the idea of the next-generation genome sequencing and made a major contribution to biology5. However, standard emulsification methods produce droplets with a broad size distribution and spontaneous fusion/fission occurs. To suppress the broad size distribution and improve the long-term stability, an amphiphilic molecule (a surfactant) is added. Accurate measurements of a single enzyme activity become increasingly difficult because the surfactant can cause conformational changes on the enzyme as well as activity changes of water. Another approach to liquid\u2013liquid systems is liposomes, which consist of unilamellar vesicles ranging in size from 100\u00a0nm7 to a few \u00b5m9 in diameter and provide the more biologically relevant environment. An ultimate ultra-small biologically relevant confinement is the viral capsid, which has an internal diameter of a few nm, equivalent to a few zL, and a single horseradish peroxidase enzyme is encapsulated inside the capsid and its activity is studied10. These simple approaches to mimic the interior of cells or viruses are very useful for understanding not only in vivo enzyme kinetics but also the condensation of biomolecules in living cells. Biomolecular condensates in cells have demonstrated a range of diverse cellular functions through associative and segregative phase transitions, e.g. transcription regulation11 and protein circuit modulation12, the methodology at micro- to nanoscale enzymology could offer a variety of options to focus on their research objectives through easy size and time control13.The first approach to a liquid\u2013liquid system was demonstrated using droplets of a water-in-oil emulsion obtained with a nebulizer consisting of a capillary tube of approximately 20\u00a0\u00b5m diameter14. This approach, which can be categorized as a liquid\u2013solid system, has been widely applied to single-molecule enzymology through improvements in plate materials and encapsulation methods. Arrays of small reaction containers, ranging from femtoliter (10\u201315\u00a0L) to attoliter (10\u201318\u00a0L), for single-molecule enzymology have been fabricated using optical fiber bundles16, polydimethylsiloxane (PDMS)18, and fused silica slides20. In this liquid\u2013solid system, individual enzyme molecules are encapsulated in the small reaction containers using a dilute solution, so that the number of reaction containers is much greater than the number of enzyme molecules present. The most critical issue in this system is how to seal the liquid to the solid surface without leaking and evaporating the liquid during the experiments. Although it is difficult to achieve perfect containment with homogeneous materials such as quartz on a quartz plate, flexible and self-adhesive polymers such as PDMS are suitable materials. Femtoliter chamber arrays have been developed on top of optical fiber bundles sealed by mechanical force with silicone gaskets22, and later an oil sealing method has been applied to ensure a tight seal and avoid evaporation of the reaction solution23. Similar oil sealing approaches have also been carried out for PDMS chambers24, but the various problems associated with the liquid interface mentioned above cannot be avoided.On the other hand, Yeung et al. challenged to fabricate small chamber arrays consisting of about 135\u00a0fL, 5\u201312\u00a0\u00b5m in diameter and 4\u20136\u00a0\u00b5m in height, on thin fused-silica glass, and then, tightly attached to flat quartz plates for encapsulation of single enzyme molecules20 and the air-operated valve25, we operated the pneumatic valve with PBS solution to prevent the solvent evaporation even during prolonged observation and demonstrated a single \u03b2-gal assay in different chamber sizes ranging from 10\u00a0\u00b5m to 800\u00a0nm in diameter, which was expected to provide more experimentally accurate and reproducible values of \u03b2-gal activity at the single molecule level were expected to obtain compared to our previous work26. The reusability of the device could contribute to accurate and reproducible measurements that require delicate handling. The main reason for choosing \u03b2-gal as a target enzyme was its appropriate turnover number to observe  and its robustness to the surrounding environment especially oxygen compared to reductases such as diaphorase28. The proton concentration in these small chambers, which indicates the location of the proton, could affect the hydrolysis reaction and was therefore investigated using a pH-sensitive fluorescent dye, Carboxy SNARF-1. The developed solid-surface enclosed system automatically acquires a long-term observation data automatically from femtoliter to attoliter scale chambers and can be simplify the discussion on the interfacial molecular interaction.Here, we used a pneumatic valve to confine individual enzyme molecules in small reaction containers ranging in volume from femtoliter to attoliter to study the fundamental enzymology in confinement. Although similar approaches have been demonstrated with the dimple machine\u03b2-D-galactosidase , fluorescein, 2-mercaptoethanol were purchased from FUJIFILM Wako Pure Chemical Corporation . A phosphate buffer  was purchased from Sigma-Aldrich, Japan . The basic assay solution was prepared by mixing \u03b2-gal and FDG  in a 100-mM phosphate buffer (pH 7.5) containing 1 mM MgCl2 and 0.2% (v/v) 2-mercaptoethanol. Fluorescence intensity was monitored using a fluorescence spectrophotometer  at an excitation/emission wavelength of 490\u00a0nm/520\u00a0nm and a temperature of 30\u00a0\u00b0C.FDG, 29. The master moulds of the micro- and nanochamber patterns were fabricated on the silicon substrate by electron-beam (EB) lithography and Bosch etching; the fabrication process is shown in Fig.\u00a0The polydimethylsiloxane (PDMS)-based microfluidic devices were fabricated using standard soft lithography methods\u03b2-gal and FDG was introduced into the fluid channel from the reservoir. The pneumatic valve enclosed the assay solution in the micro- and nanochambers by applying air pressure to the control channel, as shown in the supporting movie. The spontaneous adhesion of the PDMS ensured the perfect sealing of the solution in the micro- and nanochamber. The enclosed micro and nanochamber device was placed on the stage of the total internal reflection fluorescence microscope  that was equipped with an argon-ion laser (488\u00a0nm), after which the fluorescence in the microchambers was observed through high magnification objectives  under evanescent wave illumination. All the observations were performed at 30\u00a0\u00b0C using a stage-top incubator . Fluorescence images were captured by an EB-CCD camera 30 coupled to an image intensifier unit  and analysed using image analysis software .The surface of the cover slip was spin-coated with uncured PDMS for 60\u00a0s at 3000\u00a0rpm. The PDMS was then cured for several hours at room temperature (20\u201325\u00a0\u00b0C). A mixed solution of To suppress non-specific adsorption on the PDMS surface, 2-methacryloyloxyethyl phosphorylcholine polymer  and poly(N-hydroxyethyl acrylamide)  were used for the PDMS-based surface coating.A pH indicator, 5-(and-6)-carboxy SNARF-1 dye, was purchased from Thermo Fisher Scientific K.K. . To measure the pH of the solution encapsulated in the micro- and nanochambers, the ratio of the fluorescence intensities of the dye at two emission wavelengths, 580\u00a0nm and 640\u00a0nm, excited at 488\u00a0nm, was used by a confocal microscope  was used for quantitative determinations of the pH.33, but the permeability of water or water-soluble substrates used in this study has not been demonstrated. Migration of water or solutes may be possible but the rate should be much slower compared to the measurement time in this experiment and the influence could be ignored. Supporting movie \u03d5:9.1\u00a0\u00b5m\u2009\u00d7\u2009H:6.3\u00a0\u00b5m) and no fluorescein leakage was observed. Therefore, the following enzymology experiments were performed using the valve-equipped micro- and nanochambers.As shown in Fig.\u00a026, adsorption of enzymes encapsulated in the chambers to the chamber surface is critical in measurements for hydrolysis rate measurements. The hydrophilicity of the original PDMS surface is relatively high and causes denaturation of protein molecules including enzymes through hydrophobic interactions34. Poly(2-methacryloyloxyethyl phosphorylcholine-random-n-butyl methacrylate) (MPC polymer) and poly(N-hydroxyethylacrylamide) (PHEA), shown in Fig.\u00a0\u03b2-gal, FITC-BSA was used to trace the adsorption behavior because BSA is widely used in biochemistry and molecular biology experiments. As shown in Fig.\u00a0As we described in the previous study\u03b2-gal was measured by UV\u2013VIS spectrophotometer under different concentrations of substrate, FDG, concentrations as ensemble-averaged enzyme kinetics. The hydrolysis reaction of \u03b2-gal consists of two steps, FDG to FMG and FMG to fluorescein, and the first hydrolysis rate is known to be significantly slower than the second hydrolysis rate. Therefore, the first hydrolysis rate is a rate-limiting step and corresponds to the observed hydrolysis rate. In this study, this apparent hydrolysis rate was assumed to be the hydrolysis rate, kcat. The Michaelis\u2013Menten constant KM and the hydrolysis rate kcat were calculated according to previous methods35. The measurements, as shown in Figure KM\u2009=\u200956.5\u00a0\u00b5M and kcat\u2009=\u200955.1\u00a0s\u20131, which is three times higher than our previous reports of 18.0\u00a0s\u20131. Quantitative analysis of an enzyme activity is a sensitive experiment, and often performed by the same enzyme from different batches is assayed at different times, so a large inter-laboratory variation of the absolute value was often observed. Even in a single run, a large coefficient of variation exceeding 100% has occasionally been observed36. Experimental imperfections, such as slight differences in temperature and buffer concentration, could be responsible for the variable data. However, our focus was on the effect of reducing the size of the reaction space on the enzyme activity and not on the absolute value of the activity, and the following chamber experiments were performed with great care to suppress the variation.The time course of the fluorescence intensity of fluorescein produced by \u03b2-gal kinetic assay. The concentration of FDG and \u03b2-gal in the phosphate buffer and the incubation time were optimized according to the chamber sizes to capture the single \u03b2-gal molecule with enough FDG to maximize the hydrolysis rate. The typical three results of the single enzyme assay using 624-fL, 61-fL, and 270-aL chambers are shown in the main figures and the remainder are shown in the supplemental figures. To avoid photobleaching of the product, fluorescein, an excitation light shutter was kept closed except for the fluorescence image acquisition time. Crosstalk of the fluorescence light could be minimized by optimizing the incubation time and the excitation laser power even in the smallest 270-aL nanochamber array as shown in Fig.\u00a0Twelve different sizes of micro- and nanochambers, ranging from 270 aL(\u03d5:800\u00a0nm\u2009\u00d7\u2009H:600\u00a0nm) to 624 fL(\u03d5:10\u00a0\u00b5m\u2009\u00d7\u2009H:10\u00a0\u00b5m), were used for a single 17, the increase in fluorescence intensity can be quantified depending on the number of enclosed \u03b2-gal. The occupancy probability of the number of enclosed \u03b2-gal, X, can be assumed to follow the Poisson distribution expressed in the following equation;\u03bb is the expected number of enzymes trapped in the chamber and N! is the factorial of the probability mass function of X. When the occupancy distribution approached to the fitted line at \u03bb\u2009=\u20091, the observed ratio of chambers at the given concentration of \u03b2-gal confirmed the trapping of \u03b2-gal in the microchambers at the single enzyme level. The specific mathematical procedures are the followings: Based on the applied \u03b2-gal concentration, the occupancy probability of the number of encapsulated \u03b2-gal in each chamber, X, was calculated, assuming that loading process into the chambers followed the Poisson distribution. These probabilities yielded the expected numbers containing 0, 1, 2, and 3 of \u03b2-gal and were plotted as shown in Figs.\u00a0\u03b2-gal molecules, assuming a Poisson-distributed loading of \u03b2-gal in the microchambers. Since the fluorescence increase data were classified as different numbers of \u03b2-gal molecules, the hydrolysis rates per a single \u03b2-gal molecule were calculated by reference to the fluorescein standard curve.Three typical assay results are shown in Figs.\u00a0 kcat, of a single \u03b2-gal in 624-fL, 61-fL, and 270-aL chambers were 45.9\u2009\u00b1\u20097.5\u00a0s\u20131, 13.0\u2009\u00b1\u20090.5\u00a0s\u20131, and 0.067\u2009\u00b1\u20090.0038\u00a0s\u20131, respectively. Considering the hydrolysis rates in the bulk experiment, kcat\u2009=\u200955.1\u00a0s\u20131, the largest chamber size of 624 fL gave the similar hydrolysis rates but the other chambers showed that smaller chambers decreased the \u03b2-gal activity as shown in Fig.\u00a038. However, all the inhibition experiments were performed with galactose concentrations ranging from a few to several hundred mM in bulk. In contrast, the current single \u03b2-gal assay system typically uses 200\u00a0\u03bcM of FDG, and the product galactose should be less than this concentration even when the enzymatic reaction is complete. In addition, the total assay time was less than 5 min after mixing the \u03b2-gal and FDG. The number of FDG in the microchamber is more than 7.5\u2009\u00d7\u2009104 even in the smallest 510\u00a0aL chamber. If \u03b2-gal works as 50\u00a0s-1, which is a comparable activity to the bulk system, it will take 25\u00a0min to consume all the FDG and fill the microchamber with galactose. Therefore, we believe that the majority of the microchamber space is occupied by FDG  and that the inhibitory effect of the product, galactose, does not need to be considered during the experimental period of about 5\u00a0min.The hydrolysis rates,\u03b2-gal and FDG on the PDMS chamber surface and the accurate measurements were not achieved. Therefore, to prevent non-specific adsorption on the surface, PHEA was dynamically coated on the PDMS surface and a single \u03b2-gal activity assay was performed. As shown in Fig.\u00a0., the water structure and dynamics could influence the conformation and the function (net hydrolysis rate) of \u03b2-gal in the nanoporous structures of a silicate matrix with a mean pore diameter of 33\u2009\u00b1\u20092\u00a0nm40. Compared to the reaction space in the nanopore, our chamber system has more than 100 times the volume and appears to have less or no influence on \u03b2-gal conformation by the water structure at the chamber surface. However, if 10\u201320\u00a0nm from the chamber surface could affect the water structure, the diffusional approach of \u03b2-gal to the vicinity of the chamber surface could change the hydrolysis rate. The residence time near the chamber surface will increase according to the chamber volume reduction, so the downward tendency of \u03b2-gal hydrolysis rate might be reasonable. In addition, Tsukahara et al. reported faster proton transfer in the 10\u2013100\u00a0nm range from the quartz surface, suggesting a long-range influence on water structure than that is originally expected in nanoporous structures41. Therefore, we attempted to investigate how the PDMS surface affects the water structure inside the chambers, including proton association and dissociation at the plasma-treated PDMS surface.Another possible reason could be the non-specific adsorption of R is the ratio \u03bb1 (568\u00a0nm) and \u03bb2 (616\u00a0nm), and the subscripts A and B represent the limits at the acidic and basic endpoints of the titration, respectively. Calibration was performed using a dual emission ratio with \u03bb1\u2009=\u2009568\u00a0nm and \u03bb2\u2009=\u2009616\u00a0nm excited at 488\u00a0nm. As shown in Fig.\u00a011. Our micro- and nanochamber system will contribute to the pursuit of biological mechanisms in the confined spaces as an alternative method to liquid droplet systems42 and mesoporous systems43.To evaluate the \u201cfree\u201d proton in the chambers, the spectrum changes of the pH indicator, Carboxy SNARF-1, encapsulated in the chambers were observed. As shown in Fig.\u00a0\u03b2-gal molecules in the microchambers was assumed to follow a Poisson distribution. However, the encapsulation process might not occur in a completely independent manner without some specific or biased chemical interaction between the \u03b2-gal molecules and the PDMS surface. If the encapsulation process followed non-Poisson distribution, such as sub-Poisson or super-Poisson distribution, the obtained hydrolysis rate and the above interpretation would be changed. Therefore, with this opposite approach, an accurate comparison of the statistical interpretation and the experimental results could provide insight into the undetected interaction in this experimental system.Throughout this entire experiment, the encapsulation probability of \u03b2-gal to about one tenth depending on the reaction space. Since the PHEA-coated chamber surface could not prevent the kinetic tendency, non-specific adsorption of \u03b2-gal, substrate and product on the large specific surface area may not be the main reason. Another possibility was the silanol group of the PDMS surface dissociation, which could affect the proton localization in the solution and change the dissociation state of \u03b2-gal. The optical resolution was not sufficient to prove the hypothesis, but the micro- and nanochamber array system showed quantitative results even in such a small space from micro- to nanometer cubic scale. The proposed experimental system will help to mimic and analyze recent challenging topics such as liquid\u2013liquid phase separation and biomolecular condensates in a cell44.In this work, a pneumatic valve-based confinement system for micro- and nanochamber arrays was developed to provide technically simple and reproducible single enzyme assay. Using the system, we have experimentally demonstrated that the diffusion-limited confinement space, from micrometer to nanometer cubic, reduced the hydrolysis rates of Supplementary Information 1.Supplementary Information 2.Supplementary Video 1."}
+{"text": "Hypoxic\u2013ischemic encephalopathy (HIE) refers to cerebral hypoxic\u2013ischemic injury caused by asphyxia during perinatal period, which is one of the important causes of neonatal death and sequelae. Early and accurate diagnosis of HIE is of great significance for the prognostic evaluation of patients. The purpose of this study is to explore the efficacy of diffusion-kurtosis imaging (DKI) and diffusion-weighted imaging (DWI) in the diagnosis of early HIE.Twenty Yorkshire newborn piglets (3\u20135\u00a0days) were randomly divided into control group and experimental group. DWI and DKI scanning were performed at timepoints of 3, 6, 9, 12, 16, and 24\u00a0h after hypoxic\u2013ischemic exposure. At each timepoint, the parameter values obtained by each group scan were measured, and the lesion area of the apparent diffusion coefficient (ADC) map and mean diffusion coefficient (MDC) map were measured. . Then, we completely removed the brain for pathological examination, and observed the state of cells and mitochondria in the ADC/MDC matching area , and the mismatch area (the area around the lesion).In the experimental group, the ADC and MDC values decreased with time, but the MDC decreased more significantly and the change rate was higher. Both MDC and ADC values changed rapidly from 3 to 12\u00a0h and slowly from 12 to 24\u00a0h. The MDC and ADC images showed obvious lesions at 3\u00a0h for the first time. At this time, the area of ADC lesions was larger than that of MDC. As the lesions developed, the area of ADC maps was always larger than that of the MDC maps within 24\u00a0h. By observing the microstructure of the tissues by light microscopy, we found that the ADC and MDC matching area in the experimental group showed swelling of neurons, infiltration of inflammatory cells, and local necrotic lesions. Consistent with the observation under light microscope, pathological changes were observed in the matching ADC and MDC regions under electron microscopy as well, including collapse of mitochondrial membrane, fracture of partial mitochondrial ridge, and emergence of autophagosomes. In the mismatching region, the above pathological changes were not observed in the corresponding region of the ADC map.DKI\u2019s characteristic parameter MDC is better than ADC (parameter of DWI) to reflect the real area of the lesion. Therefore, DKI is superior to DWI in diagnosing early HIE. Hypoxic\u2013ischaemic encephalopathy (HIE) refers to cerebral hypoxic\u2013ischaemic damage caused by asphyxia during the perinatal period, which is one of the most important reasons of neonatal death and sequelae. Therefore, an early accurate diagnosis of HIE has important practical significance for the choice of treatment plan and evaluation of prognosis \u20134. The dn\u2009=\u20095; 3 male, 2 female) and experimental group , which came from the Dalian Hua-qiao Chinese Pig Farm. The experimental group was anesthetized with isoflurane, ligated bilaterally the common carotid arteries, and then suture the incision. After resuscitation, the piglet was placed in a tank, and a mixed gas (2\u00a0L/min) consisting of 4% oxygen and 96% nitrogen was delivered for 30\u00a0min. In the control group, only skin incision and blood vessel separation were performed, and bilateral common carotid artery ligation and hypoxia were not performed. Gentamicin was given 3\u00a0h before and after the operation to prevent infection.Twenty healthy Yorkshire piglets  were randomly divided into control group , and scanning time\u2009=\u200995\u00a0s. The DKI scanning parameters were TR/TE\u2009=\u20094500\u00a0ms/min, FOV\u2009=\u2009220\u2009\u00d7\u2009220\u00a0mm, matrix\u2009=\u2009128\u2009\u00d7\u2009128, NEX\u2009=\u20092, and number of layers/layer thickness/layer spacing\u2009=\u200918/3.0\u00a0mm/0.5\u00a0mm. The direction of the diffusion-sensitive gradient field was 20, there were three b values in each direction , and the scanning time was 350\u00a0s. An anchor made of cardboard holds the piglet in a prone position. Under anesthesia, the experimental and control groups underwent MR scanning at 3, 6, 9, 12, 16, and 24\u00a0h.Using GE 3.0-T Discovery MR750w scanner and 32-channel magnetic head coil, using coronal scanning, T1WI, T2WI, DWI and DKI were obtained. The DWI scanning parameters were repetition time (TR)/echo time (TE)\u2009=\u20094500\u00a0ms/min, field of view (FOV)\u2009=\u2009220\u2009\u00d7\u2009220\u00a0mm, matrix\u2009=\u2009128\u2009\u00d7\u2009128, number of layers/layer thickness/layer spacing\u2009=\u200918/3.0\u00a0mm/0.5\u00a0mm, two b values in each direction  was placed, where MK, ADC, and MDC values changed most significantly and in the corresponding area of the control group. Image J software (https://imagej.nih.gov/ij/) was used to measure the lesion area at the same level on ADC and MDC images at various timepoints, and the average value was obtained after three measurements. The above operations were performed by two experienced radiologists. A third radiologist examined the results.We used FuncTool on the GE AW4.7 workstation to post-process all raw data. Parameter images related to DKI and DWI can be obtained at the same time. Parameter maps of MK, MDC, ADC, axial kurtosis (AK), and radial kurtosis (RK) were obtained. In the experimental group, the region of interest  for observation. Two experienced pathologists evaluated histological changes and took pictures of typical lesions.After the 24\u00a0h scan, the inhaled anesthesia experimental group and control group were fixed on the operating table in a prone position. The brain tissue was completely removed by craniotomy. According to the ADC and MDC images, the lesion layer was measured with a ruler starting from the edge of the brain tissue, and the ADC and MDC matching and mismatching areas were measured, respectively. The layer corresponding to the match or mismatch areas were cut with a sterile blade into a layer with a thickness of about 3\u00a0mm and fixed in a 4% paraformaldehyde solution. After conventional dehydration, xylene transparency, and paraffin embedding, the sections were observed by light microscope. ADC/MDC matching and mismatching regions were taken from brain tissue with a volume of about 1\u00a0mmt test was used to compare the percentage changes of different indicators within and between groups. All comparisons were made using the least significant difference method after the fact. P\u2009<\u20090.05 was considered statistically significant.All statistical analyses were performed using SPSS 20.0 software . The normality and variance uniformity of the obtained parameter values were tested, and the data were expressed as mean\u2009\u00b1\u2009standard deviation. Repeated measures analysis of variance were used to calculate the difference in parameter values between different timepoints and between groups, and the P\u2009<\u20090.05), and the signals of MDC and ADC maps were relatively homogeneous. However, there was no significant change on T2WI within 24\u00a0h  were significantly increased in the experimental group, whereas the MDC and ADC values were reduced Table . The chaInterestingly, we found that the parameters of DKI (MDC and MK) changed rapidly from 3 to 12\u00a0h, while the parameter DWI (ADC) changed slowly during this period. However, both DKI parameters and DWI parameters changed slowly within 12\u201324\u00a0h   Table . The MDC Table . Under an optical microscope, compared with areas in the control group, some of the glial cell disintegration and necrosis areas were observed in the edge of lesions on the MDC/ADC matching areas of the experimental group. The surrounding glial cells were swollen, the intercellular space was widened and vasodilatation was observed locally Fig.\u00a0. In addiOur study showed that with the development of HIE, the ADC and MDC values gradually decreased, whereas the MK, AK, and RK values gradually increased. In addition, the lesions continued to deteriorate, with a large change rate mainly in the first 12\u00a0h . It revealed that HIE progresses faster in the first 12\u00a0h, suggesting that intervention measures should be taken as soon as possible. Yang et al. also found that HIE lesions progressed most rapidly in the first 12\u00a0h . It was P\u2009<\u20090.05). Studies have found that in adults with ischemic stroke, the lesion area of the ADC map was 5.1% larger than that of the MDC map [Regarding to the lesion area of the HIE model, DWI and DKI depicted the lesion in the first scan of the experimental group, but conventional MR T2WI) did not show it accurately , this study only compared the parameter MDC (DKI) with the parameter ADC (DWI). Based on previous studies on HIE, we studied only the important sites of HIE prevalence, including regions adjacent to the lateral ventricle and subcortical white matter. In the future, we will conduct studies on other sites, where lesions may occur.We used the newborn piglet as a model to compare the diagnostic ability of DKI and DWI for HIE. Our study revealed that DKI had the advantage of a more realistic image model, which could reflect the changes of brain microstructure more comprehensively and sensitively. Our research will have a broad application prospects in clinical work."
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