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Which test is used to diagnose colour synesthesia? | A standardized test battery for the study of synesthesia. We used synesthetic versions of the Stroop test with colored letters and numbers presented either in the right or the left visual field of thirty-four synesthetes. Assessment of the hemispheric lateralization of grapheme-color synesthesia with Stroop-type tests. | Synesthesia is an unusual condition in which stimulation of one modality evokes
sensation or experience in another modality. Although discussed in the
literature well over a century ago, synesthesia slipped out of the scientific
spotlight for decades because of the difficulty in verifying and quantifying
private perceptual experiences. In recent years, the study of synesthesia has
enjoyed a renaissance due to the introduction of tests that demonstrate the
reality of the condition, its automatic and involuntary nature, and its
measurable perceptual consequences. However, while several research groups now
study synesthesia, there is no single protocol for comparing, contrasting and
pooling synesthetic subjects across these groups. There is no standard battery
of tests, no quantifiable scoring system, and no standard phrasing of questions.
Additionally, the tests that exist offer no means for data comparison. To remedy
this deficit we have devised the Synesthesia Battery. This unified collection of
tests is freely accessible online (http://www.synesthete.org). It consists of a
questionnaire and several online software programs, and test results are
immediately available for use by synesthetes and invited researchers.
Performance on the tests is quantified with a standard scoring system. We
introduce several novel tests here, and offer the software for running the
tests. By presenting standardized procedures for testing and comparing subjects,
this endeavor hopes to speed scientific progress in synesthesia research. |
How does increased GDF15 affect body weight? | In humans, elevated GDF15 correlates with weight loss, and the administration of GDF15 to mice with obesity reduces body weight, at least in part, by decreasing food intake. | Growth differentiation factor 15 (GDF15), a distant member of the transforming
growth factor (TGF)-β family, is a secreted protein that circulates as a 25-kDa
dimer. In humans, elevated GDF15 correlates with weight loss, and the
administration of GDF15 to mice with obesity reduces body weight, at least in
part, by decreasing food intake. The mechanisms through which GDF15 reduces body
weight remain poorly understood, because the cognate receptor for GDF15 is
unknown. Here we show that recombit GDF15 induces weight loss in mice fed a
high-fat diet and in nonhuman primates with spontaneous obesity. Furthermore, we
find that GDF15 binds with high affinity to GDNF family receptor α-like (GFRAL),
a distant relative of receptors for a distinct class of the TGF-β superfamily
ligands. Gfral is expressed in neurons of the area postrema and nucleus of the
solitary tract in mice and humans, and genetic deletion of the receptor
abrogates the ability of GDF15 to decrease food intake and body weight in mice.
In addition, diet-induced obesity and insulin resistance are exacerbated in
GFRAL-deficient mice, suggesting a homeostatic role for this receptor in
metabolism. Finally, we demonstrate that GDF15-induced cell signaling requires
the interaction of GFRAL with the coreceptor RET. Our data identify GFRAL as a
new regulator of body weight and as the bona fide receptor mediating the
metabolic effects of GDF15, enabling a more comprehensive assessment of GDF15 as
a potential pharmacotherapy for the treatment of obesity. |
Which are the best methods for the prediction of circular RNA (circRNA)? | A circRNA prediction software for plants . Circular RNA profile in gliomas revealed by identification tool UROBORUS. Numerous algorithms that are used to detect genome-wide circRNA expression from RNA sequencing data have been developed in the past few years, but there is little overlap in their predictions and no clear gold-standard method to assess the accuracy of these algorithms. MiARma-Seq: a comprehensive tool for miRNA, mRNA and circRNA analysis. Here, we use common RNAseq datasets to scrutinize and compare the output from five different algorithms; circRNA_finder, find_circ, CIRCexplorer, CIRI, and MapSplice and evaluate the levels of bona fide and false positive circRNAs based on RNase R resistance. | MOTIVATION: Circular RNAs (circRNAs) are a poorly characterized class of
molecules that have been identified decades ago. Emerging high-throughput
sequencing methods as well as first reports on confirmed functions have sparked
new interest in this RNA species. However, the computational detection and
quantification tools are still limited.
RESULTS: We developed the software tandem, DCC and CircTest DCC uses output from
the STAR read mapper to systematically detect back-splice junctions in
next-generation sequencing data. DCC applies a series of filters and integrates
data across replicate sets to arrive at a precise list of circRNA candidates. We
assessed the detection performance of DCC on a newly generated mouse brain data
set and publicly available sequencing data. Our software achieves a much higher
precision than state-of-the-art competitors at similar sensitivity levels.
Moreover, DCC estimates circRNA versus host gene expression from counting
junction and non-junction reads. These read counts are finally used to test for
host gene-independence of circRNA expression across different experimental
conditions by our R package CircTest We demonstrate the benefits of this
approach on previously reported age-dependent circRNAs in the fruit fly.
AVAILABILITY AND IMPLEMENTATION: The source code of DCC and CircTest is licensed
under the GNU General Public Licence (GPL) version 3 and available from
https://github.com/dieterich-lab/[DCC or CircTest].
CONTACT: [email protected]
SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics
online. CircRNAs are novel members of the non-coding RNA family. For several decades
circRNAs have been known to exist, however only recently the widespread
abundance has become appreciated. Annotation of circRNAs depends on sequencing
reads spanning the backsplice junction and therefore map as non-linear reads in
the genome. Several pipelines have been developed to specifically identify these
non-linear reads and consequently predict the landscape of circRNAs based on
deep sequencing datasets. Here, we use common RNAseq datasets to scrutinize and
compare the output from five different algorithms; circRNA_finder, find_circ,
CIRCexplorer, CIRI, and MapSplice and evaluate the levels of bona fide and false
positive circRNAs based on RNase R resistance. By this approach, we observe
surprisingly dramatic differences between the algorithms specifically regarding
the highly expressed circRNAs and the circRNAs derived from proximal splice
sites. Collectively, this study emphasizes that circRNA annotation should be
handled with care and that several algorithms should ideally be combined to
achieve reliable predictions. Recent evidence suggests that many endogenous circular RNAs (circRNAs) may play
roles in biological processes. However, the expression patterns and functions of
circRNAs in human diseases are not well understood. Computationally identifying
circRNAs from total RNA-seq data is a primary step in studying their expression
pattern and biological roles. In this work, we have developed a computational
pipeline named UROBORUS to detect circRNAs in total RNA-seq data. By applying
UROBORUS to RNA-seq data from 46 gliomas and normal brain samples, we detected
thousands of circRNAs supported by at least two read counts, followed by
successful experimental validation on 24 circRNAs from the randomly selected 27
circRNAs. UROBORUS is an efficient tool that can detect circRNAs with low
expression levels in total RNA-seq without RNase R treatment. The circRNAs
expression profiling revealed more than 476 circular RNAs differentially
expressed in control brain tissues and gliomas. Together with parental gene
expression, we found that circRNA and its parental gene have diversified
expression patterns in gliomas and control brain tissues. This study establishes
an efficient and sensitive approach for predicting circRNAs using total RNA-seq
data. The UROBORUS pipeline can be accessed freely for non-commercial purposes
at http://uroborus.openbioinformatics.org/. Large-scale RNAseq has substantially changed the transcriptomics field, as it
enables an unprecedented amount of high resolution data to be acquired. However,
the analysis of these data still poses a challenge to the research community.
Many tools have been developed to overcome this problem, and to facilitate the
study of miRNA expression profiles and those of their target genes. While a few
of these enable both kinds of analysis to be performed, they also present
certain limitations in terms of their requirements and/or the restrictions on
data uploading. To avoid these restraints, we have developed a suite that offers
the identification of miRNA, mRNA and circRNAs that can be applied to any
sequenced organism. Additionally, it enables differential expression, miRNA-mRNA
target prediction and/or functional analysis. The miARma-Seq pipeline is
presented as a stand-alone tool that is both easy to install and flexible in
terms of its use, and that brings together well-established software in a single
bundle. Our suite can analyze a large number of samples due to its multithread
design. By testing miARma-Seq in validated datasets, we demonstrate here the
benefits that can be gained from this tool by making it readily accessible to
the research community. The pervasive expression of circular RNAs (circRNAs) is a recently discovered
feature of gene expression in highly diverged eukaryotes. Numerous algorithms
that are used to detect genome-wide circRNA expression from RNA sequencing
(RNA-seq) data have been developed in the past few years, but there is little
overlap in their predictions and no clear gold-standard method to assess the
accuracy of these algorithms. We review sources of experimental and
bioinformatic biases that complicate the accurate discovery of circRNAs and
discuss statistical approaches to address these biases. We conclude with a
discussion of the current experimental progress on the topic. BACKGROUND: Radon is a known human lung carcinogen, whose underlying
carcinogenic mechanism remains unclear. Recently, circular RNA (circRNA), a
class of endogenous non-protein coding RNAs that contain a circular loop, was
found to exhibit multiple biological effects. In this study, circRNA profiles in
mouse lung tissues between control and radon exposure were analyzed.
METHODS: Six mice were exposed to radon at concentration of 100,000 Bq/m3,
12 h/d, for up to cumulative doses of 60 working level months (WLM). H&E
staining and immunohistochemistry of caspase-3 were used to detect the damages
in lung tissue. The lung tissue of control and exposed group were selected for
circRNA microarray study. The circRNA/microRNA interaction was analyzed by
starBase prediction software. 5 highest expressing circRNAs were selected by
real-time PCR to validate the consistency in mouse lung tissue exposed to radon.
RESULTS: Inflammatory reaction was found in mouse lung tissue exposed to radon,
and caspase-3 expression was significantly increased. Microarray screening
revealed 107 up-regulated and 83 down-regulated circRNAs, among which top 30
circRNAs with the highest fold changes were chosen for further analysis, with 5
microRNAs binding sites listed for each circRNA. Consistency of the top 5
circRNAs with the highest expressions were confirmed in mice exposed with 60WLM
of radon.
CONCLUSION: Mouse lung tissue was severely injured when exposed to radon through
pathological diagnosis and immunohistochemical analysis. A series of
differentially expressed circRNAs demonstrated that they may play an important
role in pulmonary toxicity induced by radon. |
What is inhibited by a drug rilotumumab? | Rilotumumab is a fully human monoclonal antibody that selectively targets the hepatocyte growth factor (HGF). It is used for treatment of cancer. | PURPOSE: To evaluate the efficacy, safety, biomarkers, and pharmacokinetics of
rilotumumab, a fully human, monoclonal antibody against hepatocyte growth factor
(HGF)/scatter factor, combined with mitoxantrone and prednisone (MP) in patients
with castration-resistant prostate cancer (CRPC).
EXPERIMENTAL DESIGN: This double-blinded phase II study randomized (1:1:1)
patients with progressive, taxane-refractory CRPC to receive MP (12 mg/m(2) i.v.
day 1, 5 mg twice a day orally days 1-21, respectively) plus 15 mg/kg
rilotumumab, 7.5 mg/kg rilotumumab, or placebo (i.v. day 1) every 3 weeks. The
primary endpoint was overall survival (OS).
RESULTS: One hundred and forty-four patients were randomized. Median OS was 12.2
versus 11.1 months [HR, 1.10; 80% confidence interval (CI), 0.82-1.48] in the
combined rilotumumab versus control arms. Median progression-free survival was
3.0 versus 2.9 months (HR, 1.02; 80% CI, 0.79-1.31). Treatment appeared well
tolerated with peripheral edema (24% vs. 8%) being more common with rilotumumab.
A trend toward unfavorable OS was observed in patients with high tumor MET
expression regardless of treatment. Soluble MET levels increased in all
treatment arms. Total HGF levels increased in the rilotumumab arms. Rilotumumab
showed linear pharmacokinetics when co-administered with MP.
CONCLUSIONS: Rilotumumab plus MP had manageable toxicities and showed no
efficacy improvements in this estimation study. High tumor MET expression may
identify patients with CRPC with poorer prognosis. Rilotumumab is an investigational, fully human, monoclonal antibody
immunoglobulin G2 against hepatocyte growth factor (HGF) that blocks the binding
of HGF to its receptor MET and has shown trends toward improved survival in a
phase 2 clinical trial in gastric cancer. To characterize rilotumumab
pharmacokinetics in patients with cancer and to identify factors affecting the
pharmacokinetics, rilotumumab concentration data from seven clinical trials were
analyzed with a nonlinear mixed-effect model. We found that rilotumumab
exhibited linear and time-invariant kinetics over a dose range of 0.5-20 mg/kg.
Typical systemic clearance and central volume of distribution were 0.184 L/day
and 3.56 L, respectively. Body weight is the most significant covariate, and
sex, cancer type, coadministration of chemotherapeutics, baseline plasma HGF and
tumor MET levels, and renal and hepatic functions did not have an effect on
rilotumumab pharmacokinetics. The concentration-time profiles for the
rilotumumab clinical regimens were projected well with the model. PURPOSE: Rilotumumab is an investigational, fully human monoclonal antibody to
hepatocyte growth factor. In a randomized phase II study, trends toward improved
survival were observed with rilotumumab (7.5 or 15 mg/kg) plus epirubicin,
cisplatin, and capecitabine (ECX) versus placebo plus ECX in
gastric/gastroesophageal junction (GEJ) cancer patients, especially in
MET-positive patients. Here, we quantitatively characterized the longitudinal
exposure-response [tumor growth (TG) and overall survival (OS)] relationship for
rilotumumab.
EXPERIMENTAL DESIGN: Rilotumumab concentrations, tumor sizes, and survival time
from the phase II study were pooled to develop a longitudinal exposure versus TG
model and parametric OS model that explored predictive/prognostic/treatment
effects (MET expression, rilotumumab exposure, relative tumor size). Model
evaluation included visual predictive checks, nonparametric bootstrap, and
normalized prediction distribution errors. Simulations were undertaken to
predict the relationship between rilotumumab dose and OS.
RESULTS: Rilotumumab exhibited linear time-independent pharmacokinetics not
affected by MET expression. The TG model adequately described tumor size across
arms. A Weibull distribution best described OS. Rilotumumab exposure and change
in tumor size from baseline at week 24 were predictive of OS. MET-positive
patients showed shorter survival and responded better to rilotumumab than
MET-negative patients. Simulations predicted a median (95% confidence interval)
HR of 0.38 (0.18-0.60) in MET-positive patients treated with 15 mg/kg
rilotumumab Q3W.
CONCLUSIONS: Rilotumumab plus ECX demonstrated concentration-dependent effects
on OS, influenced by MET expression, and tumor size in gastric/GEJ cancer
patients. These findings support the phase II testing of rilotumumab 15 mg/kg
every 3 weeks in MET-positive gastric/GEJ cancer (RILOMET-1; NCT01697072). Despite improvements in systemic chemotherapy (CT), the prognosis of metastatic
adenocarcinoma of the gastroesophageal junction remains poor. Over the years,
new targeting agents have become available and were tested, with or without CT,
in first or subsequent lines of therapy. The epidermal growth factor receptor
family was targeted with monoclonal antibodies (MoAbs) (trastuzumab, cetuximab,
panitumumab) and tyrosin kinase inhibitors (TKIs) (lapatinib, erlotinib,
gefitinib). Only trastuzumab, in combination with cisplatin and
fluoropyrimidines, significantly improved overall survival (OS) in first-line
therapy (13.8 vs. 11.1 months). Angiogenesis also was targeted with MoAbs
(bevacizumab and ramucirumab); ramucirumab, a vascular endothelial growth
factor-receptor 2 antagonist, enhanced OS in two phase III studies in the first
(9.6 vs. 7.4 months) and subsequent lines of treatment (5.2 vs. 3.8 months),
while the bevacizumab study was negative. TKIs (sunitinib, sorafenib,
regorafenib, apatinib) were tested in this setting in phase II studies in the
second/third line, only showing modest antitumor activity. The hepatocyte growth
factor receptor (MET) was targeted in untreated patients in a phase III trial
with MoAb rilotumumab, with or without CT, but the study was stopped because of
mortality excess in the rilotumumab arm. Mammalian target of rapamycin (MTOR)
pathway inhibition with everolimus was tested in pretreated patients in a
placebo-controlled phase III trial who failed to improve OS (5.4 vs.
4.3 months). In conclusion, considering the modest survival gain obtained
overall, the high cost of these therapies and the quality of life issue must be
primarily considered in treating these patients. AIMS: Rilotumumab is a fully human monoclonal antibody investigated for the
treatment of MET-positive gastric cancer. The aim of this study was to evaluate
the potential pharmacokinetic (PK)-based drug-drug interaction (DDI) between
rilotumumab and epirubicin (E), cisplatin(C) and capecitabine (X).
METHODS: This was a Phase 3 double-blind, placebo-controlled study, in which
rilotumumab, epirubicin and cisplatin were administered intravenously at
15 mg kg-1 , 50 mg m-2 , and 60 mg m-2 Q3W, respectively, while capecitabine was
given orally at 625 mg m-2 twice daily. Rilotumumab PK samples were taken at
pre-dose and at the end-of-infusion from all patients in cycles 1, 3, 5 and 7.
ECX PK samples were taken in cycle 3 from patients who participated in the
intensive PK assessment. ECX PK was assessed by non-compartmental (NCA) analyses
and PK parameters were compared between two arms. Rilotumumab PK was assessed by
comparing the observed rilotumumab serum concentrations with model-predicted
concentrations using a population PK model developed from previous Phase 1 and
Phase 2 studies.
RESULTS: The study enrolled 609 patients. ECX plasma concentrations in the
presence and absence of rilotumumab were similar, as demonstrated by the
geometric mean ratios for Cmax and AUC, which were close to 1.0, suggesting ECX
PK was not affected by co-administration of rilotumumab. The observed
rilotumumab serum concentrations were similar to the values predicted by
population PK modelling on the basis of a prediction-corrected visual predictive
check, indicating rilotumumab exposure was not affected by co-administration of
ECX.
CONCLUSIONS: The results suggest lack of PK-based DDI between rilotumumab and
ECX. BACKGROUND: Dysregulated hepatocyte growth factor/mesenchymal-epithelial
transition (MET) signaling is associated with poor prognosis and resistance to
vascular endothelial growth factor inhibition in metastatic colorectal cancer
(mCRC). We report outcomes from a double-blind, multicenter phase II trial of
the MET inhibitor onartuzumab in combination with mFOLFOX-6 and bevacizumab for
mCRC (GO27827; NCT01418222).
MATERIALS AND METHODS: Patients were randomized 1:1 to receive onartuzumab (10
mg/kg intravenously [IV]) or placebo plus mFOLFOX-6 and bevacizumab (5 mg/kg
IV). Oxaliplatin was given for 8-12 cycles; other agents were continued until
disease progression, unacceptable toxicity, or death. The primary endpoint was
progression-free survival (PFS) in the intent-to-treat (ITT) and MET
immunohistochemistry (IHC) expression-positive populations.
RESULTS: Between September 2011 and November 2012, 194 patients were enrolled.
In September 2013, an interim analysis recommended stopping onartuzumab
treatment due to lack of efficacy. At the time of the final analysis in February
2014, no significant improvement in PFS was seen with onartuzumab versus placebo
in either the ITT or MET IHC-positive populations. An improvement in PFS was
noted in the MET IHC-negative population. Neither overall survival nor response
rate was improved with onartuzumab. The incidence of fatigue, peripheral edema,
and deep vein thrombosis was increased with onartuzumab relative to placebo.
CONCLUSION: Onartuzumab combined with mFOLFOX-6 and bevacizumab did not
significantly improve efficacy outcomes in either the ITT or MET IHC-positive
populations. MET expression by IHC was not a predictive biomarker in this
setting. The Oncologist 2017;22:264-271 IMPLICATIONS FOR PRACTICE: The addition
of onartuzumab to mFOLFOX-6 plus bevacizumab did not improve outcomes in
patients with previously untreated metastatic colorectal cancer in this
randomized, phase II study. Although initial results with onartuzumab were
promising, a number of phase II/III clinical trials have reported a lack of
improvement in efficacy with onartuzumab combined with standard-of-care
therapies in several tumor types. Furthermore, negative study data have been
published for rilotumumab and ficlatuzumab, both of which block hepatocyte
growth factor binding to the mesenchymal-epithelial transition (MET) receptor.
MET immunohistochemistry was not a predictive biomarker. It remains to be seen
if other biomarkers or small molecule inhibitors may be more appropriate for
inhibiting this oncogenic pathway. BACKGROUND: Activation of the mesenchymal-epidermal transition factor (MET)
tyrosine kinase and its ligand, hepatocyte growth factor (HGF), is implicated in
resistance to epidermal growth factor receptor (EGFR) inhibitors. In this phase
1/2 trial, rilotumumab (an anti-HGF antibody) combined with erlotinib was
evaluated in patients with metastatic, previously treated non-small cell lung
cancer.
METHODS: In phase 1, a dose de-escalation design was adopted with rilotumumab
starting at 15 mg/kg intravenously every 3 weeks and oral erlotinib 150 mg
daily. In phase 2, the disease control rate (DCR) (according to Response
Evaluation Criteria in Solid Tumors) of the combination was evaluated using a
Simon 2-stage design. The biomarkers examined included 10 plasma-circulating
molecules associated with the EGFR and MET pathways.
RESULTS: Without indications for de-escalation, the recommended phase 2 dose was
dose level 0. Overall, 45 response-evaluable patients were enrolled (13 with
squamous carcinoma, 32 with adenocarcinoma; 2 had confirmed EGFR mutations, 33
had confirmed wild-type [WT] EGFR, and 7 had KRAS mutations). The DCR for all
patients was 60% (90% confidence interval [CI], 47.1%-71.3%). Median
progression-free survival was 2.6 months (90% CI, 1.4-2.7 months), and median
overall survival was 6.6 months (90% CI, 5.6-8.9 months). Among patients with WT
EGFR, the DCR was 60.6% (90% CI, 46.3%-73.3%), median progression-free survival
was 2.6 months (90% CI, 1.4-2.7 months), and median overall survival was 7.0
months (90% CI, 5.6-13.4 months). Elevated baseline levels of neuregulin 1 were
associated with longer progression-free survival (hazard ratio, 0.41; 95% CI,
0.19-0.87), whereas elevated amphiregulin levels were associated with more rapid
progression (hazard ratio, 2.14; 95% CI, 1.48-3.08).
CONCLUSIONS: Combined rilotumumab and erlotinib had an acceptable safety
profile, and the DCR met the prespecified criteria for success. In the EGFR WT
group, the DCR exceeded published reports for erlotinib alone. High circulating
levels of neuregulin 1 may indicate sensitivity to this combination. Cancer
2017;123:2936-44. © 2017 American Cancer Society. |
Which RNA polymerase II subunit carries RNA cleavage activity? | The eukaryotic transcription factor TFIIS enhances elongation and nascent transcript cleavage activities of RNA polymerase II in a stalled elongation complex. | Ternary complexes of vaccinia virus RNA polymerase containing 3'-OMeGMP-arrested
transcripts were purified by native gel electrophoresis. These complexes resumed
elongation in situ when gel slices were incubated with magnesium and NTPs.
Elongation occurred in the absence of pyrophosphate, suggesting that the
blocking 3'-OMeGMP residue was removed via a novel pathway. We show that
purified elongation complexes contain an intrinsic nuclease activity that
shortens nascent RNA from the 3'-end. RNA cleavage was absolutely dependent on a
divalent cation and was stimulated by CTP. The initial 5' cleavage product
remained associated with the ternary complex and could be elongated in the
presence of NTPs. Multiple stepwise cleavages generated progressively shorter
chains. Purified ternary complexes containing 3'-OH-terminated RNAs also
displayed nuclease activity. Involvement of the vaccinia RNA polymerase subunit
rpo30 in the transcript-shortening reaction is suggested based on sequence
similarity of rpo30 to mammalian protein SII (TFIIS), an extrinsic transcription
factor required for nascent RNA cleavage by RNA polymerase II (Reines, D. (1991)
J. Biol. Chem. 267, 3795-3800). The alpha-amanitin domain or domain f of the largest subunit of RNA polymerases
is one of the most conserved of these enzymes. We have found that the C-terminal
part of domain f can be swapped between yeast RNA polymerase II and III. An
extensive mutagenesis of domain f of C160, the largest subunit of RNA polymerase
III, was carried out to better define its role and understand the mechanism
through which C160 participates in transcription. One mutant enzyme, C160-270,
showed much reduced transcription of a non-specific template at low DNA
concentrations. Abortive synthesis of trinucleotides in a dinucleotide-primed
reaction proceeded at roughly wild-type levels, indicating that the mutation did
not affect the formation of the first phosphodiester bond, but rather the
transition from abortive initiation to processive elongation. In specific
transcription assays, on the SUP4 tRNA gene, pausing was extended but the rate
of RNA elongation between pause sites was not affected. Finally, the rate of
cleavage of nascent RNA transcripts by halted mutant RNA polymerase was
increased approximately 10-fold. We propose that the domain f mutation affects
the transition between two transcriptional modes, one being adopted during
abortive transcription and at pause sites, the other during elongation between
pause sites. Budding yeast RNA polymerase III (Pol III) contains a small, essential subunit,
named C11, that is conserved in humans and shows a strong homology to TFIIS. A
mutant Pol III, heterocomplemented with Schizosaccharomyces pombe C11, was
affected in transcription termination in vivo. A purified form of the enzyme
(Pol III Delta), deprived of C11 subunit, initiated properly but ignored pause
sites and was defective in termination. Remarkably, Pol III Delta lacked the
intrinsic RNA cleavage activity of complete Pol III. In vitro reconstitution
experiments demonstrated that Pol III RNA cleavage activity is mediated by C11.
Mutagenesis in C11 of two conserved residues, which are critical for the
TFIIS-dependent cleavage activity of Pol II, is lethal. Immunoelectron
microscopy data suggested that C11 is localized on the mobile thumb-like stalk
of the polymerase. We propose that C11 allows the enzyme to switch between an
RNA elongation and RNA cleavage mode and that the essential role of the Pol III
RNA cleavage activity is to remove the kinetic barriers to the termination
process. The integration of TFIIS function into a specific Pol III subunit may
stem from the opposite requirements of Pol III and Pol II in terms of transcript
length and termination efficiency. RNA polymerase III recognizes and pauses at its terminator, an oligo(dT) tract
in non-template DNA, terminates 3' oligo(rU) synthesis within this sequence, and
releases the RNA. The pol III subunit Rpc11p (C11) mediates RNA 3'-5' cleavage
in the catalytic center of pol III during pausing. The amino and carboxyl
regions of C11 are homologous to domains of the pol II subunit Rpb9p, and the
pol II elongation and RNA cleavage factor, TFIIS, respectively. We isolated C11
mutants from Schizosaccharomyces pombe that cause pol III to readthrough
terminators in vivo. Mutant RNA confirmed the presence of terminator readthrough
transcripts. A predomit mutation site, F32, resides in the C11 Rpb9-like
domain. Another mutagenic approach confirmed the F32 mutation and also isolated
I34 and Y30 mutants. Modeling Y30, F32 and I34 of C11 in available cryoEM pol
III structures predicts a hydrophobic patch that may interface with C53/37.
Another termination mutant, Rpc2-T455I, appears to reside internally, near the
RNA-DNA hybrid. We show that the Rpb9 and TFIIS homologous mutants of C11
reflect distinct activities, that differentially affect terminator recognition
and RNA 3' cleavage. We propose that these C11 domains integrate action at the
upper jaw and center of pol III during termination. During gene transcription, the RNA polymerase (Pol) active center can catalyze
RNA cleavage. This intrinsic cleavage activity is strong for Pol I and Pol III
but very weak for Pol II. The reason for this difference is unclear because the
active centers of the polymerases are virtually identical. Here we show that Pol
II gains strong cleavage activity when the C-terminal zinc ribbon domain
(C-ribbon) of subunit Rpb9 is replaced by its counterpart from the Pol III
subunit C11. X-ray analysis shows that the C-ribbon has detached from its site
on the Pol II surface and is mobile. Mutagenesis indicates that the C-ribbon
transiently inserts into the Pol II pore to complement the active center. This
mechanism is also used by transcription factor IIS, a factor that can bind Pol
II and induce strong RNA cleavage. Together with published data, our results
indicate that Pol I and Pol III contain catalytic C-ribbons that complement the
active center, whereas Pol II contains a non-catalytic C-ribbon that is
immobilized on the enzyme surface. Evolution of the Pol II system may have
rendered mRNA transcript cleavage controllable by the dissociable factor
transcription factor IIS to enable promoter-proximal gene regulation and
elaborate 3'-processing and transcription termination. During DNA transcription, RNA polymerases often adopt inactive backtracked
states. Recovery from backtracks can occur by 1D diffusion or cleavage of
backtracked RNA, but how polymerases make this choice is unknown. Here, we use
single-molecule optical tweezers experiments and stochastic theory to show that
the choice of a backtrack recovery mechanism is determined by a kinetic
competition between 1D diffusion and RNA cleavage. Notably, RNA polymerase I
(Pol I) and Pol II recover from shallow backtracks by 1D diffusion, use RNA
cleavage to recover from intermediary depths, and are unable to recover from
extensive backtracks. Furthermore, Pol I and Pol II use distinct mechanisms to
avoid nonrecoverable backtracking. Pol I is protected by its subunit A12.2,
which decreases the rate of 1D diffusion and enables transcript cleavage up to
20 nt. In contrast, Pol II is fully protected through association with the
cleavage stimulatory factor TFIIS, which enables rapid recovery from any depth
by RNA cleavage. Taken together, we identify distinct backtrack recovery
strategies of Pol I and Pol II, shedding light on the evolution of cellular
functions of these key enzymes. |
Which is the transcriptome of RNA polymerase III? | RNA polymerase III (Pol III) transcribes a limited set of short genes in eukaryotes producing abundant small RNAs, mostly tRNA. The originally defined yeast Pol III transcriptome appears to be expanding owing to the application of new methods. Newly identified Pol III transcripts include small nucleolar RNAs, microRNAs, short interspersed nuclear element-encoded or tRNA-derived RNAs and novel classes of ncRNA that can display significant sequence complementarity to protein-coding genes and might thus regulate their expression. | RNA polymerase III (Pol III) transcribes small untranslated RNAs, such as tRNAs.
To define the Pol III transcriptome in Saccharomyces cerevisiae, we performed
genome-wide chromatin immunoprecipitation using subunits of Pol III, TFIIIB and
TFIIIC. Virtually all of the predicted targets of Pol III, as well as several
novel candidates, were occupied by Pol III machinery. Interestingly, TATA
box-binding protein occupancy was greater at Pol III targets than virtually all
Pol II targets, and the highly occupied Pol II targets are generally strongly
transcribed. The temporal relationships between factor occupancy and gene
activity were then investigated at selected targets. Nutrient deprivation
rapidly reduced both Pol III transcription and Pol III occupancy of both a tRNA
gene and RPR1. In contrast, TFIIIB remained bound, suggesting that TFIIIB
release is not a critical aspect of the onset of repression. Remarkably, TFIIIC
occupancy increased dramatically during repression. Nutrient addition generally
reestablished transcription and initial occupancy levels. Our results are
consistent with active Pol III displacing TFIIIC, and with inactivation/release
of Pol III enabling TFIIIC to bind, marking targets for later activation. These
studies reveal new aspects of the kinetics, dynamics, and targets of the Pol III
system. The role of RNA polymerase (Pol) III in eukaryotic transcription is commonly
thought of as being restricted to a small set of highly expressed, housekeeping
non-protein-coding (nc)RNA genes. Recent studies, however, have remarkably
expanded the set of known Pol III-synthesized ncRNAs, suggesting that
gene-specific Pol III regulation is more common than previously appreciated.
Newly identified Pol III transcripts include small nucleolar RNAs, microRNAs,
short interspersed nuclear element-encoded or tRNA-derived RNAs and novel
classes of ncRNA that can display significant sequence complementarity to
protein-coding genes and might thus regulate their expression. The extent of the
Pol III transcriptome, the complexity of its regulation and its influence on
cell physiology, development and disease are emerging as new areas for future
research. RNA polymerase III (Pol III) transcribes a limited set of short genes in
eukaryotes producing abundant small RNAs, mostly tRNA. The originally defined
yeast Pol III transcriptome appears to be expanding owing to the application of
new methods. Also, several factors required for assembly and nuclear import of
Pol III complex have been identified recently. Models of Pol III based on
cryo-electron microscopy reconstructions of distinct Pol III conformations
reveal unique features distinguishing Pol III from other polymerases. Novel
concepts concerning Pol III functioning involve recruitment of general Pol
III-specific transcription factors and distinctive mechanisms of transcription
initiation, elongation and termination. Despite the short length of Pol III
transcription units, mapping of transcriptionally active Pol III with nucleotide
resolution has revealed strikingly uneven polymerase distribution along all
genes. This may be related, at least in part, to the transcription factors bound
at the internal promoter regions. Pol III uses also a specific negative
regulator, Maf1, which binds to polymerase under stress conditions; however, a
subset of Pol III genes is not controlled by Maf1. Among other RNA polymerases,
Pol III machinery represents unique features related to a short transcript
length and high transcription efficiency. |
How many PML isoforms exist in the human genome? | PML, the organizer of nuclear bodies (NBs), is expressed in several isoforms designated PMLI to VII which differ in their C-terminal region due to alternative splicing of a single gene. | The small nuclear structures known as ND10 or PML nuclear bodies have been
implicated in a variety of cellular processes including response to stress and
interferons, oncogenesis, and viral infection, but little is known about their
biochemical properties. Recently, a ubiquitin-specific protease enzyme (named
HAUSP) and a ubiquitin-homology family protein (PIC1) have been found associated
with ND10. HAUSP binds strongly to Vmw110, a herpesvirus regulatory protein
which has the ability to disrupt ND10, while PIC1 was identified as a protein
which interacts with PML, the prototype ND10 protein. We have investigated the
role of ubiquitin-related pathways in the mechanism of ND10 disruption by Vmw110
and the effect of virus infection on PML stability. The results show that the
disruption of ND10 during virus infection correlates with the loss of several
PML isoforms and this process is dependent on active proteasomes. The PML
isoforms that are most sensitive to virus infection correspond closely to those
which have recently been identified as being covalently conjugated to PIC1. In
addition, a large number of PIC1-protein conjugates can be detected following
transfection of a PIC1 expression plasmid, and many of these are also eliminated
in a Vmw110-dependent manner during virus infection. These observations provide
a biochemical mechanism to explain the observed effects of Vmw110 on ND10 and
suggest a simple yet powerful mechanism by which Vmw110 might function during
virus infection. Mucosal human papillomaviruses (HPVs) are the causative agents of a number of
human pathologies, including benign condylomas, as well as of the majority of
cervical cancers and their high-grade precursor lesions. Although the viral E6
protein is known to be essential for driving maligt progression of
HPV-infected cells, there are still many uncertainties about its mode of action.
In this study, we have analysed the intracellular distribution of the E6
oncoproteins from the high-risk HPV-18 and the low-risk HPV-11. We show that
both E6 proteins localize within the nucleus in nuclear bodies that are confocal
with the promyelocytic leukaemia (PML) protein. Using a panel of different PML
isoforms, we demonstrate specific co-localization between the E6 proteins and
PML isoforms I-IV, but not with PML isoforms V and VI. We also demonstrate the
interaction between E6 and a subset of PML isoforms in vivo. As a consequence of
this interaction, the insoluble form of PML IV is destabilized by HPV-18 E6
through a proteasome-dependent pathway. Interestingly, both HPV-11 E6 and HPV-18
E6 can readily overcome PML IV-induced cellular senescence in primary cells.
These results show separable functions for different PML isoforms that are
specifically targeted by the HPV E6 oncoproteins. Intrinsic antiviral resistance mediated by constitutively expressed cellular
proteins is one arm of defence against virus infection. Promyelocytic leukaemia
nuclear bodies (PML-NBs, also known as ND10) contribute to host restriction of
herpes simplex virus type 1 (HSV-1) replication via mechanisms that are
counteracted by viral regulatory protein ICP0. ND10 assembly is dependent on
PML, which comprises several different isoforms, and depletion of all PML
isoforms decreases cellular resistance to ICP0-null mutant HSV-1. We report that
individual expression of PML isoforms I and II partially reverses the increase
in ICP0-null mutant HSV-1 plaque formation that occurs in PML-depleted cells.
This activity of PML isoform I is dependent on SUMO modification, its SUMO
interaction motif (SIM), and each element of its TRIM domain. Detailed analysis
revealed that the punctate foci formed by individual PML isoforms differ subtly
from normal ND10 in terms of composition and/or Sp100 modification.
Surprisingly, deletion of the SIM motif from PML isoform I resulted in increased
colocalisation with other major ND10 components in cells lacking endogenous PML.
Our observations suggest that complete functionality of PML is dependent on
isoform-specific C-terminal sequences acting in concert. PML, the organizer of nuclear bodies (NBs), is expressed in several isoforms
designated PMLI to VII which differ in their C-terminal region due to
alternative splicing of a single gene. This variability is important for the
function of the different PML isoforms. PML NB formation requires the covalent
linkage of SUMO to PML. Arsenic trioxide (As₂O₃) enhances PML SUMOylation
leading to an increase in PML NB size and promotes its interaction with RNF4, a
poly-SUMO-dependent ubiquitin E3 ligase responsible for proteasome-mediated PML
degradation. Furthermore, the presence of a bona fide SUMO Interacting Motif
(SIM) within the C-terminal region of PML seems to be required for recruitment
of other SUMOylated proteins within PML NBs. This motif is present in all PML
isoforms, except in the nuclear PMLVI and in the cytoplasmic PMLVII. Using a
bioluminescence resoce energy transfer (BRET) assay in living cells, we found
that As₂O₃ enhanced the SUMOylation and interaction with RNF4 of nuclear PML
isoforms (I to VI). In addition, among the nuclear PML isoforms, only the one
lacking the SIM sequence, PMLVI, was resistant to As₂O₃-induced PML degradation.
Similarly, mutation of the SIM in PMLIII abrogated its sensitivity to
As₂O₃-induced degradation. PMLVI and PMLIII-SIM mutant still interacted with
RNF4. However, their resistance to the degradation process was due to their
inability to be polyubiquitinated and to recruit efficiently the 20S core and
the β regulatory subunit of the 11S complex of the proteasome in PML NBs. Such
resistance of PMLVI to As₂O₃-induced degradation was alleviated by
overexpression of RNF4. Our results demonstrate that the SIM of PML is
dispensable for PML SUMOylation and interaction with RNF4 but is required for
efficient PML ubiquitination, recruitment of proteasome components within NBs
and proteasome-dependent degradation of PML in response to As₂O₃. The tumor suppressor promyelocytic leukemia (PML) protein is fused to the
retinoic acid receptor alpha in patients suffering from acute promyelocytic
leukemia (APL). Treatment of APL patients with arsenic trioxide (As2O3) reverses
the disease phenotype by a process involving the degradation of the fusion
protein via its PML moiety. Several PML isoforms are generated from a single PML
gene by alternative splicing. They share the same N-terminal region containing
the RBCC/tripartite motif but differ in their C-terminal sequences. Recent
studies of all the PML isoforms reveal the specific functions of each. Here, we
review the nomenclature and structural organization of the PML isoforms in order
to clarify the various designations and classifications found in different
databases. The functions of the PML isoforms and their differential roles in
antiviral defense also are reviewed. Finally, the key players involved in the
degradation of the PML isoforms in response to As2O3 or other inducers are
discussed. Arsenic is a clinically effective treatment for acute promyelocytic leukaemia
(APL) in which the promyelocytic leukaemia (PML) protein is fused to retinoic
receptor alpha (RARα). PML-RARα is degraded by the proteasome by a
SUMO-dependent, ubiquitin-mediated pathway in response to arsenic treatment,
curing the disease. Six major PML isoforms are expressed as a result of
alternative splicing, each of which encodes a unique C-terminal region. Using a
system in which only a single EYFP-linked PML isoform is expressed, we
demonstrate that PMLI, PMLII and PMLVI accumulate in the cytoplasm following
arsenic treatment, whereas PMLIII, PMLIV and PMLV do not. 3D structured
illumination was used to obtain super-resolution images of PML bodies, revealing
spherical shells of PML along with associated SUMO. Arsenic treatment results in
dramatic isoform-specific changes to PML body ultrastructure. After extended
arsenic treatment most PML isoforms are degraded, leaving SUMO at the core of
the nuclear bodies. A high-content imaging assay identifies PMLV as the isoform
most readily degraded following arsenic treatment, and PMLIV as relatively
resistant to degradation. Immunoprecipitation analysis demonstrates that all PML
isoforms are modified by SUMO and ubiquitin after arsenic treatment, and by
using siRNA, we demonstrate that arsenic-induced degradation of all PML isoforms
is dependent on the ubiquitin E3 ligase RNF4. Intriguingly, depletion of RNF4
results in marked accumulation of PMLV, suggesting that this isoform is an
optimal substrate for RNF4. Thus the variable C-terminal domain influences the
rate and location of degradation of PML isoforms following arsenic treatment. Although modulation of the cellular tumor-suppressor p53 is considered to have
the major role in E1A/E1B-55K-mediated tumorigenesis, other promyelocytic
leukemia nuclear body (PML-NB)/PML oncogenic domain (POD)-associated factors
including SUMO, Mre11, Daxx, as well as the integrity of these nuclear bodies
contribute to the transformation process. However, the biochemical consequences
and oncogenic alterations of PML-associated E1B-55K by SUMO-dependent PML-IV and
PML-V interaction have so far remained elusive. We performed mutational analysis
to define a PML interaction motif within the E1B-55K polypeptide. Our results
showed that E1B-55K/PML binding is not required for p53, Mre11 and Daxx
interaction. We also observed that E1B-55K lacking subnuclear PML localization
because of either PML-IV or PML-V-binding deficiency was no longer capable of
mediating E1B-55K-dependent SUMOylation of p53, inhibition of p53-mediated
transactivation or efficiently transforming primary rodent cells. These results
together with the observation that E1B-55K-dependent SUMOylation of p53 is
required for efficient cell transformation, provides evidence for the idea that
the SUMO ligase activity of the E1B-55K viral oncoprotein is intimately linked
to its growth-promoting oncogenic activities. |
Do cephalopods use RNA editing less frequently than other species? | Extensive messenger RNA editing generates transcript and protein diversity in genes involved in neural excitability, as previously described, as well as in genes participating in a broad range of other cellular functions. | Coleoid cephalopods like squids have a camera-type eye similar to vertebrates.
On the other hand, Nautilus (Nautiloids) has a pinhole eye that lacks lens and
cornea. Since pygmy squid and Nautilus are closely related species they are
excellent model organisms to study eye evolution. Having being able to collect
Nautilus embryos, we employed next-generation RNA sequencing using Nautilus and
pygmy squid developing eyes. Their transcriptomes were compared and analyzed.
Enrichment analysis of Gene Ontology revealed that contigs related to nucleic
acid binding were largely up-regulated in squid, while the ones related to
metabolic processes and extracellular matrix-related genes were up-regulated in
Nautilus. These differences are most likely correlated with the complexity of
tissue organization in these species. Moreover, when the analysis focused on the
eye-related contigs several interesting patterns emerged. First, contigs from
both species related to eye tissue differentiation and morphogenesis as well as
to cilia showed best hits with their Human counterparts, while contigs related
to rabdomeric photoreceptors showed the best hit with their Drosophila
counterparts. This bolsters the idea that eye morphogenesis genes have been
generally conserved in evolution, and compliments other studies showing that
genes involved in photoreceptor differentiation clearly follow the
diversification of invertebrate (rabdomeric) and vertebrate (ciliated)
photoreceptors. Interestingly some contigs showed as good a hit with Drosophila
and Human homologues in Nautilus and squid samples. One of them, capt/CAP1, is
known to be preferentially expressed in Drosophila developing eye and in
vertebrate lens. Importantly our analysis also provided evidence of gene
duplication and diversification of their function in both species. One of these
genes is the Neurofibromatosis 1 (NF1/Nf1), which in mice has been implicated in
lens formation, suggesting a hitherto unsuspected role in the evolution of the
lens in molluscs. In the mutualistic relationship between the squid Euprymna tasmanica and the
bioluminescent bacterium Vibrio fischeri, several host factors, including
immune-related proteins, are known to interact and respond specifically and
exclusively to the presence of the symbiont. In squid and octopus, the white
body is considered to be an immune organ mainly due to the fact that blood
cells, or hemocytes, are known to be present in high numbers and in different
developmental stages. Hence, the white body has been described as the site of
hematopoiesis in cephalopods. However, to our knowledge, there are no studies
showing any molecular evidence of such functions. In this study, we performed a
transcriptomic analysis of white body tissue of the Southern dumpling squid, E.
tasmanica. Our primary goal was to gain insights into the functions of this
tissue and to test for the presence of gene transcripts associated with
hematopoietic and immune processes. Several hematopoiesis genes including CPSF1,
GATA 2, TFIID, and FGFR2 were found to be expressed in the white body. In
addition, transcripts associated with immune-related signal transduction
pathways, such as the toll-like receptor/NF-κβ, and MAPK pathways were also
found, as well as other immune genes previously identified in E. tasmanica's
sister species, E. scolopes. This study is the first to analyze an immune organ
within cephalopods, and to provide gene expression data supporting the white
body as a hematopoietic tissue. Coleoid cephalopods (octopus, squid and cuttlefish) are active, resourceful
predators with a rich behavioural repertoire. They have the largest nervous
systems among the invertebrates and present other striking morphological
innovations including camera-like eyes, prehensile arms, a highly derived early
embryogenesis and a remarkably sophisticated adaptive colouration system. To
investigate the molecular bases of cephalopod brain and body innovations, we
sequenced the genome and multiple transcriptomes of the California two-spot
octopus, Octopus bimaculoides. We found no evidence for hypothesized
whole-genome duplications in the octopus lineage. The core developmental and
neuronal gene repertoire of the octopus is broadly similar to that found across
invertebrate bilaterians, except for massive expansions in two gene families
previously thought to be uniquely enlarged in vertebrates: the protocadherins,
which regulate neuronal development, and the C2H2 superfamily of zinc-finger
transcription factors. Extensive messenger RNA editing generates transcript and
protein diversity in genes involved in neural excitability, as previously
described, as well as in genes participating in a broad range of other cellular
functions. We identified hundreds of cephalopod-specific genes, many of which
showed elevated expression levels in such specialized structures as the skin,
the suckers and the nervous system. Finally, we found evidence for large-scale
genomic rearrangements that are closely associated with transposable element
expansions. Our analysis suggests that substantial expansion of a handful of
gene families, along with extensive remodelling of genome linkage and repetitive
content, played a critical role in the evolution of cephalopod morphological
innovations, including their large and complex nervous systems. RNA editing, a post-transcriptional process, allows the diversification of
proteomes beyond the genomic blueprint; however it is infrequently used among
animals for this purpose. Recent reports suggesting increased levels of RNA
editing in squids thus raise the question of the nature and effects of these
events. We here show that RNA editing is particularly common in behaviorally
sophisticated coleoid cephalopods, with tens of thousands of evolutionarily
conserved sites. Editing is enriched in the nervous system, affecting molecules
pertinent for excitability and neuronal morphology. The genomic sequence
flanking editing sites is highly conserved, suggesting that the process confers
a selective advantage. Due to the large number of sites, the surrounding
conservation greatly reduces the number of mutations and genomic polymorphisms
in protein-coding regions. This trade-off between genome evolution and
transcriptome plasticity highlights the importance of RNA recoding as a strategy
for diversifying proteins, particularly those associated with neural function.
PAPERCLIP. RNA editing can yield protein products that differ from those directly encoded
by genomic DNA. This process is pervasive in the mitochondria of many
eukaryotes, where it predomitly results in the restoration of ancestral
protein sequences. Nuclear mRNAs in metazoans also undergo editing
(adenosine-to-inosine or 'A-to-I' substitutions), and most of these edits appear
to be nonadaptive 'misfirings' of adenosine deaminases. However, recent analysis
of cephalopod transcriptomes found that many editing sites are shared by
anciently divergent lineages within this group, suggesting they play some
adaptive role. Recent discoveries have also revealed that some fungi have an
independently evolved A-to-I editing mechanism, resulting in extensive recoding
of their nuclear mRNAs. Here, phylogenetic comparisons were used to determine
whether RNA editing generally restores ancestral protein sequences or creates
derived variants. Unlike in mitochondrial systems, RNA editing in metazoan and
fungal nuclear transcripts overwhelmingly leads to novel sequences not found in
inferred ancestral proteins. Even for the subset of RNA editing sites shared by
deeply divergent cephalopod lineages, the primary effect of nuclear editing is
an increase-not a decrease-in protein divergence. These findings suggest
fundamental differences in the forces responsible for the evolution of RNA
editing in nuclear versus mitochondrial systems. The adenosine-to-inosine (A-to-I) RNA editomes have been systematically
characterized in various metazoan species, and many editing sites were found in
clusters. However, it remains unclear whether the clustered editing sites tend
to be linked in the same RNA molecules or not. By adopting a method originally
designed to detect linkage disequilibrium of DNA mutations, we examined the
editomes of ten metazoan species and detected extensive linkage of editing in
Drosophila and cephalopods. The prevalent linkages of editing in these two
clades, many of which are conserved between closely related species and might be
associated with the adaptive proteomic recoding, are maintained by natural
selection at the cost of genome evolution. Nevertheless, in worms and humans, we
only detected modest proportions of linked editing events, the majority of which
were not conserved. Furthermore, the linkage of editing in coding regions of
worms and humans might be overall deleterious, which drives the evolution of DNA
sites to escape promiscuous editing. Altogether, our results suggest that the
linkage landscape of A-to-I editing has evolved during metazoan evolution. This
present study also suggests that linkage of editing should be considered in
elucidating the functional consequences of RNA editing. |
Is there an RNAi drug being developed to treat amyloidosis? | Yes, patisiran is an investigational RNA interference (RNAi) therapeutic in development for the treatment of hereditary ATTR (hATTR) amyloidosis. | BACKGROUND: Patisiran is an investigational RNA interference (RNAi) therapeutic
in development for the treatment of hereditary ATTR (hATTR) amyloidosis, a
progressive disease associated with significant disability, morbidity, and
mortality.
METHODS: Here we describe the rationale and design of the Phase 3 APOLLO study,
a randomized, double-blind, placebo-controlled, global study to evaluate the
efficacy and safety of patisiran in patients with hATTR amyloidosis with
polyneuropathy. Eligible patients are 18-85 years old with hATTR amyloidosis,
investigator-estimated survival of ≥2 years, Neuropathy Impairment Score (NIS)
of 5-130, and polyneuropathy disability score ≤IIIb. Patients are randomized 2:1
to receive either intravenous patisiran 0.3 mg/kg or placebo once every 3 weeks.
The primary objective is to determine the efficacy of patisiran at 18 months
based on the difference in the change in modified NIS+7 (a composite measure of
motor strength, sensation, reflexes, nerve conduction, and autonomic function)
between the patisiran and placebo groups. Secondary objectives are to evaluate
the effect of patisiran on Norfolk-Diabetic Neuropathy quality of life
questionnaire score, nutritional status (as evaluated by modified body mass
index), motor function (as measured by NIS-weakness and timed 10-m walk test),
and autonomic symptoms (as measured by the Composite Autonomic Symptom Score-31
questionnaire). Exploratory objectives include assessment of cardiac function
and pathologic evaluation to assess nerve fiber innervation and amyloid burden.
Safety of patisiran will be assessed throughout the study.
DISCUSSION: APOLLO represents the largest randomized, Phase 3 study to date in
patients with hATTR amyloidosis, with endpoints that capture the multisystemic
nature of this disease.
TRIAL REGISTRATION: This trial is registered at clinicaltrials.gov ( NCT01960348
); October 9, 2013. |
Which receptor does GDF15 bind? | GDF15 binds specifically to GDNF family receptor α-like (GFRAL) | Growth differentiation factor 15 (GDF15; also known as MIC-1) is a divergent
member of the TGF-β superfamily and is associated with body-weight regulation in
humans and rodents. However, the cognate receptor of GDF15 is unknown. Here we
show that GDF15 binds specifically to GDNF family receptor α-like (GFRAL) with
high affinity, and that GFRAL requires association with the coreceptor RET to
elicit intracellular signaling in response to GDF15 stimulation. We also found
that GDF15-mediated reductions in food intake and body weight of mice with
obesity were abolished in GFRAL-knockout mice. We further found that GFRAL
expression was limited to hindbrain neurons and not present in peripheral
tissues, which suggests that GDF15-GFRAL-mediated regulation of food intake is
by a central mechanism. Lastly, given that GDF15 did not increase energy
expenditure in treated mice with obesity, the anti-obesity actions of the
cytokine are likely driven primarily by a reduction in food intake. |
Is there an association between carcinoid syndrome and mitral valve disease? | Yes, mitral valve damage was reported in patients with carcinoid syndrome. | An observation of carcinoid syndrome in a woman of 47 suffering from maligt
carcinoid of the ileum with metastases into the liver and right ovary is
described. The clinical picture included diarrhea, heat waves, bronchospasms,
hypertension, hyperserotoninemia, affection of the mitral valve and left atrium.
"Carcinoid plaques" in the endocardium formed due to excessive proliferation
under the influence of serotonin and kinins of polypotent subendothelial cells
followed by their differentiation into fibroblast-like and smooth-muscle
elements and production of basophilic interstitial substance. The receding
rheumatic affection of the mitral valve may be the cause of the predomit
involvement of the left part of the heart. We report two observations of significant left heart involvement in patients
with the carcinoid syndrome assessed by transthoracic and transoesophageal
echocardiography. Echocardiographic lesions of this kind have only been reported
twice. In the present cases, there was mitral involvement with mitral
regurgitation in one case and a mitro-aortic involvement with mitral and aortic
regurgitation in the other. The mechanism of left heart lesions is unclear since
in both cases no right-to-left cardiac shunt was present, as attested by colour
Doppler and saline contrast transoesophageal echocardiography. The location of
the primary tumour was unknown in one case and ileal in the other; no pulmonary
metastasis was detected. The use of transoesophageal echocardiography might make
it possible to detect left-sided cardiac lesions more frequently since they were
found in anatomical series, in 30% of patients with carcinoid syndrome. BACKGROUND: Carcinoid involvement of left-sided heart valves has been reported
in patients with a patent foramen ovale, carcinoid tumor of the lung, and active
carcinoid syndrome with high levels of serotonin. The present study details the
clinical features and surgical management of patients with carcinoid heart
disease affecting both left- and right-sided valves.
METHODS AND RESULTS: Eleven patients (7 men, 4 women) with symptomatic carcinoid
heart disease underwent surgery for left- and right-sided valve disease between
1989 and 1999. Mean age was 57+/-9 years, and median preoperative NYHA class was
3. All patients had metastatic carcinoid tumors and were on somatostatin analog.
Of 11 patients, 5 (45%) had a patent foramen ovale; 1 of these also had a
primary lung carcinoid tumor. Surgery included tricuspid valve replacement in
all patients, pulmonary valve replacement in 3 and valvectomy in 7, mitral valve
replacement in 6 and repair in 1, aortic valve replacement in 4 and repair in 2,
CABG in 2, and patent foramen ovale closure in 5. One myocardial metastatic
carcinoid tumor was removed. There were 2 perioperative deaths. At a mean
follow-up of 41 months, 4 additional patients were dead. All but 1 surgical
survivor initially improved >/=1 functional class. No patient required
reoperation.
CONCLUSIONS: Carcinoid heart disease may affect left- and right-sided valves and
occurred without intracardiac shunting in 55% of this surgical series. Despite
metastatic disease that limits longevity, operative survivors had improvement in
functional capacity. Cardiac surgery should be considered for select patients
with carcinoid heart disease affecting left- and right-sided valves. Heart valves exhibit a highly-conserved stratified structure exquisitely
designed to counter biomechanical forces delivered over a lifetime. Heart valve
structure and competence is maintained by heart valve cells through a process of
continuous turnover extracellular matrix (ECM). Degenerative (myxomatous) mitral
valve disease (DMVD) is an important disease associated with aging in both dogs
and humans. DMVD is increasingly regarded as a disease with identifiable
signaling mechanisms that control key genes associated with regulation and
dysregulation of ECM homeostasis. Initiating stimuli for these signaling
pathways have not been fully elucidated but likely include both mechanical and
chemical stimuli. Signaling pathways implicated in DMVD include serotonin,
transforming growth factor β (TGFβ), and heart valve developmental pathways.
High circulating serotonin (carcinoid syndrome) and serotoninergic drugs are
known to cause valvulopathy that shares pathologic features with DMVD. Recent
evidence supports a local serotonin signaling mechanism, possibly triggered by
high tensile loading on heart valves. Serotonin initiates TGFβ signaling, which
in turn has been strongly implicated in canine DMVD. Recent evidence suggests
that degenerative aortic and mitral valve disease may involve pathologic
processes that mimic osteogenesis and chondrogenesis, respectively. These
processes may be mediated by developmental pathways shared by heart valves,
bone, and cartilage. These pathways include bone morphogenic protein (BMP) and
Wnt signaling. Other signaling pathways implicated in heart valve disease
include Notch, nitric oxide, and angiotensin II. Ultimately, increased
understanding of signaling mechanisms could point to therapeutic strategies
aimed at slowing or halting disease progression. BACKGROUND: Symptoms and survival of patients with carcinoid syndrome have
improved, but development of carcinoid heart disease (CaHD) continues to
decrease survival.
OBJECTIVES: This study aimed to analyze patient outcomes after valve surgery for
CaHD during a 27-year period at 1 institution to determine early and late
outcomes and opportunities for improved patient care.
METHODS: We retrospectively studied the short-term and long-term outcomes of all
consecutive patients with CaHD who underwent valve replacement at our
institution between 1985 and 2012.
RESULTS: The records of 195 patients with CaHD were analyzed. Pre-operative New
York Heart Association class was III or IV in 125 of 178 patients (70%). All had
tricuspid valve replacement (159 bioprostheses, 36 mechanical), and
157 underwent a pulmonary valve operation. Other concomitant operations included
mitral valve procedure (11%), aortic valve procedure (9%), patent foramen ovale
or atrial septal defect closure (23%), cardiac metastasectomies or biopsy (4%),
and simultaneous coronary artery bypass (11%). There were 20 perioperative
deaths (10%); after 2000, perioperative mortality was 6%. Survival rates (95%
confidence intervals) at 1, 5, and 10 years were 69% (63% to 76%), 35% (28% to
43%), and 24% (18% to 32%), respectively. Overall mortality was associated with
older age, cytotoxic chemotherapy, and tobacco use; 75% of survivors had
symptomatic improvement at follow-up. Presymptomatic valve operation was not
associated with late survival benefit.
CONCLUSIONS: Operative mortality associated with valve replacement surgery for
CaHD has decreased. Symptomatic and survival benefit is noted in most patients
when CaHD is managed by an experienced multidisciplinary team. |
How many of the human PML isoforms are cytosolic? | Using a system in which only a single EYFP-linked PML isoform is expressed, we demonstrate that PMLI, PMLII and PMLVI accumulate in the cytoplasm following arsenic treatment, whereas PMLIII, PMLIV and PMLV do not the PML isoforms that are most sensitive to virus infection correspond closely to those which have recently been identified as being covalently conjugated to PIC1. | The small nuclear structures known as ND10 or PML nuclear bodies have been
implicated in a variety of cellular processes including response to stress and
interferons, oncogenesis, and viral infection, but little is known about their
biochemical properties. Recently, a ubiquitin-specific protease enzyme (named
HAUSP) and a ubiquitin-homology family protein (PIC1) have been found associated
with ND10. HAUSP binds strongly to Vmw110, a herpesvirus regulatory protein
which has the ability to disrupt ND10, while PIC1 was identified as a protein
which interacts with PML, the prototype ND10 protein. We have investigated the
role of ubiquitin-related pathways in the mechanism of ND10 disruption by Vmw110
and the effect of virus infection on PML stability. The results show that the
disruption of ND10 during virus infection correlates with the loss of several
PML isoforms and this process is dependent on active proteasomes. The PML
isoforms that are most sensitive to virus infection correspond closely to those
which have recently been identified as being covalently conjugated to PIC1. In
addition, a large number of PIC1-protein conjugates can be detected following
transfection of a PIC1 expression plasmid, and many of these are also eliminated
in a Vmw110-dependent manner during virus infection. These observations provide
a biochemical mechanism to explain the observed effects of Vmw110 on ND10 and
suggest a simple yet powerful mechanism by which Vmw110 might function during
virus infection. Arsenic is a clinically effective treatment for acute promyelocytic leukaemia
(APL) in which the promyelocytic leukaemia (PML) protein is fused to retinoic
receptor alpha (RARα). PML-RARα is degraded by the proteasome by a
SUMO-dependent, ubiquitin-mediated pathway in response to arsenic treatment,
curing the disease. Six major PML isoforms are expressed as a result of
alternative splicing, each of which encodes a unique C-terminal region. Using a
system in which only a single EYFP-linked PML isoform is expressed, we
demonstrate that PMLI, PMLII and PMLVI accumulate in the cytoplasm following
arsenic treatment, whereas PMLIII, PMLIV and PMLV do not. 3D structured
illumination was used to obtain super-resolution images of PML bodies, revealing
spherical shells of PML along with associated SUMO. Arsenic treatment results in
dramatic isoform-specific changes to PML body ultrastructure. After extended
arsenic treatment most PML isoforms are degraded, leaving SUMO at the core of
the nuclear bodies. A high-content imaging assay identifies PMLV as the isoform
most readily degraded following arsenic treatment, and PMLIV as relatively
resistant to degradation. Immunoprecipitation analysis demonstrates that all PML
isoforms are modified by SUMO and ubiquitin after arsenic treatment, and by
using siRNA, we demonstrate that arsenic-induced degradation of all PML isoforms
is dependent on the ubiquitin E3 ligase RNF4. Intriguingly, depletion of RNF4
results in marked accumulation of PMLV, suggesting that this isoform is an
optimal substrate for RNF4. Thus the variable C-terminal domain influences the
rate and location of degradation of PML isoforms following arsenic treatment. |
Which tendons are affected in the Dequervain's tenosynovitis? | DeQuervain's tenosynovitis is a common cause of radial-sided wrist pain. Symptoms result from a narrow first dorsal compartment and associated tendinosis of the enclosed extensor pollicis brevis and/or abductor pollicis longus. | DeQuervain tenosynovitis, which involves the abductor pollicis longus and
extensor pollicis brevis tendons, is much more common in women than men and is
due to repetitive movements of the hand such as grasping and twisting.
Housewives and persons involved in manual occupations using the hands and wrists
account for most cases in previous series. In this series, six of 24 female
patients (25%) were pregt or postpartum at the time of onset. In five of the
six, activities of infant care aggravated symptoms. Both pregcy, per se, and
mechanical factors appear to play a role in causing this condition. DeQuervain's disease of the first dorsal compartment of the wrist, is a common
wrist pathology, pain results from resisted gliding of the abductor pollicis
longus and the extensor pollicis brevis tendon in the fibroosseous canal.
Management of resistant cases of DeQuervain's disease with failed conservative
treatment treated by surgical decompression yield satisfactory outcomes. A large
number of patients being dissatisfied with the medical treatment, still present
with persistent pain and positive clinical finding. Surgical decompression is an
effective method for the treatment of resistant cases of DeQuervain's disease.
Outcome variables were measured by Scheller, Forget and Macey evaluation
criteria. Most of our patients were female 28(93.3%), housewife 17(56.7%) with
mean age of 41.57 years, ranging from 25-60 years. Right sided involvement was
20(66.7%) and Left sided involvement was 10(33.3%). Restricted movement of thumb
in 30(100%) were the predomit symptoms. One (3.3%) patient develop chronic
tenosynovitis, 1(3.3%) patient develop hypertrophic scar. There was no wound
infection in the follow-up period of 3-18 months. Satisfactory results were
found in 29(96.7%). DeQuervain's tenosynovitis is a common cause of radial-sided wrist pain.
Symptoms result from a narrow first dorsal compartment and associated tendinosis
of the enclosed extensor pollicis brevis and/or abductor pollicis longus (APL).
Surgical intervention, offered when conservative measures fail to adequately
relieve symptoms, requires a detailed understanding of potentially aberrant
anatomy in order to avoid persistence or recurrence of symptoms. We describe a
case whereby the patient presented with complaints of thumb triggering in
extension and associated disabling first dorsal compartment tendinosis.
Intraoperatively, after supernumerary tendons were identified and addressed, the
APL was at risk for subluxation over a prominent fibroosseous ridge. Routine
first dorsal compartment release alone may have failed to address all of this
patient's pathology. |
Are there RNAi approaches considered for the treatment of kidney injury? | Yes, RNAi approaches are being considered for the treatment of kidney injury. | RNA interference has tremendous yet unrealized potential to treat a wide range
of illnesses. Innovative solutions are needed to protect and selectively deliver
small interfering RNA (siRNA) cargo to and within a target cell to fully exploit
siRNA as a therapeutic tool in vivo. Herein, we describe ammonium-functionalized
carbon otube (fCNT)-mediated transport of siRNA selectively and with high
efficiency to renal proximal tubule cells in animal models of acute kidney
injury (AKI). fCNT enhanced siRNA delivery to tubule cells compared to siRNA
alone and effectively knocked down the expression of several target genes,
includingTrp53,Mep1b,Ctr1, andEGFP A clinically relevant cisplatin-induced
murine model of AKI was used to evaluate the therapeutic potential of
fCNT-targeted siRNA to effectively halt the pathogenesis of renal injury.
Prophylactic treatment with a combination of fCNT/siMep1band
fCNT/siTrp53significantly improved progression-free survival compared to
controls via a mechanism that required concurrent reduction of meprin-1β and p53
expression. The fCNT/siRNA was well tolerated, and no toxicological consequences
were observed in murine models. Toward clinical application of this platform,
fCNTs were evaluated for the first time in nonhuman primates. The rapid and
kidney-specific pharmacokinetic profile of fCNT in primates was comparable to
what was observed in mice and suggests that this approach is amenable for use in
humans. The ocarbon-mediated delivery of siRNA provides a therapeutic means
for the prevention of AKI to safely overcome the persistent barrier of
nephrotoxicity during medical intervention. |
What does davunetide do to microtubules? | Davunetide or NAP is a microtubule-stabilizer. | |
Is propranolol used for treatment of infantile hemangioma? | Yes, propranolol is becoming the treatment of choice for complicated infantile hemangioma. | The objective of this study is to describe the initial use of propranolol as the
sole treatment for focal infantile airway hemangiomas, and to report on
available literature describing the use of propranolol for airway lesions. This
retrospective case series was carried out at a tertiary pediatric medical
center. We obtained the following results: two children demonstrated significant
response to oral propranolol therapy and avoided not only invasive surgical
procedures, but also long-term administration of oral corticosteroids. This is
the first report of treating infantile airway hemangiomas with only propranolol
without additional surgical intervention or corticosteroid use. Review of
literature reveals initial case series with similar, successful results using
propranolol as an adjuvant treatment along with other medications and surgical
interventions. We conclude that the initial use of propranolol as the sole
treatment for infantile airway hemangioma is promising. Literature review
reveals that propranolol as the sole treatment for most head and neck
hemangiomas shows significant promise based on early case reports. Further
studies are needed to determine the long-term effectiveness, dosing strategies,
and side effect profile of propranolol treatment for hemangiomas. Propranolol treatment was recently reported to be successful for the management
of severe infantile hemangioma. Known adverse effects of propranolol treatment
include transient bradycardia, hypotension, hypoglycemia, and bronchospasm (in
patients with underlying spastic respiratory illnesses), which led to a general
recommendation to gradually increase propranolol dosage and closely monitor
patients' hemodynamics at the onset of therapy. To date, no serious or
unexpected adverse effects that required specific intervention have been
reported. In this report, we describe the case of a 17-week-old female preterm
infant who presented with a large, ulcerated, cutaneous-subcutaneous hemangioma
of the right lateral thoracic wall, which we treated successfully with
propranolol. A few days into therapy, a potentially life-threatening adverse
effect, severe hyperkalemia, was observed and required treatment with loop
diuretics, fluids, and nebulized salbutamol to normalize her serum potassium
levels. This therapy could be gradually tapered and finally discontinued only
after several weeks of propranolol treatment. Our case report indicates that, at
least during the initial phase of the propranolol treatment of infantile
hemangioma, close monitoring of serum electrolytes, besides the monitoring of
hemodynamics and blood glucose, is necessary. Propranolol has been used successfully in a limited number of children with
infantile hemangiomas. This multicenter retrospective study describes the
efficacy and adverse effects of propranolol in infantile hemangioma. Seventy-one
infants with infantile hemangiomas were treated with oral propranolol, 1
mg/kg/12 hours, for at least 12 weeks. A photograph based severity scoring
assessment was performed by five observers to evaluate efficacy, utilizing a
scoring system of 10 as the original infantile hemangioma before treatment and 0
as completely normal skin. The mean of the five independent measurements was
used in the analysis. Propranolol was a rapid and effective treatment for
infantile hemangiomas at 4 weeks (p < 0.001), at 8 weeks (p < 0.001 compared to
the 4 wks value), at 12 weeks (p < 0.05 compared to the 8 wks value), and
thereafter up to 32 weeks (p < 0.01 compared to the 16 wks value). The response
of infantile hemangiomas to propranolol was similar regardless of sex, age at
onset of treatment, type of involvement (segmental and nonsegmental), facial
segments affected, special locations (eyelid, nasal tip, and parotid region),
ulceration, and depth of infantile hemangiomas. Very few side effects were
reported; mainly agitated sleep in 10 of 71 patients. In the series of patients
in this study, oral propranolol 2 mg/kg/day was a well-tolerated and effective
treatment for infantile hemangiomas. Prospective studies are needed to establish
the exact role of propranolol in the treatment of infantile hemangiomas. OBJECTIVE: To investigate the clinical results of the treatment of severe
infantile hemangioma with high-dose propranolol in Chinese.
METHODS: 56 cases with severe infantile hemangioma were treated with
propranolol. Clinical evaluation, electrocardiography, and experimental
examination of liver function and heart function were performed before
treatment. The daily dose of propranolol was increased from 1 mg/kg at the first
day to 1.5 mg/kg at the second day, and to 2 mg/kg at the third day. The
propranolol was given twice a day. The treatment was lasted for six months. The
patients were visited every month.
RESULTS: The lesion color was changed after 2-4 days of treatment in all the
cases. All the lesions were dramatically improved after one month of treatment.
The ulceration were healed, except one case. Until now, complete regression was
achieved in 10 cases and marked improvement in 46 cases. Side effects were
happened in 3 cases, including one case of abnormal liver function, one case of
CK-MB increase and one case of continuous increase of CK-MB, LDH, ALT, GGT.
CONCLUSIONS: High-dose Propranolol is very effective in the treatment of
infantile hemangioma with minor side effects and short disease period. It might
he used as the first-line treatment for infantile hemangioma. Infantile hemangiomas (IH) are common benign tumors in infancy, affecting 5-10%
of all infants and they can still cause disfigurement and serious complications
depending on their location and size, which can be associated with ulcerations
and haemorrhage. Since 2008, propranolol has become the first choice of therapy
for complicated IH, compared to conventional approach with systemic
corticosteroid therapy as first-line treatment and then interferon or
vincristine as second- or third-line therapeutic agents. We report three cases
of hemangioma, successfully treated with propranolol. Oral propranolol was given
for a period of 6 months with monthly follow up. All cases showed dramatic
response without any relapse after stopping the treatment. Propranolol is novel
and safe medication for treatment of infantile hemangioma. BACKGROUND: Hemangioma in infants has a benign self-limited course, but the 10%
of cases with complications need further treatment. Successful treatment with
propranolol in western countries has been reported over the past few years. We
evaluated the efficacy of propranolol for treating infantile hemangioma in
Taiwanese newborns and young infants.
METHODS: Patients below 1 year of age treated with propanolol between November
2009 and March 2011 were enrolled. Demographic data, clinical features, imaging
findings, treatment regimens of propranolol, and outcome were investigated.
RESULTS: Thirteen patients were treated with propranolol at a dose of 2-3
mg/kg/day. Seven (53.8%) patients had solitary hemangioma and six had multiple
ones. The indications for treatment were risk of local event in nine patients,
functional risk in four, local complication in one, and life-threatening
complication in one. The median age for starting propranolol was 4 months
(range: 1-11 months). Responses to propranolol, such as decolorization,
regression in tumor size, or improvement of hemangioma-associated complications
were observed in all patients within 1-2 weeks after treatment.
Propranolol-associated adverse effects occurred in two patients. One infant had
occasional tachypnea, and the other had occasional pale-looking appearance. The
symptoms resolved after dosage tapering.
CONCLUSION: Propranolol may be a promising therapeutic modality for infantile
hemangioma. Therapeutic strategies are needed to evaluate the optimal treatment
protocol and long-term adverse effects. Propranolol hydrochloride is a nonselective β-blocker that is used for the
treatment of hypertension, arrhythmia, and angina pectoris. In Japan, it was
recently approved for the treatment of childhood arrhythmia. It has been
observed to produce drastic involution of infantile hemangiomas. The aim of this
prospective study was to examine propranolol's superiority to classical therapy
with pulsed dye laser and/or cryosurgery in treating proliferating infantile
hemangiomas. Fifteen patients between the ages of 1 and 4 months with
proliferating infantile hemangiomas received grinded propranolol tablets 2 mg/kg
per day divided in three doses. Twelve patients with proliferating infantile
hemangiomas receiving pulsed dye laser and/or cryosurgery were enrolled as
controls. Baseline electrocardiogram, echocardiogram, and chest x-ray were
performed. Monitoring of heart rate, blood pressure, and blood glucose was
performed every 2 weeks. Efficacy was assessed by performing blinded volume
measurements and taking photographs at every visit. Propranolol induced
significantly earlier involution and redness reduction of infantile hemangiomas,
compared to pulsed dye laser and cryosurgery. Adverse effects such as
hypoglycemia, hypotension, or bradycardia did not occur.
CONCLUSION: The dramatic response of infantile hemangiomas to propranolol and
few side effects suggest that early treatment of infantile hemangiomas could
result in decreased disfigurement. Propranolol should be considered as a
first-line treatment of infantile hemangiomas. IMPORTANCE: Propranolol therapy is changing the treatment paradigm for infantile
hemangioma. This study addresses the effect of propranolol therapy on the
treatment of nasal infantile hemangioma (NIH), an area that often does not
respond to medical therapy.
OBJECTIVE: To determine if propranolol treatment is associated with fewer
invasive treatments for NIH.
DESIGN, SETTING, AND PARTICIPANTS: Retrospective cohort study conducted within a
single pediatric institution's multidisciplinary vascular anomaly program for
patients with NIH treated between January 1, 2003, and December 31, 2011. Three
NIH cohorts were compared: prepropranolol (20 in group 1; 2003-2009),
propranolol (25 in group 2; 2009-2011), and nonpropranolol (13 in group 3;
2009-2011) treatment.
INTERVENTIONS: Analysis of systemic medical, laser, or surgical therapies for
NIH.
MAIN OUTCOMES AND MEASURES: The study plan was created to detect a change in
invasive therapy for NIH. Data collected included presenting age, sex, affected
nasal subunits, infantile hemangioma morphologic characteristics, treatment type
and number, and primary treating service. An NIH grading system, based on nasal
subunit involvement, helped quantify treatment change. Descriptive statistics
summarized data, and a Cox proportional hazards regression model evaluated
propranolol use and the likelihood of invasive treatments (surgical excision or
laser).
RESULTS: Of the 95 patients identified, 58 met inclusion criteria: 20 in group 1
(mean age, 4.8 months), 25 in group 2 (mean age, 4.9 months), and 13 in group 3
(mean age, 4.9 months). Nasal infantile hemangiomas involved the nasal tip
subunit in 33 of 58 patients (56.9%). Eight of 13 patients (61.5%) in group 3
frequently had small NIH (grade 1). Patients in group 2 were less likely to
undergo any invasive treatments (relative risk, 0.44; 95% CI, 0.27-0.73), have
surgical excision only (0.45; 0.15-1.38), or undergo laser treatment only (0.44;
0.27-0.78) compared with those in group 1. Patients with higher-grade NIH had
more medical or invasive therapy, but invasive procedures were carried out in
each subgroup defined by grade.
CONCLUSIONS AND RELEVANCE: Patients with isolated propranolol-treated NIH were
less likely to undergo invasive treatment, but despite its implementation, the
need for invasive treatment was not totally supplanted by its use. PURPOSE: The successful use of nadolol as an alternative to propranolol therapy
in three cases of infantile hemangioma is reported.
SUMMARY: Infantile hemangioma is a benign vascular neoplastic disorder that
affects up to 10% of newborns and can lead to deformity or local complications
in severe cases. Propranolol, administered alone or in combination with
corticosteroids, is increasingly used to treat infantile hemangioma, but its
ability to cross the blood-brain barrier and potentially cause central nervous
system adverse effects has prompted research on alternative β-blocker therapies
for the disorder that have more favorable safety profiles, including nadolol.
This article describes the use of nadolol to treat three pediatric patients with
a buccal or genital hemangioma who developed adverse reactions (mainly,
irritability and sleep disturbances) or resistance to initial treatment with
propranolol. The patients were 10 months, 12 months, and 4 years of age,
respectively, when hemangioma treatment was initiated. The results of nadolol
therapy were favorable, with involution of lesions and gradual disappearance of
propranolol-associated adverse effects occurring in all three cases. As with any
use of β-blocker therapy in a pediatric patient, a cardiac workup is advised
before the start of nadolol therapy; blood pressure and heart rate monitoring
should be performed at one and two hours after the first dose and continued
during dose escalation.
CONCLUSION: Nadolol was an effective alternative to propranolol in three
pediatric patients with hemangiomas. PURPOSE: Genital infantile hemangiomas are vascular anomalies that often require
complex management and interdisciplinary care. Propranolol was first used to
treat patients with infantile hemangiomas in 2008 and has since gained
acceptance as first-line therapy.
MATERIALS AND METHODS: We review the presentation, course, management and
outcomes of all cases of genital infantile hemangiomas managed by propranolol
administration at a single institution from April 2010 to July 2014.
RESULTS: During the study period 9 patients with genital infantile hemangiomas
were referred to our hemangioma treatment clinic. Propranolol was initially
administered under careful outpatient monitoring at a dose of 1 mg/kg daily in 8
patients. One patient, a 700 gm premature infant, was started on therapy in the
inpatient setting at 0.5 mg/kg daily, given the history of prematurity. All
patients underwent successful increase of dose to at least 2 mg/kg for the
observation phase after tolerating the starting doses. One patient discontinued
propranolol prematurely per parental request due to concern regarding peripheral
vasoconstriction. Otherwise, no patient demonstrated significant hypotension,
symptomatic bradycardia, hypoglycemia or other major side effect requiring
treatment discontinuation. All patients who continued the treatment protocol had
excellent response to therapy.
CONCLUSIONS: Propranolol therapy for genital infantile hemangiomas was
successfully initiated and the dosage increased in 9 young children without
significant side effects and with marked improvement in all patients who
continued on treatment. Propranolol is the only Food and Drug Administration
approved therapy for treatment of patients with this vascular anomaly and should
be considered first-line therapy for genital infantile hemangiomas. BACKGROUND: Intracranial infantile hemangiomas are extremely rare, with only 36
patients reported in literature. Treatment for intracranial infantile
hemangiomas has been mostly limited to surgery, steroids, and interferon
therapy. Propranolol, which is often used to treat cutaneous infantile
hemangiomas, is not currently standard treatment for intracranial infantile
hemangiomas.
PATIENT DESCRIPTION: We present a one-month old boy with an intracranial
infantile hemangioma treated with propranolol.
RESULTS: This boy was being treated with oral propranolol for a supraclavicular
infantile hemangioma. Subsequent brain magnetic resoce imaging (MRI) scan
showed evidence of an associated intracranial infantile hemangioma in the right
cerebellopontine angle. Repeat brain MRI scan after two months of propranolol
treatment demonstrated a significant reduction in the size of the intracranial
infantile hemangioma.
CONCLUSIONS: This is the first report of successful therapy of an intracranial
infantile hemangioma with propranolol. BACKGROUND: More and more infantile hemangiomas (IH) are being treated with
propranolol, but the effectiveness, dosage, and treatment course are still in
dispute. The aim of this observational study was to describe the therapeutic
response, tolerance, and safety of low-dose propranolol in 23 children with IH
of the head and neck.
METHODS: Data were collected from the medical charts of patients treated with
low-dose propranolol from December 2009 through November 2011. Oral dose was
1-1.5 mg/kg once per day. Blood pressure and heart rate were monitored during
the first 24 h of treatment. In the absence of side-effects, treatment was
continued at home and the child was re-evaluated every month.
RESULTS: All patients had a good response, even if treated with corticosteroid
previously. Color and growth changes within 1 week were noted. Treatment
continued for a mean total duration of 6 months until the IH had totally
disappeared or stabilized. There were no severe adverse reactions. Side-effects
were limited and mild, including blood pressure decrease, somnolence, and
nausea. No relapse was noted.
CONCLUSIONS: Low-dose propranolol appears to be effective and safe for IH,
especially for those patients previously treated with corticosteroid and who had
no response or severe side-effects. IMPORTANCE: Propranolol hydrochloride has become the primary medical treatment
for problematic infantile hemangioma; however, the expression of propranolol's
target receptors during growth, involution, and treatment of hemangioma remains
unclear.
OBJECTIVE: To measure and compare the expression of β1-, β2-, and β3-adrenergic
receptors (ADBR1, ADBR2, and ADBR3, respectively) in proliferative (n = 10),
involuted (n = 11), and propranolol-responsive (n = 12) hemangioma tissue.
DESIGN, SETTING, AND PARTICIPANTS: Infantile hemangioma specimens were harvested
for molecular investigation. Messenger RNA (mRNA) expression of the ADBR1,
ADBR2, and ADBR3 genes was detected by real-time polymerase chain reaction.
Protein level expression was measured by Western blot and standardized with
densitometry. A total of 33 specimens were collected from patients in a tertiary
pediatric hospital who underwent excision of problematic hemangiomas. This study
was conducted from January 18, 2011, to September 24, 2013, and data analysis
was performed from February 25, 2015, to June 25, 2016.
RESULTS: Of the 33 patients included, 21 were female (64%). The mean (SD)
patient age at the time of excision was 7 (2.5) months for the proliferative
group lesions, 23.5 (10) months for the involuted group, and 16 (10) months for
the propranolol group. The mean level of ADBR1 mRNA expression was significantly
higher in proliferative hemangioma than in propranolol-responsive hemangioma
(1.05 [0.56] vs 0.52 [0.36]; P = .01; 95% CI, 0.12-0.94). There was no
difference in ADBR2 expression among the groups. Protein expression of ADBR3 was
significantly higher in involuted (0.64 [0.12] vs 0.26 [0.04]; P < .01; 95% CI,
0.26-0.49) and propranolol-responsive hemangioma (0.66 [0.31] vs 0.26 [0.04];
P = .01; 95% CI, 0.16-0.68) compared with proliferative hemangioma.
CONCLUSIONS AND RELEVANCE: These data demonstrate the variable expression of
ADBR subtypes among infantile hemangiomas during growth, involution, and
response to treatment. These findings may have clinical implications regarding
the use of selective vs nonselective β-blockade.
LEVEL OF EVIDENCE: 2. BACKGROUND: Infantile hemangiomas (IHs) are the most common benign vascular
tumors of childhood. Propranolol is an effective drug in treating IH. A reliable
and complementary instrument is necessary to evaluate IH response to propranolol
in addition to clinical and photographic assessments. Ultrasonography is a
simple and non-invasive technique that enables precise measurements of tumor
size and contributes to objective follow-up.
OBJECTIVE: To demonstrate the use of serial ultrasonography as an adjunctive
tool for assessment of IH treatment with propranolol.
PATIENTS AND METHODS: A retrospective study of 19 patients with IH treated with
propranolol was conducted from January 2009 to March 2014. Data of individual IH
volume at the beginning and at least 6 months after the onset of treatment and
overall volume reduction by ultrasonographic measurement were obtained.
RESULTS: We observed a statistically significant IH volume reduction of
approximately 0.51 cm3 . This volume corresponds to an average reduction of 47%
in the final volume compared with the initial volume.
CONCLUSION: Ultrasonographic measurements contribute to demonstrate tumor
regression and IH response to propranolol. Thus, ultrasonography is an important
instrument to guide therapeutic strategies. |
Can the CEP290 gene mutations be targeted by AAV-mediated gene therapy? | The large size of the CEP290 gene prevents its use in adeno-associated virus (AAV)-mediated gene augmentation therapy. | As the most common subtype of Leber congenital amaurosis (LCA), LCA10 is a
severe retinal dystrophy caused by mutations in the CEP290 gene. The most
frequent mutation found in patients with LCA10 is a deep intronic mutation in
CEP290 that generates a cryptic splice donor site. The large size of the CEP290
gene prevents its use in adeno-associated virus (AAV)-mediated gene augmentation
therapy. Here, we show that targeted genomic deletion using the clustered
regularly interspaced short palindromic repeats (CRISPR)/Cas9 system represents
a promising therapeutic approach for the treatment of patients with LCA10
bearing the CEP290 splice mutation. We generated a cellular model of LCA10 by
introducing the CEP290 splice mutation into 293FT cells and we showed that guide
RNA pairs coupled with SpCas9 were highly efficient at removing the intronic
splice mutation and restoring the expression of wild-type CEP290. In addition,
we demonstrated that a dual AAV system could effectively delete an intronic
fragment of the Cep290 gene in the mouse retina. To minimize the immune response
to prolonged expression of SpCas9, we developed a self-limiting CRISPR/Cas9
system that minimizes the duration of SpCas9 expression. These results support
further studies to determine the therapeutic potential of CRISPR/Cas9-based
strategies for the treatment of patients with LCA10. |
Which proteins are controlling sterol metabolism in S. cerevisiae? | The yeast genome encodes seven oxysterol binding protein homologs, Osh1p-Osh7p, which have been implicated in regulating intracellular lipid and vesicular transport The yeast genome encodes seven oxysterol binding protein homologs, Osh1p-Osh7p, which have been implicated in regulating intracellular lipid and vesicular transport Upc2p, a transcription factor of the zinc cluster family, is an important regulator of sterol biosynthesis and azole drug resistance in Candida albicans | There is an intimate association between sterol biosynthesis in yeast and
aerobicity. Besides the requirement for molecular oxygen for the epoxidation of
squalene, cytochrome hemoproteins are involved in demethylation and desaturation
steps. Regulatory effects of hemes on sterol formation have been demonstrated
using specifically defective mutants of yeast. Heme competency participates in a
mechanism whereby wild-type cells are prevented from taking exogenous sterols
from the growth media. The multiple interactions of hemes and sterols appear to
be associated with the variously defined functions for sterols in the yeast
cells. Upc2p, a transcription factor of the zinc cluster family, is an important
regulator of sterol biosynthesis and azole drug resistance in Candida albicans.
To better understand Upc2p function in C. albicans, we used genomewide location
profiling to identify the transcriptional targets of Upc2p in vivo. A triple
hemagglutinin epitope, introduced at the C terminus of Upc2p, conferred a
gain-of-function effect on the fusion protein. Location profiling identified 202
bound promoters (P < 0.05). Overrepresented functional groups of genes whose
promoters were bound by Upc2p included 12 genes involved in ergosterol
biosynthesis (NCP1, ERG11, ERG2, and others), 18 genes encoding ribosomal
subunits (RPS30, RPL32, RPL12, and others), 3 genes encoding drug transporters
(CDR1, MDR1, and YOR1), 4 genes encoding transcription factors (INO2, ACE2,
SUT1, and UPC2), and 6 genes involved in sulfur amino acid metabolism (MET6,
SAM2, SAH1, and others). Bioinformatic analyses suggested that Upc2p binds to
the DNA motif 5'-VNCGBDTR that includes the previously characterized Upc2p
binding site 5'-TCGTATA. Northern blot analysis showed that increased binding
correlates with increased expression for the analyzed Upc2p targets (ERG11,
MDR1, CDR1, YOR1, SUT1, SMF12, and CBP1). The analysis of ERG11, MDR1, and CDR1
transcripts in wild-type and upc2Delta/upc2Delta strains grown under
Upc2p-activating conditions (lovastatin treatment and hypoxia) showed that Upc2p
regulates its targets in a complex manner, acting as an activator or as a
repressor depending upon the target and the activating condition. Taken
together, our results indicate that Upc2p is a key regulator of ergosterol
metabolism. They also suggest that Upc2p may contribute to azole resistance by
regulating the expression of drug efflux pump-encoding genes in addition to
ergosterol biosynthesis genes. |
Which enzymes are responsible for base J creation in Trypanosoma brucei? | The base is synthesized in a two-step pathway. Initially, a thymidine residue in DNA is hydroxylated by a thymidine hydroxylase (TH). This intermediate (HOMedU) is then glucosylated to form base J. Two proteins involved in J synthesis, JBP1 (J binding protein 1) and JBP2, contain a putative TH domain related to the family of Fe(2+)/2-oxoglutarate-dependent hydroxylases. JBP2 is a chromatin re-modeling protein that induces de novo J-synthesis, allowing JBP1, a J-DNA binding protein, to stimulate additional J-synthesis. A recent computational screen identified a possible candidate for the base J-associated glucosyltransferase (JGT) in trypanosomatid genomes. | Synthesis of the modified thymine base, beta-d-glucosyl-hydroxymethyluracil or
J, within telomeric DNA of Trypanosoma brucei correlates with the bloodstream
form specific epigenetic silencing of telomeric variant surface glycoprotein
genes involved in antigenic variation. In order to analyze the function of base
J in the regulation of antigenic variation, we are characterizing the regulatory
mechanism of J biosynthesis. We have recently proposed a model in which
chromatin remodeling by a SWI2/SNF2-like protein (JBP2) regulates the
developmental and de novo site-specific localization of J synthesis within
bloodstream form trypanosome DNA. Consistent with this model, we now show that
JBP2 (-/-) bloodstream form trypanosomes contain five-fold less base J and are
unable to stimulate de novo J synthesis in newly generated telomeric arrays. Base J is a hypermodified DNA base localized primarily to telomeric regions of
the genome of Trypanosoma brucei. We have previously characterized two
thymidine-hydroxylases (TH), JBP1 and JBP2, which regulate J-biosynthesis. JBP2
is a chromatin re-modeling protein that induces de novo J-synthesis, allowing
JBP1, a J-DNA binding protein, to stimulate additional J-synthesis. Here, we
show that both JBP2 and JBP1 are capable of stimulating de novo J-synthesis. We
localized the JBP1- and JBP2-stimulated J by anti-J immunoprecipitation and
high-throughput sequencing. This genome-wide analysis revealed an enrichment of
base J at regions flanking polymerase II polycistronic transcription units (Pol
II PTUs) throughout the T. brucei genome. Chromosome-internal J deposition is
primarily mediated by JBP1, whereas JBP2-stimulated J deposition at the
telomeric regions. However, the maintece of J at JBP1-specific regions is
dependent on JBP2 SWI/SNF and TH activity. That similar regions of Leishmania
major also contain base J highlights the functional importance of the modified
base at Pol II PTUs within members of the kinetoplastid family. The regulation
of J synthesis/localization by two THs and potential biological function of J in
regulating kinetoplastid gene expression is discussed. Base J is a DNA modification found in the genome of Trypanosoma brucei and all
other kinetoplastids analyzed, where it replaces a small fraction of Ts, mainly
in telomeric and chromosome-internal transcription initiation and termination
regions. The synthesis of base J is a two-step process whereby a specific T is
converted to HOMedU (hydroxymethyldeoxyuridine) and subsequently glucosylated to
generate J. The thymidine hydroxylases (JPB1 and JBP2) that catalyze the first
step have been characterized, but the identity of the glucosyltransferase
catalyzing the second step has proven elusive. Recent bioinformatic analysis by
Iyer et al. (Nucleic Acids Res 2013;41:7635) suggested that Tb927.10.6900
encodes the glucosyltransferase (HmdUGT) responsible for converting HOMedU to J
in T. brucei. We now present experimental evidence to validate this hypothesis;
null mutants of Tb927.10.6900 are unable to synthesize base J. Orthologues from
related kinetoplastids show only modest conservation, with several insertion
sequences found in those from Leishmania and related genera. |
Which method for subsampling of NGS reads requires only gene counts? | SamExploreR : exploring reproducibility and robustness of RNA-seq results based on SAM files.We introduce the subseq r package, which uses a novel efficient approach to perform this subsampling and to calculate informative metrics at each depth required to inform a broad range of functional and evolutionary studies.Our methods are broadly applicable for polymorphism discovery in moderate to large genomes even at highly diverged loci, and we established by subsampling the illumina sbs coverage depth related questions for the experimental design.SubSeq : determining appropriate sequencing depth through efficient read subsampling. | Whole-genome hybridization studies have suggested that the nuclear genomes of
accessions (natural strains) of Arabidopsis thaliana can differ by several
percent of their sequence. To examine this variation, and as a first step in the
1001 Genomes Project for this species, we produced 15- to 25-fold coverage in
Illumina sequencing-by-synthesis (SBS) reads for the reference accession, Col-0,
and two divergent strains, Bur-0 and Tsu-1. We aligned reads to the reference
genome sequence to assess data quality metrics and to detect polymorphisms.
Alignments revealed 823,325 unique single nucleotide polymorphisms (SNPs) and
79,961 unique 1- to 3-bp indels in the divergent accessions at a specificity of
>99%, and over 2000 potential errors in the reference genome sequence. We also
identified >3.4 Mb of the Bur-0 and Tsu-1 genomes as being either extremely
dissimilar, deleted, or duplicated relative to the reference genome. To obtain
sequences for these regions, we incorporated the Velvet assembler into a
targeted de novo assembly method. This approach yielded 10,921 high-confidence
contigs that were anchored to flanking sequences and harbored indels as large as
641 bp. Our methods are broadly applicable for polymorphism discovery in
moderate to large genomes even at highly diverged loci, and we established by
subsampling the Illumina SBS coverage depth required to inform a broad range of
functional and evolutionary studies. Our pipeline for aligning reads and
predicting SNPs and indels, SHORE, is available for download at
http://1001genomes.org. High-throughput quantitative DNA sequencing enables the parallel phenotyping of
pools of thousands of mutants. However, the appropriate analytical methods and
experimental design that maximize the efficiency of these methods while
maintaining statistical power are currently unknown. Here, we have used Bar-seq
analysis of the Saccharomyces cerevisiae yeast deletion library to
systematically test the effect of experimental design parameters and sequence
read depth on experimental results. We present computational methods that
efficiently and accurately estimate effect sizes and their statistical
significance by adapting existing methods for RNA-seq analysis. Using simulated
variation of experimental designs, we found that biological replicates are
critical for statistical analysis of Bar-seq data, whereas technical replicates
are of less value. By subsampling sequence reads, we found that when using
four-fold biological replication, 6 million reads per condition achieved 96%
power to detect a two-fold change (or more) at a 5% false discovery rate. Our
guidelines for experimental design and computational analysis enables the study
of the yeast deletion collection in up to 30 different conditions in a single
sequencing lane. These findings are relevant to a variety of pooled genetic
screening methods that use high-throughput quantitative DNA sequencing,
including Tn-seq. MOTIVATION: Next-generation sequencing experiments, such as RNA-Seq, play an
increasingly important role in biological research. One complication is that the
power and accuracy of such experiments depend substantially on the number of
reads sequenced, so it is important and challenging to determine the optimal
read depth for an experiment or to verify whether one has adequate depth in an
existing experiment.
RESULTS: By randomly sampling lower depths from a sequencing experiment and
determining where the saturation of power and accuracy occurs, one can determine
what the most useful depth should be for future experiments, and furthermore,
confirm whether an existing experiment had sufficient depth to justify its
conclusions. We introduce the subSeq R package, which uses a novel efficient
approach to perform this subsampling and to calculate informative metrics at
each depth.
AVAILABILITY AND IMPLEMENTATION: The subSeq R package is available at
http://github.com/StoreyLab/subSeq/. Author information:
(1)Computational Biology and Machine Learning Laboratory, Center for Cancer
Research and Cell Biology, School of Medicine, Dentistry and Biomedical
Sciences, Faculty of Medicine Health and Life Sciences, Queen's University
Belfast, BT9 7AE Belfast, UK.
(2)Computational Biology and Machine Learning Laboratory, Center for Cancer
Research and Cell Biology, School of Medicine, Dentistry and Biomedical
Sciences, Faculty of Medicine Health and Life Sciences, Queen's University
Belfast, BT9 7AE Belfast, UK School of Mathematics and Physics, Queen's
University Belfast, BT7 1NN Belfast, UK.
(3)Northern Ireland Molecular Pathology Laboratory, Centre for Cancer Research
and Cell Biology, Queen's University Belfast, BT9 7AE Belfast, UK.
(4)Division of Biomedical Informatics, University of Arkansas for Medical
Sciences, Little Rock, AR 72205, USA.
(5)Department of Biomedical Computer Science and Mechatronics, UMIT, Hall in
Tirol, Austria College of Computer and Control Engineering, Nankai University,
Tianjin, P.R. China.
(6)Computational Biology and Machine Learning Laboratory, Center for Cancer
Research and Cell Biology, School of Medicine, Dentistry and Biomedical
Sciences, Faculty of Medicine Health and Life Sciences, Queen's University
Belfast, BT9 7AE Belfast, UK Predictive Medicine and Analytics Lab, Department
of Signal Processing, Tampere University of Technology, 33720 Tampere, Finland
Institute of Biosciences and Medical Technology, 33720 Tampere, Finland. |
Which topoisomerase is essential in yeast? | Eukaryotic DNA topoisomerase II is an abundant nuclear enzyme that is essential for cell proliferation. Yeast DNA topoisomerase II is encoded by a single-copy, essential gene. | The decatenation activity of DNA topoisomerase II is essential for viability as
eukaryotic cells traverse mitosis. Phosphorylation has been shown to stimulate
topoisomerase II activity in vitro. Here we show that topoisomerase II is a
phosphoprotein in yeast and that the level of incorporated phosphate is
significantly higher at mitosis than in G1. Comparison of tryptic phosphopeptide
maps reveals that the major phosphorylation sites in vivo are targets for casein
kinase II. Incorporation of phosphate into topoisomerase II is nearly
undetectable at the non-permissive temperature in a conditional casein kinase II
mutant. The sites modified by casein kinase II are located in the extreme
C-terminal domain of topoisomerase II. This domain is absent in prokaryotic and
highly divergent among eukaryotic type II topoisomerases, and may serve to
regulate functions of topoisomerase II that are unique to eukaryotic cells. The gene encoding topoisomerase II in yeast is unique and essential, required
for both mitotic and meiotic proliferation. The use of temperature-sensitive
mutants in topoisomerase II have demonstrated roles in the relaxation of
tortional stress, reduction of recombination rates, and in the separation of
sister chromatids after replication. In vertebrate cells, topoisomerase II was
shown to be the most abundant component of the metaphase chromosomal scaffold,
and has been shown to play a role in chromosome condensation in vitro. The cell
cycle control of chromosome condensation may well require phosphorylation of
topoisomerase II, since the enzyme is more highly phosphorylated in metaphase
than in G1. Recent studies have identified casein kinase II as the major enzyme
phosphorylating topoisomerase II in intact yeast cells. The target sites of CKII
are exclusively in the C-terminal 400 amino acids of topoisomerase II, the
region that is most divergent among the eukaryotic type II enzymes and which is
absent in the bacterial gyrase homologues. Studies with yeast DNA topoisomerase mutants indicate that neither topoisomerase
I nor II appears to be essential for transcription by RNA polymerase II.
However, plasmids carrying transcriptionally active genes are found to be
extremely negatively supercoiled when isolated from mutants lacking
topoisomerase I. Supercoiling occurs during transcriptional elongation rather
than during transcriptional activation. It takes place in the absence of
topoisomerase I and does not seem to be dependent on topoisomerase II since it
can occur at the nonpermissive temperature in a top1-top2 ts mutant. Whether
this change in linking number is due to an unusual form of topoisomerase II or
whether it is due to a new enzyme has yet to be determined. The results suggest
that topoisomerase I is normally required to relax transcriptionally induced
supercoils. A model is discussed which considers the role of topoisomerases in
the movement of RNA polymerase along the DNA template. Since DNA topoisomerase II (EC 5.99.1.3) is an essential enzyme in yeast,
heterologous topoisomerase II gene expression in yeast cells can provide a
system for analyzing the structure and function of topoisomerase II genes from
other species. A series of yeast expression plasmids was constructed in which
segments of the cDNA sequences encoding Drosophila DNA topoisomerase II were
inserted under the transcriptional control of yeast GAL1 promoter. Expression of
the functional form of Drosophila topoisomerase II cDNA can complement
conditionally lethal, temperature-sensitive mutations in the yeast topoisomerase
II gene (TOP2), as well as mutations in which the TOP2 locus was disrupted. The
survival of these yeast cells depends upon the continuous expression of
Drosophila topoisomerase II. Repression of Drosophila gene expression by glucose
causes these yeast cells to cease dividing after a few generations. In addition
to these genetic complementation data, the expression of the Drosophila
topoisomerase II gene in yeast cells with a disruption in TOP2 can also be
detected by immunochemical methods with an antibody specific for Drosophila
topoisomerase II. We have isolated mutants defective in DNA topoisomerases and an endonuclease
from the fission yeast Schizosaccharomyces pombe by screening individual
extracts of mutagenized cells. Two type I topoisomerase mutants (top1) and three
endonuclease mutants (end1) were all viable. The double mutant top1 end1 was
also viable and, in its extract, Mg2+- and ATP- dependent type II activity could
be detected. Three temperature-sensitive (ts-) mutants having heat-sensitive
(hs-) type II enzymes were isolated, and the ts- marker cosegregated with the
hs- type II activity. All the ts- mutations fell in one gene (top2) tightly
linked to leul in chromosome II. The nuclear division of single top2 mutants was
blocked at the restrictive temperature, but the formation of a septum was not
inhibited so that the nucleus was cut across with the cell plate. In contrast,
the double top1 top2 mutants were rapidly arrested at various stages of the cell
cycle, showing a strikingly altered nuclear chromatin region. The type II
topoisomerase may have an essential role in the compaction and/or segregation of
chromosomes during the nuclear division but also complement the defect of the
type I enzyme whose major function is the maintece of chromatin organization
throughout the cell cycle. The gene TOP2 encoding yeast topoisomerase II has been cloned by immunological
screening of a yeast genomic library constructed in the phage lambda expression
vector, lambda gt11. The ends of the message encoded by the cloned DNA fragment
were delimited by the Berk and Sharp procedure (S1 nuclease mapping) for the 5'
end and mapping of the polyA tail portion of a cDNA fragment for the 3' end. The
predicted size of the message agrees with the length of the message as
determined by Northern blot hybridization analysis. The identity of the gene was
confirmed by expressing the gene in E. coli from the E. coli promoter lac UV5 to
give catalytically active yeast DNA topoisomerase II. Disruption of one copy of
the gene in a diploid yeast creates a recessive lethal mutation, indicating that
the single DNA topoisomerase II gene of yeast has an essential function. Eukaryotic DNA topoisomerase II is an abundant nuclear enzyme that is essential
for cell proliferation. This homodimeric enzyme catalyzes the cleavage and
re-ligation of double-stranded DNA required to separate replicated sister
chromatids. Both biochemical and genetic studies show that its catalytic
activity is required for chromosome condensation and segregation, and that its
decatenation activity can be stimulated by a variety of protein kinases in
vitro. In budding yeast, topoisomerase II is most highly phosphorylated in
metaphase, and casein kinase II (CKII) was shown to be the major kinase
modifying topoisomerase II. We have investigated the effects of phosphorylation
of yeast topoisomerase II by CKII in vitro, by means of gel-retardation and
filter binding assays. The phosphorylation of the C terminus of topoisomerase II
by CKII appears to increase the stability of the complex formed with linear DNA
fragments, while dephosphorylation has the opposite effect. Rephosphorylation of
phosphatase-treated topoisomerase II by chicken casein kinase II restores a
stable protein-DNA complex using a linear DNA fragment. The enhanced stability
of the topoisomerase II-DNA complex is also observed with relaxed circular DNA,
but not with supercoiled minicircles, in agreement with published results using
topoisomerase II from Drosophila. Limited proteolysis and probing with
domain-specific antibodies shows that, with the exception of a weakly modified
residue between amino acid residues 660 and 1250, all residues modified by
casein kinase II are in the last 180 amino acid residues of yeast topoisomerase
II. Topoisomerase II is a ubiquitous enzyme that removes knots and tangles from the
genetic material by generating transient double-strand DNA breaks. While the
enzyme cannot perform its essential cellular functions without cleaving DNA,
this scission activity is inherently dangerous to chromosomal integrity. In
fact, etoposide and other clinically important anticancer drugs kill cells by
increasing levels of topoisomerase II-mediated DNA breaks. Cells rely heavily on
recombination to repair double-strand DNA breaks, but the specific pathways used
to repair topoisomerase II-generated DNA damage have not been defined.
Therefore, Saccharomyces cerevisiae was used as a model system to delineate the
recombination pathways that repair DNA breaks generated by topoisomerase II.
Yeast cells that expressed wild-type or a drug-hypersensitive mutant
topoisomerase II or overexpressed the wild-type enzyme were examined. Based on
cytotoxicity and recombination induced by etoposide in different
repair-deficient genetic backgrounds, double-strand DNA breaks generated by
topoisomerase II appear to be repaired primarily by the single-strand invasion
pathway of homologous recombination. Non-homologous end joining also was
triggered by etoposide treatment, but this pathway was considerably less active
than single-strand invasion and did not contribute significantly to cell
survival in S.cerevisiae. Topoisomerase II (Topo II) performs topological modifications on double-stranded
DNA molecules that are essential for chromosome condensation, resolution, and
segregation. In mammals, G2 and metaphase cell cycle delays induced by Topo II
poisons have been proposed to be the result of checkpoint activation in response
to the catenation state of DNA. However, the apparent lack of such controls in
model organisms has excluded genetic proof that Topo II checkpoints exist and
are separable from the conventional DNA damage checkpoint controls. But here, we
define a Topo II-dependent G2/M checkpoint in a genetically amenable eukaryote,
budding yeast, and demonstrate that this checkpoint enhances cell survival.
Conversely, a lack of the checkpoint results in aneuploidy. Neither DNA
damage-responsive pathways nor Pds1/securin are needed for this checkpoint.
Unusually, spindle assembly checkpoint components are required for the Topo II
checkpoint, but checkpoint activation is not the result of failed chromosome
biorientation or a lack of spindle tension. Thus, compromised Topo II function
activates a yeast checkpoint system that operates by a novel mechanism. Type II topoisomerases are essential for resolving topologically entwined
double-stranded DNA. Although anti-topoisomerase 2 (Top2) drugs are clinically
important antibiotics and chemotherapies, to our knowledge, the mechanisms of
cell killing by Top2 depletion and inactivation have never been directly
compared. We show that depletion of Top2 protein from budding yeast cells
prevents DNA decatenation during S phase. Cells complete DNA replication and
enter the ensuing mitosis on schedule, suffering extensive chromosome
missegregation. Cytokinesis through incompletely segregated chromosomes causes
lethal DNA damage. By contrast, expression of catalytically inactive Top2 causes
a stable G2 arrest requiring an intact DNA damage checkpoint. Checkpoint
activation correlates with an inability to complete DNA replication, resulting
in hypercatenated, gapped daughter DNA molecules. Thus, Top2 depletion and
inactivation kill cells by different mechanisms, which has implications for
understanding the nature of the catenation checkpoint, how DNA replication
terminates, how anti-Top2 drugs work, and how new drugs might be designed. DNA topoisomerases are specialized nuclear enzymes that perform topological
modifications on double-stranded DNA (dsDNA) and hence are essential for DNA
metabolism such as replication, transcription, recombination, condensation and
segregation. In a genetic screen, we identified a temperature-sensitive mutant
allele of topoisomerase 2 that exhibits conditional synthetic lethality with a
chk1 knockout strain. The mutant allele of topoisomerase 2 is defective in
chromosome segregation at a non-permissive temperature and there was increase in
chromosome segregation defects in the double mutant of top2-10 and chk1 delete
at a non-permissive temperature. More importantly, topoisomearse 2 mutant cells
mildly delay the mitotic progression at non-permissive temperature that is
mediated by checkpoint protein kinase Chk1. Additionally, top2-10 mutant cells
also activate the Chk1 at a non-permissive temperature and this activation of
Chk1 takes place at the time of mitosis. Interestingly, top2-10 mutant cells
retain their viability at a non-permissive temperature if the cells are not
allowed to enter into mitosis. Taking together our results, we speculate that in
the top2-10 mutant, the segregation of entangled chromatids during mitosis could
result in delaying the mitotic progression through the activation of Chk1
kinase. Eukaryotic topoisomerase II (topo II) is the essential decatenase of newly
replicated chromosomes and the main relaxase of nucleosomal DNA. Apart from
these general tasks, topo II participates in more specialized functions. In
mammals, topo IIα interacts with specific RNA polymerases and
chromatin-remodeling complexes, whereas topo IIβ regulates developmental genes
in conjunction with chromatin remodeling and heterochromatin transitions. Here
we show that in budding yeast, topo II regulates the expression of specific gene
subsets. To uncover this, we carried out a genomic transcription run-on shortly
after the thermal inactivation of topo II. We identified a modest number of
genes not involved in the general stress response but strictly dependent on topo
II. These genes present distinctive functional and structural traits in
comparison with the genome average. Yeast topo II is a positive regulator of
genes with well-defined promoter architecture that associates to chromatin
remodeling complexes; it is a negative regulator of genes extremely
hypo-acetylated with complex promoters and undefined nucleosome positioning,
many of which are involved in polyamine transport. These findings indicate that
yeast topo II operates on singular chromatin architectures to activate or
repress DNA transcription and that this activity produces functional responses
to ensure chromatin stability. |
What is the genetic basis of Ohdo syndrome? | MED12 cause X-linked Ohdo syndromeIn | We report a series of eight patients with the Say/Barber/Biesecker/Young-Simpson
(SBBYS) type of Ohdo syndrome, which is the largest cohort described to date. We
expand on the type, frequency and severity of the clinical characteristics in
this condition; comment on the natural history of Ohdo syndrome and further
refine previously published diagnostic criteria. Cytogenetic investigations and
microarray CGH analysis undertaken in this cohort of patients failed to identify
a chromosomal aetiology. It remains possible that this rare condition is
heterogeneous and therefore caution must be undertaken during counselling until
the underlying genetic mechanism(s) is (are) identified. Ohdo syndrome comprises a heterogeneous group of disorders characterized by
intellectual disability (ID) and typical facial features, including
blepharophimosis. Clinically, these blepharophimosis-ID syndromes have been
classified in five distinct subgroups, including the Maat-Kievit-Brunner (MKB)
type, which, in contrast to the others, is characterized by X-linked inheritance
and facial coarsening at older age. We performed exome sequencing in two
families, each with two affected males with Ohdo syndrome MKB type. In the two
families, MED12 missense mutations (c.3443G>A [p.Arg1148His] or c.3493T>C
[p.Ser1165Pro]) segregating with the phenotype were identified. Upon subsequent
analysis of an additional cohort of nine simplex male individuals with Ohdo
syndrome, one additional de novo missense change (c.5185C>A [p.His1729Asn]) in
MED12 was detected. The occurrence of three different hemizygous missense
mutations in three unrelated families affected by Ohdo syndrome MKB type shows
that mutations in MED12 are the underlying cause of this X-linked form of Ohdo
syndrome. Together with the recently described KAT6B mutations resulting in Ohdo
syndrome Say/Barber/Biesecker/Young/Simpson type, our findings point to aberrant
chromatin modification as being central to the pathogenesis of Ohdo syndrome. FG syndrome, Lujan syndrome, and Ohdo syndrome, the Maat-Kievit-Brunner type,
have been described as distinct syndromes with overlapping non-specific features
and different missense mutations of the MED12 gene have been reported in all of
them. We report a family including 10 males and 1 female affected with profound
non-specific intellectual disability (ID) which was linked to a 30-cM region
extending from Xp11.21 (ALAS2) to Xq22.3 (COL4A5). Parallel sequencing of all
X-chromosome exons identified a frameshift mutation (c.5898dupC) of MED12.
Mutated mRNA was not affected by non-sense mediated RNA decay and induced an
additional abnormal isoform due to activation of cryptic splice-sites in exon
41. Dysmorphic features common to most affected males were long narrow face,
high forehead, flat malar area, high nasal bridge, and short philtrum. Language
was absent or very limited. Most patients had a friendly personality. Cognitive
impairment, varying from borderline to profound ID was similarly observed in
seven heterozygous females. There was no correlation between cognitive function
and X-chromosome inactivation profiles in blood cells. The severe degree of ID
in male patients, as well as variable cognitive impairment in heterozygous
females suggests that the duplication observed in the present family may have a
more severe effect on MED12 function than missense mutations. In a cognitively
impaired male from this family, who also presented with tall stature and
dysmorphism and did not have the MED12 mutation, a 600-kb duplication at 17p13.3
including the YWHAE gene, was found in a mosaic state. MED12: is a member of the large Mediator complex, which has a critical and
central role in RNA polymerase II transcription. As a multiprotien complex,
Mediator regulates signals involved in cell growth, development, and
differentiation, and it is involved in a protein network required for
extraneuronal gene silencing and also functions as a direct suppressor of
Gli3-dependent Sonic hedgehog signaling. This may explain its role in several
different X-linked intellectual disability syndromes that share some overlapping
clinical features. This review will compare and contrast four different clinical
conditions that have been associated with different mutations in MED12, which is
located at Xq13. To date, these conditions include Opitz-Kaveggia (FG) syndrome,
Lujan syndrome, Ohdo syndrome (Maat-Kievit-Brunner type, or OSMKB), and one
large family with profound X-linked intellectual disability due to a novel
c.5898insC frameshift mutation that unlike the other three syndromes, resulted
in affected female carriers and truncation of the MED12 protein. It is likely
that more MED12 mutations will be detected in sporadic patients and X-linked
families with intellectual disability and dysmorphic features as exome
sequencing becomes more commonly utilized, and this overview of MED12-related
disorders may help to correlate MED12 genotypes with clinical findings. We report on two male sibs, a fetus and a newborn, with short humeri and
dysmorphic facial features including blepharophimosis. The newborn also had
Hirschsprung disease. Goldberg-Shprintzen syndrome and the
Say-Barber-Biesecker-Young-Simpson type of Ohdo syndrome were suspected but
direct sequencing of KBP and KAT6B failed to identify a mutation. Finally,
direct sequencing of MED12, the gene mutated in Opitz-Kaveggia syndrome,
Lujan-Fryns syndrome and X-linked Ohdo syndrome identified in the two sibs the
missense mutation c.3443G>A (p.Arg1148His) inherited from the mother. This
report further expands the phenotypic spectrum of MED12 mutations. |
Which drugs were tested in the KEYNOTE-006 study? | KEYNOTE-006 study compared pembrolizumab versus ipilimumab for advanced melanoma. | BACKGROUND: The immune checkpoint inhibitor ipilimumab is the standard-of-care
treatment for patients with advanced melanoma. Pembrolizumab inhibits the
programmed cell death 1 (PD-1) immune checkpoint and has antitumor activity in
patients with advanced melanoma.
METHODS: In this randomized, controlled, phase 3 study, we assigned 834 patients
with advanced melanoma in a 1:1:1 ratio to receive pembrolizumab (at a dose of
10 mg per kilogram of body weight) every 2 weeks or every 3 weeks or four doses
of ipilimumab (at 3 mg per kilogram) every 3 weeks. Primary end points were
progression-free and overall survival.
RESULTS: The estimated 6-month progression-free-survival rates were 47.3% for
pembrolizumab every 2 weeks, 46.4% for pembrolizumab every 3 weeks, and 26.5%
for ipilimumab (hazard ratio for disease progression, 0.58; P<0.001 for both
pembrolizumab regimens versus ipilimumab; 95% confidence intervals [CIs], 0.46
to 0.72 and 0.47 to 0.72, respectively). Estimated 12-month survival rates were
74.1%, 68.4%, and 58.2%, respectively (hazard ratio for death for pembrolizumab
every 2 weeks, 0.63; 95% CI, 0.47 to 0.83; P=0.0005; hazard ratio for
pembrolizumab every 3 weeks, 0.69; 95% CI, 0.52 to 0.90; P=0.0036). The response
rate was improved with pembrolizumab administered every 2 weeks (33.7%) and
every 3 weeks (32.9%), as compared with ipilimumab (11.9%) (P<0.001 for both
comparisons). Responses were ongoing in 89.4%, 96.7%, and 87.9% of patients,
respectively, after a median follow-up of 7.9 months. Efficacy was similar in
the two pembrolizumab groups. Rates of treatment-related adverse events of grade
3 to 5 severity were lower in the pembrolizumab groups (13.3% and 10.1%) than in
the ipilimumab group (19.9%).
CONCLUSIONS: The anti-PD-1 antibody pembrolizumab prolonged progression-free
survival and overall survival and had less high-grade toxicity than did
ipilimumab in patients with advanced melanoma. (Funded by Merck Sharp & Dohme;
KEYNOTE-006 ClinicalTrials.gov number, NCT01866319.). BACKGROUND: Recent clinical trials have shown that pembrolizumab significantly
prolonged progression-free survival and overall survival compared with
ipilimumab in ipilimumab-naïve patients with unresectable or metastatic
melanoma. However, there has been no published evidence on the
cost-effectiveness of pembrolizumab for this indication.
OBJECTIVE: To assess the long-term cost-effectiveness of pembrolizumab versus
ipilimumab in ipilimumab-naïve patients with unresectable or meta-static
melanoma from a U.S. integrated health system perspective.
METHODS: A partitioned-survival model was developed, which divided overall
survival time into progression-free survival and postprogression survival. The
model used Kaplan-Meier estimates of progression-free survival and overall
survival from a recent randomized phase 3 study (KEYNOTE-006) that compared
pembrolizumab and ipilimumab. Extrapolation of progression-free survival and
overall survival beyond the clinical trial was based on parametric functions and
literature data. The base-case time horizon was 20 years, and costs and health
outcomes were discounted at a rate of 3% per year. Clinical data-including
progression-free survival and overall survival data spanning a median follow-up
time of 15 months, as well as quality of life and adverse event data from the
ongoing KEYNOTE-006 trial-and cost data from public sources were used to
populate the model. Costs included those of drug acquisition, treatment
administration, adverse event management, and disease management of advanced
melanoma. The incremental cost-effectiveness ratio (ICER) expressed as cost
difference per quality-adjusted life-year (QALY) gained was the main outcome,
and a series of sensitivity analyses were performed to test the robustness of
the results.
RESULTS: In the base case, pembrolizumab was projected to increase the life
expectancy of U.S. patients with advanced melanoma by 1.14 years, corresponding
to a gain of 0.79 discounted QALYs over ipilimumab. The model also projected an
average increase of $63,680 in discounted perpatient costs of treatment with
pembrolizumab versus ipilimumab. The corresponding ICER was $81,091 per QALY
($68,712 per life-year) over a 20-year time horizon. With $100,000 per QALY as
the threshold, when input parameters were varied in deterministic one-way
sensitivity analyses, the use of pembrolizumab was cost-effective relative to
ipilimumab in most ranges. Further, in a comprehensive probabilistic sensitivity
analysis, the ICER was cost-effective in 83% of the simulations.
CONCLUSIONS: Compared with ipilimumab, pembrolizumab had higher expected QALYs
and was cost-effective for the treatment of patients with unresectable or
metastatic melanoma from a U.S. integrated health system perspective.
DISCLOSURES: This study was supported by funding from Merck & Co., which
reviewed and approved the manuscript before journal submission. Wang,
Pellissier, Xu, Stevinson, and Liu are employees of, and own stock in, Merck &
Co. Chmielowski has served as a paid consultant for Merck & Co. and received a
consultant fee for clinical input in connection with this study. Chmielowski
also reports receiving advisory board and speaker bureau fees from multiple
major pharmaceutical companies. Wang led the modeling and writing of the
manuscript. Chmielowski, Xu, Stevinson, and Pellissier contributed substantially
to the modeling design and methodology. Liu led the data collection work and
contributed substantially to writing the manuscript. In conducting the analysis
and writing the manuscript, the authors followed Merck publication polices and
the "cost-effectiveness analysis alongside clinical trials-good research
practices and the CHEERS reporting format as recommended by the International
Society for Pharmacoeconomics and Outcomes Research. Author information:
(1)Sunnybrook Health Sciences Centre, University of Toronto, 2075 Bayview Ave,
T2-041, Toronto, ON, M4N 3M5, Canada. Electronic address:
[email protected].
(2)Gustave Roussy and Université Paris-Sud, 114 Rue Edouard Vaillant, 94800
Villejuif, France. Electronic address: [email protected].
(3)Medical University of Graz, Auenbruggerpl. 2, 8036 Graz, Graz, Austria.
Electronic address: [email protected].
(4)Segal Cancer Centre, Jewish General Hospital, Rossy Cancer Network, and
McGill University, 3755 Ch de la Côte-Sainte-Catherine, Montreal, QC, H3T 1E2,
Canada. Electronic address: [email protected].
(5)Karolinska Institute, Solnavägen 1, 171 77 Solna, Stockholm, Sweden.
Electronic address: [email protected].
(6)Princess Alexandra Hospital and The University of Queensland, 199 Ipswich Rd,
Woolloongabba, Brisbane, QLD 4102, Australia. Electronic address:
[email protected].
(7)APHP, Dermatology and CIC, Université Paris Diderot, Hôpital Saint-Louis, 1
Avenue Claude Vellefaux, 75010 Paris, France. Electronic address:
[email protected].
(8)Queen Elizabeth Hospital Birmingham, Mindelsohn Way, Birmingham B15 2TH, UK.
Electronic address: [email protected].
(9)The Churchill Hospital and The University of Oxford, Old Rd, Headington,
Oxford OX3 7LE, UK. Electronic address: [email protected].
(10)Merck & Co., Inc., 2000 Galloping Hill Road, Kenilworth, NJ 07033, USA.
Electronic address: [email protected].
(11)Merck & Co., Inc., 2000 Galloping Hill Road, Kenilworth, NJ 07033, USA.
Electronic address: [email protected].
(12)Merck & Co., Inc., 2000 Galloping Hill Road, Kenilworth, NJ 07033, USA.
Electronic address: [email protected].
(13)Olivia Newton-John Cancer Research Institute, Austin Health, School of
Cancer Medicine, La Trobe University, 145 Studley Road, Heidelberg VIC 3084,
Melbourne, Australia. Electronic address: [email protected]. |
What is the purpose of the FRAX scale? | The FRAX score (The WHO Fracture Risk Assessment Tool), is a free web-based clinical scale assessing the 10-year probability of major osteoporotic fracture risk and need for lifestyle advice/reassurance, dual X-ray absorptiometry (DEXA) scanning or preventive treatment. | AIM: Osteoporosis is a frequent comorbidity in patients with chronic obstructive
pulmonary disease (COPD). We have studied the risk of major osteoporotic
fracture and hip fracture in patients with COPD.
PATIENTS AND METHODS: A multicenter cross-sectional study was performed in Spain
in 26 hospitals of 16 regional communities. Patients diagnosed with COPD who
required admission to the Internal Medicine Service due to exacerbation of their
respiratory disease were enrolled. COPD was confirmed by post-bronchodilator
spirometry in stable state: maximum expiratory volume in the first second (FEV₁)
< 80% of the theoretical value and quotient FEV(1)/FVC < 0.70 and percent
predicted after the administration of a bronchodilator. Dyspnea was evaluated
with the modified Medical Research Council (mMRC) dyspnea scale. The principal
variable was the likelihood of fracture evaluated with the FRAX® tool for the
Spanish population.
RESULTS: Three hundred and ninety two patients, 347 (88%) men, with a mean (SD)
age of 73.7 (8.9) years and a mean FEV₁ of 1.23 liters (43.3% of predicted) were
enrolled. Only 37 patients (9.4%), 27 men and 10 women had been diagnosed
previously of osteoporosis. Overall, 1.8% (95% CI: 0.9-3.6) had a 10-year
probability of major osteoporotic fracture ≥ 20% and 49.7% (95% CI: 44.8-54.7)
had a probability of hip fracture ≥ 3%. No relationship was observed between the
probability of fracture and GOLD stage or mMRC dyspnea scale.
CONCLUSIONS: The diagnosis of osteoporosis is uncommon in our COPD patients.
However, half of them have a high probability of a hip fracture in the next 10
years. Bone mineral density (BMD) alone does not reliably predict osteoporotic
fractures. The Fracture Risk Assessment Tool (FRAX) was developed to estimate
the risk of fracture in the general population. This study was designed to
identify predictors of osteoporosis and vertebral fractures in patients
presenting with chronic obstructive pulmonary disease (COPD). We studied 85
patients (mean age = 75 years; 92% men) with moderate to very severe COPD.
Osteoporosis and vertebral fractures were diagnosed with dual energy X-ray
absorptiometric scan and vertebral X-rays, respectively. Patient
characteristics, including age, gender, body mass index (BMI), and results of
pulmonary function tests, chest computed tomography scan, blood and urinary
biomarkers of bone turnover were recorded, and a FRAX score was calculated by a
computer-based algorithm. Osteoporosis, defined as a T score < -2.5, found in 20
patients (24%), was associated with female gender, BMI, dyspnea scale, long-term
oxygen therapy (LTOT), vital capacity (VC), emphysema score on computed
tomography, measurements of serum and urinary biomarkers of bone turnover.
Vertebral fractures, diagnosed in 29 patients (35%), were strongly correlated
with age, LTOT, VC, and forced expiratory volume in 1 sec, treatment with oral
corticosteroid or warfarin, and weakly associated with the presence of
osteoporosis. There was no correlation between FRAX score and prevalence of
vertebral fractures, suggesting that neither BMD alone nor FRAX score would
predict the presence of vertebral fractures in COPD patients. A disease-specific
algorithm to predict osteoporotic fractures is needed to improve the management
of patients suffering from COPD. The WHO Fracture Risk Assessment Tool (FRAX; http://www.shef.ac.uk/FRAX)
estimates the 10-year probability of major osteoporotic fracture. Clodronate and
bazedoxifene reduced nonvertebral and clinical fracture more effectively on a
relative scale in women with higher FRAX scores. We used data from the Fracture
Intervention Trial (FIT) to evaluate the interaction between FRAX score and
treatment with alendronate. We combined the Clinical Fracture (CF) arm and
Vertebral Fracture (VF) arm of FIT. The CF and VF arm of FIT randomized 4432 and
2027 women, respectively, to placebo or alendronate for 4 and 3 years,
respectively. FRAX risk factors were assessed at baseline. FRAX scores were
calculated by WHO. We used Poisson regression models to assess the interaction
between alendronate and FRAX score on the risk of nonvertebral, clinical, major
osteoporotic, and radiographic vertebral fractures. Overall, alendronate
significantly reduced the risk of nonvertebral fracture (incidence rate ratio
[IRR] 0.86; 95% confidence interval [CI], 0.75-0.99), but the effect was greater
for femoral neck (FN) bone mineral density (BMD) T-score ≤ -2.5 (IRR 0.76; 95%
CI, 0.62-0.93) than for FN T-score > -2.5 (IRR 0.96; 95% CI, 0.80-1.16)
(p = 0.02, interaction between alendronate and FN BMD). However, there was no
evidence of an interaction between alendronate and FRAX score with FN BMD for
risk of nonvertebral fracture (interaction p = 0.61). The absolute benefit of
alendronate was greatest among women with highest FRAX scores. Results were
similar for clinical fractures, major osteoporotic fractures, and radiographic
vertebral fractures and whether or not FRAX scores included FN BMD. Among this
cohort of women with low bone mass there was no significant interaction between
FRAX score and alendronate for nonvertebral, clinical or major osteoporotic
fractures, or radiographic vertebral fractures. These results suggest that the
effect of alendronate on a relative scale does not vary by FRAX score. A
randomized controlled trial testing the effect of antifracture agents among
women with high FRAX score but without osteoporosis is warranted. OBJECTIVE: Osteoporosis and hypogonadism are common in men with HIV infection.
Ageing Male Symptoms (AMS) scale measures symptoms related to hypogonadism. FRAX
provides 10-year probability of major fractures. We investigated the role of AMS
scale combined with FRAX without bone mineral density (BMD), in identifying HIV
men with bone fragility.
DESIGN: Cross-sectional observational study.
METHODS: Fifty HIV-positive men treated with highly active antiretroviral
therapy and 27 controls underwent hormonal evaluation, BMD scan and spine X-ray.
The AMS questionnaire was administered.
RESULTS: Osteoporosis was found in 24·0% of HIV patients and in 3·7% of controls
(P = 0·05). In HIV patients, 9 radiological vertebral fractures were found (none
in controls, P = 0·04). Calculated free testosterone suggested hypogonadism in
26% of HIV patients vs 4% of controls (P = 0·04); an abnormal AMS score (≥27)
was found in 62% HIV patients compared with 41% controls (P = 0·04). ROC curves
showed that FRAX for major fracture had a 23% sensitivity and a 100% specificity
in identifying HIV patients with bone fragility (P = 0·002, with the threshold
of 7% at which bisphosphonate therapy is cost-effective). Considering a value of
AMS ≥27, we obtained an 82·6% sensitivity and a 42·9% specificity (P = 0·04).
The combination of AMS and FRAX score achieved a 77·3% sensitivity and a 69%
specificity (P = 0·02, cut-off 34).
CONCLUSION: Combination of FRAX (without BMD) and AMS improved sensitivity of
FRAX alone in identifying HIV patients at fracture risk, at the expense of
reduced specificity. INTRODUCTION: In 1998, the first Japanese practice guidelines on osteoporosis
was published. It has been updated several times, with the most recent being the
full-scale 2011 edition and its abridged edition. The present guidelines provide
information for the managements of primary osteoporosis in postmenopausal women
and men over 50 years old, a summary of the evidence for the treatment of
secondary osteoporosis, and a summary of the evidence for the prevention of
osteoporosis in younger people.
METHOD: The present Executive Summary is primarily based on the content of the
2011 Japanese abridged edition. One of the key changes is revision of the
criteria for initiation of pharmacological treatment, along with an introduction
of the fracture risk factors used in FRAX®. Key figures and tables were selected
from the Japanese abridged edition and a reference list was added.
RESULT AND CONCLUSIONS: The essential points of the Japanese practice guidelines
on osteoporosis were translated into English for the first time. It is hoped
that the content of the guidelines becomes known throughout the world. INTRODUCTION: Currently identification, and therefore, management of patients at
risk of osteoporotic fracture in the UK is suboptimal. As the majority of
patients who fracture have fallen, it follows that people who fall can usefully
be targeted in any programme that aims to reduce osteoporotic fracture.
Targeting vulnerable patients who are likely to benefit from intervention may
help shift the management of fracture prevention into primary care, away from
emergency departments. Paramedics who attend to patients who have fallen may be
well placed to assess future fracture risk, using the Fracture Risk Assessment
Tool (FRAX) and communicate that information directly to general practitioners
(GPs).
METHODS AND ANALYSIS: This feasibility study takes the form of a pragmatic,
randomised controlled trial aimed at exploring and refining issues of study
design, recruitment, retention, sample size and acceptability preceding a
large-scale study with fracture as the end point. Patients (aged >50) who fall,
call an ambulance, are attended by a study paramedic and give verbal consent
will be asked FRAX and fall questions. Patients who subsequently formally
consent to participation will be randomised to control (usual care) or
intervention groups. Intervention will constitute transmission of calculated
future fracture risk to the patients' GP with suitable, evidence-based
recommendations for investigation or treatment. 3 months after the index fall,
data (proportion of patients in each group undergoing investigation or starting
new treatment, quality of life and health economic) will be collected and
analysed using descriptive statistics. A nested qualitative study will explore
issues of acceptability and study design with patients, paramedics and GPs.
ETHICS AND DISSEMINATION: This protocol was approved by NRES Committee South
Central Oxford C in October 2012. Research Ethics Committee ref.12/SC/0604. The
study findings will be disseminated through peer-reviewed journals, conference
presentations and local public events. A publication plan and authorship
criteria have been preagreed.
TRIAL REGISTRATION NUMBER ISRCTN: 36245726. PURPOSE: The WHO fracture risk prediction tool (FRAX®) utilises clinical risk
factors to estimate the probability of fracture over a 10-year period. Although
falls increase fracture risk, they have not been incorporated into FRAX. It is
currently unclear if FRAX captures falls risk and whether addition of falls
would improve fracture prediction. We aimed to investigate the association of
falls risk and Australian-specific FRAX.
METHODS: Clinical risk factors were documented for 735 men and 602 women (age
40-90 yr) assessed at follow-up (2006-2010 and 2000-2003, respectively) of the
Geelong Osteoporosis Study. FRAX scores with and without BMD were calculated. A
falls risk score was determined at the time of BMD assessment and self-reported
incident falls were documented from questionnaires returned one year later.
Multivariable analyses were performed to determine: (i) cross-sectional
association between FRAX scores and falls risk score (Elderly Falls Screening
Test, EFST) and (ii) prospective relationship between FRAX and time to a fall.
RESULTS: There was an association between FRAX (hip with BMD) and EFST scores (β
= 0.07, p < 0.001). After adjustment for sex and age, the relationship became
non-significant (β = 0.00, p = 0.79). The risk of incident falls increased with
increasing FRAX (hip with BMD) score (unadjusted HR 1.04, 95% CI 1.02, 1.07).
After adjustment for age and sex, the relationship became non-significant (1.01,
95% CI 0.97, 1.05).
CONCLUSIONS: There is a weak positive correlation between FRAX and falls risk
score, that is likely explained by the inclusion of age and sex in the FRAX
model. These data suggest that FRAX score may not be a robust surrogate for
falls risk and that inclusion of falls in fracture risk assessment should be
further explored. In this study, we compared subjective fracture risks of Hungarian women with
osteoporosis to FRAX®-based estimates. Patients with a previous fracture,
parental hip fracture, low femoral T-score, higher age, and higher BMI were more
likely to underestimate their risks. Patients also failed to associate risk
factors with an increased risk of fractures.
PURPOSE: The main objectives were to explore associations between self-perceived
10-year fracture risks of women with osteoporosis (OP) and their risks
calculated by the FRAX® algorithm and to identify determits of the
underestimation of risk.
METHODS: We carried out a cross-sectional study in 11 OP centers in Hungary and
collected data on the risk factors considered by the FRAX® calculator. Patients
estimated their subjective 10-year probability of any major osteoporotic and hip
fracture numerically, in percentages and also on a visual analog scale (VAS). We
compared subjective and FRAX® estimates and applied logistic regression to
analyze the determits of the underestimation of risk. Associations between
risk factors and subjective risk were explored using linear probability models.
RESULTS: Nine hundred seventy-two OP patients were included in the analysis.
Major OP and hip fracture risk by FRAX® were on average 20.1 and 10.5%, while
subjective estimates were significantly higher, 30.0 and 24.7%, respectively.
Correlations between FRAX® and subjective measures were very weak
(r = 0.12-0.16). Underestimation of major OP fracture risk was associated with
having had a single previous fracture (OR = 2.0), parental hip fracture
(OR = 3.4), femoral T-score ≤-2.5 (OR = 4.2), higher age, body mass index, and
better general health state. We did not find significant associations between
subjective risk estimates and most of the risk factors except for previous
fractures.
CONCLUSIONS: Hungarian OP patients fail to recognize most of the risk factors of
fractures. Thus, education of patients about these risk factors would be
beneficial especially for the elderly with a low femoral T-score and parental
hip fracture history. INTRODUCTION: Osteoporosis and cardiovascular diseases (CVD) are more common in
the elderly population and have similar risk factors. THE GOAL: was an
evaluation of the correlation between 10-year risk of death from CVD and 10-year
bone fracture risk (FRAX).
MATERIAL AND METHODS: A total of 79 patients of the Regional Centre of Menopause
and Osteoporosis of the Military Teaching Hospital in Lodz (Poland), aged 50-83
years, consulted for osteoporosis were divided into two groups: study group -
with osteoporosis (O; T-score ≤ -2.5 SD) and control - without osteoporosis
(T-sc > -2.5). Bone mineral density was evaluated by densitometric scanning of
spine (L2-L4 T-score) and/or femoral neck (Neck T-score) and/or total hip (Total
Hip T-score). Total cholesterol (TC), fasting glucose, arterial blood pressure,
medical history, and family history were obtained. The risk of fatal-CVD was
assessed by Euro Heart Score (EHS), and major osteoporotic (MOFR) and hip
fracture risk (HFR) by the FRAX scale.
RESULTS: 80% of the patients (32/40) with osteoporosis and 51% (20/39) of the
patients without osteoporosis revealed a HeartScore ≥ 5%. There was correlation
in the group of all patients between EHS and Neck T-score (p < 0.05; Spearman
rank correlation coefficient (Rs) = -0.3806), L2-L4 T-score (p < 0.05; Rs =
-0.2891), and Total Hip T-score (p < 0.005; Rs = -0.3561), and in the control
group - between EHS and Neck T-score (p < 0.05; Rs = -0.3502). There was a 2.33%
difference between the average EHS level to the disadvantage of patients with
osteoporosis (p < 0.05). EHS positively correlated with MOFR (p < 0.001) and HFR
(p < 0.001) in the whole study popula-tion and with MOFR (p < 0.05) and HFR (p <
0.01) in the group of osteoporotic patients. There were differences between
groups in TC (p < 0.001) and BMI (p < 0.001) levels.
CONCLUSIONS: The 10-year risk of fatal-CVD correlated with osteoporosis and with
the 10-year osteoporotic fracture risk. This conclusion may help identify
patients who require extended cardiotherapy protocols. |
Are mutations in the nf1 gene associated with memory? | Yes, distinct functional domains of neurofibromatosis type 1 regulate immediate versus long-term memory formation. | Neurofibromatosis type 1 (NF1) is a domit genetic disorder that causes tumors
of the peripheral nervous system. In addition, >40% of afflicted children have
learning difficulties. The NF1 protein contains a highly conserved
GTPase-activating protein domain that inhibits Ras activity, and the C-terminal
region regulates cAMP levels via G-protein-dependent activation of adenylyl
cyclase. Behavioral analysis indicates that learning is disrupted in both
Drosophila and mouse NF1 models. Our previous work has shown that defective cAMP
signaling leads to the learning phenotype in Drosophila Nf1 mutants. In the
present report, our experiments showed that in addition to learning, long-term
memory was also abolished in Nf1 mutants. However, altered NF1-regulated Ras
activity is responsible for this defect rather than altered cAMP levels.
Furthermore, by expressing clinically relevant human NF1 mutations and deletions
in Drosophila Nf1-null mutants, we demonstrated that the GAP-related domain of
NF1 was necessary and sufficient for long-term memory, whereas the C-terminal
domain of NF1 was essential for immediate memory. Thus, we show that two
separate functional domains of the same protein can participate independently in
the formation of two distinct memory components. Neurofibromatosis 1 (NF1) is a common single-gene disorder that causes learning
impairments in patients. Neurofibromin encoded by the NF1 causal gene regulates
Ras/MAPK and cAMP signaling pathways. These signaling pathways play critical
roles in controlling gene transcription during synaptic plasticity and memory
formation. We hypothesized that NF1 mutations disturb the expression of genes
important for memory formation. To test this hypothesis, we performed DNA
microarray analysis on the hippocampus of NF1(+/-) mice, the mouse model for NF1
learning disabilities. Our results indicated that genes involved in a wide
spectrum of biological processes are dysregulated in the NF1(+/-) hippocampus.
Many of the NF1-affected genes play critical roles in synaptic plasticity, such
as Rabs, synaptotagmins, NMDAR1, CaMKII, and CREB1. Because NF1-associated
learning disabilities can be reversed by lovastatin, we also determined the
effect of lovastatin treatment on genome-wide expression patterns of the
NF1(+/-) hippocampus. We found that lovastatin altered the expression of a large
number of genes, including those disturbed by NF1 mutations. Our results reveal
a genome-wide overview of the molecular abnormalities in the NF1(+/-)
hippocampus and should be useful for further identifying the novel molecular
pathways that cause NF1 learning deficits. |
Has Cas9 gene editing the potential to correct inhereted hearing loss? | CRISPR/Cas9-mediated genome editing can be efficiently performed in the mammalian inner ear in vivo.
The genetic correction of induced pluripotent stem cells (iPSCs) induced from somatic cells of patients with sensorineural hearing loss (caused by hereditary factors) is a promising method for its treatment. The correction of gene mutations in iPSCs could restore the normal function of cells and provide a rich source of cells for transplantation. | Author information:
(1)Department of Otolaryngology, University of Miami Miller School of Medicine,
Miami, FL 33136, USA.
(2)Department of Biology, University of Miami, Miami, FL 33146, USA.
(3)Department of Otology and Laryngology, Harvard Medical School and
Eaton-Peabody Laboratory, Massachusetts Eye and Ear Infirmary, Boston 02114,
USA; Department of Otology and Skull Base Surgery, Eye, Ear, Nose and Throat
Hospital, Shanghai Medical College, Fudan University, Shanghai, China.
(4)Department of Otology and Laryngology, Harvard Medical School and
Eaton-Peabody Laboratory, Massachusetts Eye and Ear Infirmary, Boston 02114,
USA; Department of Otolaryngology, Union Hospital, Tongji Medical College,
Huazhong University of Science and Technology, Wuhan, China.
(5)Department of Otolaryngology, Xiangya Hospital, Central South University,
Changsha, Hu, China.
(6)Department of Otolaryngology-Head and Neck Surgery, The Second Xiangya
Hospital Central South University, Changsha, Hu, China.
(7)Department of Otolaryngology, Union Hospital, Tongji Medical College,
Huazhong University of Science and Technology, Wuhan, China.
(8)Department of Otolaryngology-Head and Neck Surgery, Chinese PLA General
Hospital, Beijing, China.
(9)Department of Otology and Laryngology, Harvard Medical School and
Eaton-Peabody Laboratory, Massachusetts Eye and Ear Infirmary, Boston 02114,
USA. Electronic address: [email protected].
(10)Department of Otolaryngology, University of Miami Miller School of Medicine,
Miami, FL 33136, USA; Department of Otolaryngology, Xiangya Hospital, Central
South University, Changsha, Hu, China; Department of Otolaryngology-Head and
Neck Surgery, The Second Xiangya Hospital Central South University, Changsha,
Hu, China; Department of Otolaryngology-Head and Neck Surgery, Chinese PLA
General Hospital, Beijing, China. Electronic address: [email protected]. BACKGROUND: Nuclease-based technologies have been developed that enable
targeting of specific DNA sequences directly in the zygote. These approaches
provide an opportunity to modify the genomes of inbred mice, and allow the
removal of strain-specific mutations that confound phenotypic assessment. One
such mutation is the Cdh23 (ahl) allele, present in several commonly used inbred
mouse strains, which predisposes to age-related progressive hearing loss.
RESULTS: We have used targeted CRISPR/Cas9-mediated homology directed repair
(HDR) to correct the Cdh23 (ahl) allele directly in C57BL/6NTac zygotes.
Employing offset-nicking Cas9 (D10A) nickase with paired RNA guides and a
single-stranded oligonucleotide donor template we show that allele repair was
successfully achieved. To investigate potential Cas9-mediated 'off-target'
mutations in our corrected mouse, we undertook whole-genome sequencing and
assessed the 'off-target' sites predicted for the guide RNAs (≤4 nucleotide
mis-matches). No induced sequence changes were identified at any of these sites.
Correction of the progressive hearing loss phenotype was demonstrated using
auditory-evoked brainstem response testing of mice at 24 and 36 weeks of age,
and rescue of the progressive loss of sensory hair cell stereocilia bundles was
confirmed using scanning electron microscopy of dissected cochleae from
36-week-old mice.
CONCLUSIONS: CRISPR/Cas9-mediated HDR has been successfully utilised to
efficiently correct the Cdh23 (ahl) allele in C57BL/6NTac mice, and rescue the
associated auditory phenotype. The corrected mice described in this report will
allow age-related auditory phenotyping studies to be undertaken using
C57BL/6NTac-derived models, such as those generated by the International Mouse
Phenotyping Consortium (IMPC) programme. Deafness or hearing loss is a major issue in human health. Inner ear hair cells
are the main sensory receptors responsible for hearing. Defects in hair cells
are one of the major causes of deafness. A combination of induced pluripotent
stem cell (iPSC) technology with genome-editing technology may provide an
attractive cell-based strategy to regenerate hair cells and treat hereditary
deafness in humans. Here, we report the generation of iPSCs from members of a
Chinese family carrying MYO15A c.4642G>A and c.8374G>A mutations and the
induction of hair cell-like cells from those iPSCs. The compound heterozygous
MYO15A mutations resulted in abnormal morphology and dysfunction of the derived
hair cell-like cells. We used a CRISPR/Cas9 approach to genetically correct the
MYO15A mutation in the iPSCs and rescued the morphology and function of the
derived hair cell-like cells. Our data demonstrate the feasibility of generating
inner ear hair cells from human iPSCs and the functional rescue of gene
mutation-based deafness by using genetic correction. The genetic correction of induced pluripotent stem cells (iPSCs) induced from
somatic cells of patients with sensorineural hearing loss (caused by hereditary
factors) is a promising method for its treatment. The correction of gene
mutations in iPSCs could restore the normal function of cells and provide a rich
source of cells for transplantation. In the present study, iPSCs were generated
from a deaf patient with compound heterozygous MYO7A mutations (c.1184G>A and
c.4118C>T; P-iPSCs), the asymptomatic father of the patient (MYO7A c.1184G>A
mutation; CF-iPSCs), and a normal donor (MYO7A(WT/WT); C-iPSCs). One of MYO7A
mutation sites (c.4118C>T) in the P-iPSCs was corrected using CRISPR/Cas9. The
corrected iPSCs (CP-iPSCs) retained cell pluripotency and normal karyotypes.
Hair cell-like cells induced from CP-iPSCs showed restored organization of
stereocilia-like protrusions; moreover, the electrophysiological function of
these cells was similar to that of cells induced from C-iPSCs and CF-iPSCs.
These results might facilitate the development of iPSC-based gene therapy for
genetic disorders.
SIGNIFICANCE: Induced pluripotent stem cells (iPSCs) were generated from a deaf
patient with compound heterozygous MYO7A mutations (c.1184G>A and c.4118C>T).
One of the MYO7A mutation sites (c.4118C>T) in the iPSCs was corrected using
CRISPR/Cas9. The genetic correction of MYO7A mutation resulted in morphologic
and functional recovery of hair cell-like cells derived from iPSCs. These
findings confirm the hypothesis that MYO7A plays an important role in the
assembly of stereocilia into stereociliary bundles. Thus, the present study
might provide further insight into the pathogenesis of sensorineural hearing
loss and facilitate the development of therapeutic strategies against monogenic
disease through the genetic repair of patient-specific iPSCs. |
How may CTCF mediate splicing? | Two different mechanisms convey DNA methylation information into the regulation of alternative splicing. The first involves modulation of the elongation rate of RNA polymerase II (Pol II) by CCCTC-binding factor (CTCF) and methyl-CpG binding protein 2 (MeCP2); the second involves the formation of a protein bridge by heterochromatin protein 1 (HP1) that recruits splicing factors onto transcribed alternative exons. | Alternative splicing of pre-messenger RNA is a key feature of transcriptome
expansion in eukaryotic cells, yet its regulation is poorly understood.
Spliceosome assembly occurs co-transcriptionally, raising the possibility that
DNA structure may directly influence alternative splicing. Supporting such an
association, recent reports have identified distinct histone methylation
patterns, elevated nucleosome occupancy and enriched DNA methylation at exons
relative to introns. Moreover, the rate of transcription elongation has been
linked to alternative splicing. Here we provide the first evidence that a
DNA-binding protein, CCCTC-binding factor (CTCF), can promote inclusion of weak
upstream exons by mediating local RNA polymerase II pausing both in a mammalian
model system for alternative splicing, CD45, and genome-wide. We further show
that CTCF binding to CD45 exon 5 is inhibited by DNA methylation, leading to
reciprocal effects on exon 5 inclusion. These findings provide a mechanistic
basis for developmental regulation of splicing outcome through heritable
epigenetic marks. Acute myeloid leukemia (AML) is characterized by molecular heterogeneity. As
commonly altered genomic regions point to candidate genes involved in
leukemogenesis, we used microarray-based comparative genomic hybridization and
single nucleotide polymorphism profiling data of 391 AML cases to further narrow
down genomic regions of interest. Targeted resequencing of 1000 genes located in
the critical regions was performed in a representative cohort of 50 AML samples
comprising all major cytogenetic subgroups. We identified 120 missense/nonsense
mutations as well as 60 insertions/deletions affecting 73 different genes (∼ 3.6
tumor-specific aberrations/AML). While most of the newly identified alterations
were nonrecurrent, we observed an enrichment of mutations affecting genes
involved in epigenetic regulation including known candidates like TET2, TET1,
DNMT3A, and DNMT1, as well as mutations in the histone methyltransferases NSD1,
EZH2, and MLL3. Furthermore, we found mutations in the splicing factor SFPQ and
in the nonclassic regulators of mRNA processing CTCF and RAD21. These
splicing-related mutations affected 10% of AML patients in a mutually exclusive
manner. In conclusion, we could identify a large number of alterations in genes
involved in aberrant splicing and epigenetic regulation in genomic regions
commonly altered in AML, highlighting their important role in the molecular
pathogenesis of AML. The Wilms tumor 1 (WT1) gene plays an essential role in early development and
differentiation of the urinary tract, particularly the kidneys. Aberrant
transcriptional activity of WT1 is a key finding in the genesis of Wilms tumors
(WTs). However, the mechanisms responsible for this alteration remain poorly
understood. In the present study, we examined the methylation pattern of a
putative CCCTC-binding factor (CTCF) binding site downstream of the WT1 gene as
a potential cause of WT1 misregulation in 44 native WT specimens. We found that
16 WT cases exhibited a much higher WT1 expression compared to normal kidney
tissue, and that the high mRNA expression of WT1 is strongly correlated with a
high degree of DNA methylation of the CTCF binding site near the WT1 promoter.
However, there was no correlation between the KTS+/KTS- splicing variants of WT1
and the methylation status of the CpGs of the CTCF binding site. Our results
demonstrated an aberrant methylation pattern at a CTCF binding site downstream
the WT1 gene, which is associated with an elevated WT1 transcriptional activity.
Thus, methylation of the CTCF binding site may be partially responsible for the
transcriptional activation of the WT1 locus and hypermethylation of this site
may be an important oncogenic mechanism in the genesis of WT. CTCF plays a vital role in chromatin structure and function. CTCF is
ubiquitously expressed and plays diverse roles in gene regulation, imprinting,
insulation, intra/interchromosomal interactions, nuclear compartmentalisation,
and alternative splicing. CTCF has a single paralogue, the testes-specific
CTCF-like gene (CTCFL)/BORIS. CTCF and BORIS can be deregulated in cancer. The
tumour suppressor gene CTCF can be mutated or deleted in cancer, or CTCF DNA
binding can be altered by epigenetic changes. BORIS is aberrantly expressed
frequently in cancer, leading some to propose a pro-tumourigenic role for BORIS.
However, BORIS can inhibit cell proliferation, and is mutated in cancer
similarly to CTCF suggesting BORIS activation in cancer may be due to global
genetic or epigenetic changes typical of maligt transformation. Although DNA methylation was originally thought to only affect transcription,
emerging evidence shows that it also regulates alternative splicing. Exons, and
especially splice sites, have higher levels of DNA methylation than flanking
introns, and the splicing of about 22% of alternative exons is regulated by DNA
methylation. Two different mechanisms convey DNA methylation information into
the regulation of alternative splicing. The first involves modulation of the
elongation rate of RNA polymerase II (Pol II) by CCCTC-binding factor (CTCF) and
methyl-CpG binding protein 2 (MeCP2); the second involves the formation of a
protein bridge by heterochromatin protein 1 (HP1) that recruits splicing factors
onto transcribed alternative exons. These two mechanisms, however, regulate only
a fraction of such events, implying that more underlying mechanisms remain to be
found. All viruses target host cell factors for successful life cycle completion.
Transcriptional control of DNA viruses by host cell factors is important in the
temporal and spatial regulation of virus gene expression. Many of these factors
are recruited to enhance virus gene expression and thereby increase virus
production, but host cell factors can also restrict virus gene expression and
productivity of infection. CCCTC binding factor (CTCF) is a host cell DNA
binding protein important for the regulation of genomic chromatin boundaries,
transcriptional control and enhancer element usage. CTCF also functions in RNA
polymerase II regulation and in doing so can influence co-transcriptional
splicing events. Several DNA viruses, including Kaposi's sarcoma-associated
herpesvirus (KSHV), Epstein-Barr virus (EBV) and human papillomavirus (HPV)
utilize CTCF to control virus gene expression and many studies have highlighted
a role for CTCF in the persistence of these diverse oncogenic viruses. CTCF can
both enhance and repress virus gene expression and in some cases CTCF increases
the complexity of alternatively spliced transcripts. This review article will
discuss the function of CTCF in the life cycle of DNA viruses in the context of
known host cell CTCF functions. Author information:
(1)Section of Virology, Division of Infectious Diseases, Imperial College,
London W2 1PG, United Kingdom; Centre for AIDS Research, Kumamoto University,
Kumamoto 860-0811, Japan; Priority Organization for Innovation and Excellence,
Kumamoto University, Kumamoto 860-0811, Japan; International Research Centre for
Medical Science, Kumamoto University, Kumamoto 860-0811, Japan; Core Research
for Evolutionary Science and Technology, Japan Science and Technology Agency,
Saitama 332-0012, Japan; [email protected] [email protected].
(2)Centre for AIDS Research, Kumamoto University, Kumamoto 860-0811, Japan;
Priority Organization for Innovation and Excellence, Kumamoto University,
Kumamoto 860-0811, Japan; International Research Centre for Medical Science,
Kumamoto University, Kumamoto 860-0811, Japan;
(3)Priority Organization for Innovation and Excellence, Kumamoto University,
Kumamoto 860-0811, Japan; Core Research for Evolutionary Science and Technology,
Japan Science and Technology Agency, Saitama 332-0012, Japan;
(4)Section of Virology, Division of Infectious Diseases, Imperial College,
London W2 1PG, United Kingdom;
(5)Cancer Centre, Kumamoto University Hospital, Kumamoto University, Kumamoto
860-0811, Japan;
(6)Department of Medical Cell Biology, Institute of Molecular Embryology and
Genetics, Kumamoto University, Kumamoto 860-0811, Japan.
(7)Core Research for Evolutionary Science and Technology, Japan Science and
Technology Agency, Saitama 332-0012, Japan; Department of Medical Cell Biology,
Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto
860-0811, Japan.
(8)Section of Virology, Division of Infectious Diseases, Imperial College,
London W2 1PG, United Kingdom; [email protected]
[email protected]. The role of the zinc finger protein CTCF in organizing the genome within the
nucleus is now well established. Widely separated sites on DNA, occupied by both
CTCF and the cohesin complex, make physical contacts that create large loop
domains. Additional contacts between loci within those domains, often also
mediated by CTCF, tend to be favored over contacts between loci in different
domains. A large number of studies during the past 2 years have addressed the
questions: How are these loops generated? What are the effects of disrupting
them? Are there rules governing large-scale genome organization? It now appears
that the strongest and evolutionarily most conserved of these CTCF interactions
have specific rules for the orientation of the paired CTCF sites, implying the
existence of a nonequilibrium mechanism of generation. Recent experiments that
invert, delete, or inactivate one of a mating CTCF pair result in major changes
in patterns of organization and gene expression in the surrounding regions. What
remain to be determined are the detailed molecular mechanisms for
re-establishing loop domains and maintaining them after replication and mitosis.
As recently published data show, some mechanisms may involve interactions with
noncoding RNAs as well as protein cofactors. Many CTCF sites are also involved
in other functions such as modulation of RNA splicing and specific regulation of
gene expression, and the relationship between these activities and loop
formation is another uswered question that should keep investigators occupied
for some time. |
Is ACI-35 a passive vaccine? | No, ACI-35 is an active vaccine. | Author information:
(1)Neurodegenerative Disease Unit, Department of Basic Medicine, Neuroscience, &
Sense Organs, University of Bari Aldo Moro, Bari, Italy.
(2)Department of Clinical Research in Neurology, University of Bari Aldo Moro,
'Pia Fondazione Cardinale G. Panico,' Tricase, Lecce, Italy.
(3)Geriatric Unit & Laboratory of Gerontology & Geriatrics, Department of
Medical Sciences, IRCCS 'Casa Sollievo della Sofferenza,' San Giovanni Rotondo,
Foggia, Italy.
(4)Geriatric Medicine-Memory Unit & Rare Disease Centre, University of Bari Aldo
Moro, Bari, Italy.
(5)Research & Development Department, Chiesi Farmaceutici, Parma, Italy.
(6)Physical Medicine & Rehabilitation Section, 'OORR' Hospital, University of
Foggia, Foggia, Italy.
(7)Psychiatric Unit, Department of Basic Medicine, Neuroscience, & Sense Organs,
University of Bari Aldo Moro, Bari, Italy.
(8)Department of OrthoGeriatrics, Rehabilitation & Stabilization, Frailty Area,
E.O. Galliera NR-HS Hospital, Genova, Italy.
(9)Institute of Neurology, Catholic University of Sacred Heart, Rome, Italy. |
Which main ribotype of Clostridium difficile is responsible of the recent outbreak? | The outbreak of the hypervirulent strain belonging to ribotype 027 has increased the incidence and severity of CDI in some countries. | Author information:
(1)Department of clinical laboratory, China-Japan Friendship Hospital, No. 2
Yinghua Dongjie, Chaoyang, Beijing, 100029, China.
(2)Institute of Infectious Diseases, Beijing Ditan Hospital, Capital Medical
University, and Beijing Key Laboratory of Emerging infectious Diseases, No. 8
Jingshundongjie, Beijing, 100015, China.
(3)State Key Laboratory for Infectious Disease Prevention and Control, and
National Institute for Communicable Disease Control and Prevention, Chinese
Center for Disease Control and Prevention, Beijing, 102206, China.
(4)Collaborative Innovation Center for Diagnosis and Treatment of Infectious
Diseases, Hangzhou, 310003, China.
(5)National Reference Laboratory for Clostridium difficile, Faculté de Médecine
Pierre et Marie Curie and Hôpital Saint-Antoine, Assistance Publique-Hôpitaux de
Paris, Paris, 75012, France.
(6)The Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101,
China.
(7)Department of clinical laboratory, China-Japan Friendship Hospital, No. 2
Yinghua Dongjie, Chaoyang, Beijing, 100029, China. [email protected].
(8)Key Laboratory of Surveillance and Early-warning on Infectious Disease,
Division of Infectious Disease, Chinese Center for Disease Control and
Prevention, 155 Changbai Road, Changping, Beijing, 102206, China.
[email protected].
(9)Institute of Infectious Diseases, Beijing Ditan Hospital, Capital Medical
University, and Beijing Key Laboratory of Emerging infectious Diseases, No. 8
Jingshundongjie, Beijing, 100015, China. [email protected].
(10)State Key Laboratory for Infectious Disease Prevention and Control, and
National Institute for Communicable Disease Control and Prevention, Chinese
Center for Disease Control and Prevention, Beijing, 102206, China.
[email protected]. BACKGROUND: An outbreak of Clostridium difficile ribotype 027 infection (CDI)
occurred at an university hospital, involving 19 departments. To determine what
hospital-associated factors drove the outbreak of this particular strain we
performed a case-control study.
METHODS: Cases (n = 79), diagnosed with CDI due to C. difficile ribotype 027
were matched for age and treating medical specialty to four control patients (n
= 316). Patients diagnosed with CDI due to other ribotypes were included as a
second control group. A random selection of C. difficile ribotype 027 strains (n
= 10) was genotyped by Whole Genome Sequencing (WGS).
FINDINGS: WGS showed the outbreak was likely caused by a single strain of C.
difficile (two or less single-nucleotide variants between isolates). Ninety-five
percent of cases had used antibiotics, compared to 56% of controls. Previous
admission to the intensive care unit (ICU) (OR: 2.4, 95% CI 1.0-5.6), longer
length of stay (LOS), and recent hospital admission were associated with CDI
ribotype 027. Cases were less likely to have been admitted to a ward with a
known isolated CDI patient (OR: 0.2, 95% CI 0.1-0.6). Analysis of patients who
stayed at the ICU (35 cases; 51 controls), indicated that the use of selective
decontamination of the digestive tract (SDD) and a longer LOS in the ICU were
associated with CDI risk.
INTERPRETATION: In this large outbreak, any antibiotic use, including SDD use,
appeared as a prerequisite for acquisition of the outbreak strain. The role of
use of SDD and prolonged stay on the ICU could not be disentangled, but both
factors can play a biologically plausible role in C. difficile acquisition and
infection. Author information:
(1)Servicio de Microbiología y Enfermedades Infecciosas, Hospital General
Universitario Gregorio Marañón, Madrid, España; Centro de Investigación
Biomédica en Red de Enfermedades Respiratorias (CIBERES CD06/06/0058), Madrid,
España; Instituto de Investigación Sanitaria Gregorio Marañón, Madrid, España.
Electronic address: [email protected].
(2)Servicio de Microbiología y Enfermedades Infecciosas, Hospital General
Universitario Gregorio Marañón, Madrid, España; Instituto de Investigación
Sanitaria Gregorio Marañón, Madrid, España.
(3)Servicio de Microbiología y Enfermedades Infecciosas, Hospital General
Universitario Gregorio Marañón, Madrid, España; Centro de Investigación
Biomédica en Red de Enfermedades Respiratorias (CIBERES CD06/06/0058), Madrid,
España; Instituto de Investigación Sanitaria Gregorio Marañón, Madrid, España;
Facultad de Medicina, Universidad Complutense de Madrid (UCM), Madrid, España. An outbreak of Clostridium difficile infection (CDI) caused by ribotype 027
(B1/NAP1) began in our hospital in November 2014, and produced 141 episodes in
the following months. The aim of this study is to describe this outbreak, assess
risk factors for recurrence of CDI-027 and to analyze the implementation of a
novel treatment strategy. This is a prospective study of all patients with
CDI-027, from November 2014 to November 2015. The epidemiological data were
collected daily for each patient. We compared clinical characteristics and
treatment between patients with and without recurrence of CDI-027.
Interestingly, liver cirrhosis was present in 22% of the patients, and most of
them received prophylaxis for hepatic encephalopathy with rifaximin. Patients
were also taking antimicrobial drugs (93.6%) and proton pump inhibitors (80.1%).
Overall, 27 (23.5%) patients had a first recurrence of CDI-027. Liver cirrhosis
increased the risk of recurrence (44.4% vs 14.8%). Patients treated with a
prolonged oral vancomycin regimen vs the conventional regimen (oral
metronidazole or 10 days of vancomycin) had fewer recurrences (8.6 versus 44.7%
[p ≤ 0.01]; OR, 0.91; 95% CI, 0.028-0.294) and less attributable mortality (0%
versus 7.1%; p = 0.058). We report an outbreak of CDI-027, mainly in patients
with liver cirrhosis. Recurrence of CDI-027 was more common in those patients. A
novel approach involving high-dose prolonged vancomycin taper as a first-line
treatment, together with a bundle of outbreak measures, seemed to reduce the
number of cases of CDI-027, recurrences, and attributable mortality.
Nevertheless, this approach warrants further investigation. OBJECTIVE Estimating the risk of a complicated course of Clostridium difficile
infection (CDI) might help doctors guide treatment. We aimed to validate 3
published prediction models: Hensgens (2014), Na (2015), and Welfare (2011).
METHODS The validation cohort comprised 148 patients diagnosed with CDI between
May 2013 and March 2014. During this period, 70 endemic cases of CDI occurred as
well as 78 cases of CDI related to an outbreak of C. difficile ribotype 027.
Model calibration and discrimination were assessed for the 3 prediction rules.
RESULTS A complicated course (ie, death, colectomy, or ICU admission due to CDI)
was observed in 31 patients (21%), and 23 patients (16%) died within 30 days of
CDI diagnosis. The performance of all 3 prediction models was poor when applied
to the total validation cohort with an estimated area under the curve (AUC) of
0.68 for the Hensgens model, 0.54 for the Na model, and 0.61 for the Welfare
model. For those patients diagnosed with CDI due to non-outbreak strains, the
prediction model developed by Hensgens performed the best, with an AUC of 0.78.
CONCLUSION All 3 prediction models performed poorly when using our total cohort,
which included CDI cases from an outbreak as well as endemic cases. The
prediction model of Hensgens performed relatively well for patients diagnosed
with CDI due to non-outbreak strains, and this model may be useful in endemic
settings. Infect Control Hosp Epidemiol 2017;38:897-905. OBJECTIVE: A frequent complication of Clostridium difficile infection (CDI) is
recurrent disease. The aim of this study was to determine whether early
recurrence risk was higher after infection with ribotype 027 (outbreak strain)
compared with infection with endemic strain types of C. difficile.
METHODS: Consecutive patients diagnosed with CDI between May 2013 and March 2014
were included (outbreak strain, and non-outbreak strains). Patients who
developed recurrent CDI within 30 days after completion of CDI treatment, were
compared with patients without a recurrence. Medical charts were reviewed for
demographic and clinical characteristics. General practitioners were contacted
to complete data about the occurrence of recurrent CDI, and the use of
medication after hospital discharge.
RESULTS: In total, 135 patients were at risk for the development of recurrent
CDI; 74 patients were infected by ribotype 027, and 61 patients by other
ribotypes. Thirty-nine patients (29%) developed recurrent CDI within 30 days
after completion of CDI treatment. In multivariable analysis, age ≥70 years (HR
3.05, 95% CI 1.54-6.03), and a duration of CDI treatment ≥11 days (HR 1.92, 95%
CI 1.00-3.69) were clearly associated with recurrence; infection with ribotype
027 showed a HR of 1.72 (95% CI 0.88-3.33).
CONCLUSION: During this outbreak of C. difficile in a tertiary care centre, age
and a prolonged duration of CDI therapy (which is most likely a marker of
underlying disease severity) were the main risk factors for recurrent CDI. This
points to host factors as more important predictors for recurrent CDI than
strain type or antibiotic use. |
What is the function of the TFIIS transcriptional factor (Dst1) in yeast? | TFIIS, an elongation factor encoded by DST1 in Saccharomyces cerevisiae, stimulates transcript cleavage in arrested RNA polymerase II. | TFIIS promotes the intrinsic ability of RNA polymerase II to cleave the 3'-end
of the newly synthesized RNA. This stimulatory activity of TFIIS, which is
dependent upon Rpb9, facilitates the resumption of transcription elongation when
the polymerase stalls or arrests. While TFIIS has a pronounced effect on
transcription elongation in vitro, the deletion of DST1 has no major effect on
cell viability. In this work we used a genetic approach to increase our
knowledge of the role of TFIIS in vivo. We showed that: (1) dst1 and rpb9
mutants have a synthetic growth defective phenotype when combined with fyv4,
gim5, htz1, yal011w, ybr231c, soh1, vps71, and vps72 mutants that is exacerbated
during germination or at high salt concentrations; (2) TFIIS and Rpb9 are
essential when the cells are challenged with microtubule-destabilizing drugs;
(3) among the SDO (synthetic with Dst one), SOH1 shows the strongest genetic
interaction with DST1; (4) the presence of multiple copies of TAF14, SUA7,
GAL11, RTS1, and TYS1 alleviate the growth phenotype of dst1 soh1 mutants; and
(5) SRB5 and SIN4 genetically interact with DST1. We propose that TFIIS is
required under stress conditions and that TFIIS is important for the transition
between initiation and elongation in vivo. TFIIS, an elongation factor encoded by DST1 in Saccharomyces cerevisiae,
stimulates transcript cleavage in arrested RNA polymerase II. Two components of
the RNA polymerase II machinery, Med13 (Srb9) and Spt8, were isolated as
two-hybrid partners of the conserved TFIIS N-terminal domain. They belong to the
Cdk8 module of the Mediator and to a subform of the SAGA co-activator,
respectively. Co-immunoprecipitation experiments showed that TFIIS can bind the
Cdk8 module and SAGA in cell-free extracts. spt8Delta and dst1Delta mutants were
sensitive to nucleotide-depleting drugs and epistatic to null mutants of the RNA
polymerase II subunit Rpb9, suggesting that their elongation defects are
mediated by Rpb9. rpb9Delta, spt8Delta and dst1Delta were lethal in cells
lacking the Rpb4 subunit. The TFIIS N-terminal domain is also strictly required
for viability in rpb4Delta, although it is not needed for binding to RNA
polymerase II or for transcript cleavage. It is proposed that TFIIS and the
Spt8-containing form of SAGA co-operate to rescue RNA polymerase II from
unproductive elongation complexes, and that the Cdk8 module temporarily blocks
transcription during transcript cleavage. The eukaryotic transcript elongation factor TFIIS is encoded by a nonessential
gene, PPR2, in Saccharomyces cerevisiae. Disruptions of PPR2 are lethal in
conjunction with a disruption in the nonessential gene TAF14/TFG3. While
investigating which of the Taf14p-containing complexes may be responsible for
the synthetic lethality between ppr2Delta and taf14Delta, we discovered genetic
interactions between PPR2 and both TFG1 and TFG2 encoding the two larger
subunits of the TFIIF complex that also contains Taf14p. Mutant alleles of tfg1
or tfg2 that render cells cold sensitive have improved growth at low temperature
in the absence of TFIIS. Remarkably, the amino-terminal 130 amino acids of
TFIIS, which are dispensable for the known in vitro and in vivo activities of
TFIIS, are required to complement the lethality in taf14Delta ppr2Delta cells.
Analyses of deletion and chimeric gene constructs of PPR2 implicate
contributions by different regions of this N-terminal domain. No strong common
phenotypes were identified for the ppr2Delta and taf14Delta strains, implying
that the proteins are not functionally redundant. Instead, the absence of Taf14p
in the cell appears to create a dependence on an undefined function of TFIIS
mediated by its N-terminal region. This region of TFIIS is also at least in part
responsible for the deleterious effect of TFIIS on tfg1 or tfg2 cold-sensitive
cells. Together, these results suggest a physiologically relevant functional
connection between TFIIS and TFIIF. TFIIS is a transcription elongation factor that stimulates transcript cleavage
activity of arrested RNA polymerase II (Pol II). Recent studies revealed that
TFIIS has also a role in Pol II transcription initiation. To improve our
understanding of TFIIS function in vivo, we performed genome-wide location
analysis of this factor. Under normal growth conditions, TFIIS was detected on
Pol II-transcribed genes, and TFIIS occupancy was well correlated with that of
Pol II, indicating that TFIIS recruitment is not restricted to NTP-depleted
cells. Unexpectedly, TFIIS was also detected on almost all Pol III-transcribed
genes. TFIIS and Pol III occupancies correlated well genome-wide on this novel
class of targets. In vivo, some dst1 mutants were partly defective in tRNA
synthesis and showed a reduced Pol III occupancy at the restrictive temperature.
In vitro transcription assays suggested that TFIIS may affect Pol III start site
selection. These data provide strong in vivo and in vitro evidence in favor of a
role of TFIIS as a general Pol III transcription factor. The catalytic center of yeast RNA polymerase II and III contains an acidic loop
borne by their second largest subunit (Rpb2-(832)GYNQED(837),
Rpc128-(764)GYDIED(769)) and highly conserved in all cellular and viral
DNA-dependent RNA polymerases. A site-directed mutagenesis of this dicarboxylic
motif reveals its strictly essential character in RNA polymerase III, with a
slightly less stringent pattern in RNA polymerase II, where rpb2-E836Q and other
substitutions completely prevent growth, whereas rpb2-E836A combines a domit
growth defect with severe lethal sectoring. A mild but systematic increase in
RNA polymerase occupancy and a strict dependency on the transcript cleavage
factor TFIIS (Dst1) also suggest a slower rate of translocation or higher
probability of transcriptional stalling in this mutation. A conserved nucleotide
triphosphate funnel domain binds the Rpb2-(832)GYNQED(837) loop by an
Rpb2-R(1020)/Rpb2-D(837) salt-bridge. Molecular dynamic simulations reveal a
second bridge (Rpb1-K(752)/Rpb2-E(836)), which may account for the critical role
of the invariant Rpb2-E(836). Rpb2-E(836) and the funnel domain are not found
among the RNA-dependent eukaryotic RNA polymerases and may thus represent a
specific adaptation to double-stranded DNA templates. Transcription factor IIS (TFIIS) stimulates RNA cleavage by RNA polymerase II by
allowing backtracked enzymes to resume transcription elongation. Yeast cells do
not require TFIIS for viability, unless they suffer severe transcriptional
stress due to NTP-depleting drugs like 6-azauracil or mycophenolic acid. In
order to broaden our knowledge on the role of TFIIS under transcriptional
stress, we carried out a genetic screening for suppressors of TFIIS-lacking
cells' sensitivity to 6-azauracil and mycophenolic acid. Five suppressors were
identified, four of which were related to the transcriptional regulation of
those genes encoding ribosomal components [rRNAs and ribosomal proteins (RP)],
including global regulator SFP1. This led us to discover that RNA polymerase II
is hypersensitive to the absence of TFIIS under NTP scarcity conditions when
transcribing RP genes. The absence of Sfp1 led to a profound alteration of the
transcriptional response to NTP-depletion, thus allowing the expression of RP
genes to resist these stressful conditions in the absence of TFIIS. We discuss
the effect of transcriptional stress on ribosome biogenesis and propose that
TFIIS contributes to prevent a transcriptional imbalance between rDNA and RP
genes. |
Is DNA methylation correlated with nucleosome occupancy? | DNA methylation can determine nucleosome positioning. Using a novel bioinformatics pipeline, we show a striking anti-correlation between nucleosome occupancy and DNA methylation at CTCF regions that is not present at promoters. DNA methylation determines nucleosome occupancy in the 5'-CpG islands of tumor suppressor genes. In contrast, exons demonstrate a high degree of methylation and nucleosome occupancy. Although global DNA demethylation has been observed after treatment, it is unclear to what extent demethylation induces changes in nucleosome occupancy, a key determinant of gene expression. In order to systematically evaluate potential diversities among CGIs and ultimately to illuminate the link between diversity of CGIs and their epigenetic variation, we analyzed the nucleotide-resolution DNA methylation maps of multiple cellular origins. Using this global approach, we observe the dependency of nucleosome occupancy upon the DNA methylation status. Exonic DNA methylation seems to function together with exonic nucleosomes and H3K36me3 for the proper splicing of transcripts with different expression levels. Transcription, histone modifications, and DNA methylation alter this "ground state" by having distinct effects on both nucleosome positioning and occupancy. | BACKGROUND: CpG islands and nucleosome-free regions are both found in promoters.
However, their association has never been studied. On the other hand, DNA
methylation is absent in promoters but is enriched in gene bodies. Intragenic
nucleosomes and their modifications have been recently associated with RNA
splicing. Because the function of intragenic DNA methylation remains unclear, I
explored the possibility of its involvement in splicing regulation.
RESULTS: Here I show that CpG islands were associated not only with
methylation-free promoters but also with nucleosome-free promoters.
Nucleosome-free regions were observed only in promoters containing a CpG island.
However, the DNA sequences of CpG islands predicted the opposite pattern,
implying a limitation of sequence programs for the determination of nucleosome
occupancy. In contrast to the methylation-and nucleosome-free states of
CpG-island promoters, exons were densely methylated at CpGs and packaged into
nucleosomes. Exon-enrichment of DNA methylation was specifically found in
spliced exons and in exons with weak splice sites. The enrichment patterns were
less pronounced in initial exons and in non-coding exons, potentially reflecting
a lower need for their splicing. I also found that nucleosomes, DNA methylation,
and H3K36me3 marked the exons of transcripts with low, medium, and high gene
expression levels, respectively.
CONCLUSIONS: Human promoters containing a CpG island tend to remain
nucleosome-free as well as methylation-free. In contrast, exons demonstrate a
high degree of methylation and nucleosome occupancy. Exonic DNA methylation
seems to function together with exonic nucleosomes and H3K36me3 for the proper
splicing of transcripts with different expression levels. Despite the fact that 45% of all human gene promoters do not contain CpG
islands, the role of DNA methylation in control of non-CpG island promoters is
controversial and its relevance in normal and pathological processes is poorly
understood. Among the few studies which investigate the correlation between DNA
methylation and expression of genes with non-CpG island promoters, the majority
do not support the view that DNA methylation directly leads to transcription
silencing of these genes. Our reporter assays and gene reactivation by
5-aza-2'-deoxycytidine, a DNA demethylating agent, show that DNA methylation
occurring at CpG poor LAMB3 promoter and RUNX3 promoter 1(RUNX3 P1) can directly
lead to transcriptional silencing in cells competent to express these genes in
vitro. Using Nucleosome Occupancy Methylome- Sequencing, NOMe-Seq, a
single-molecule, high-resolution nucleosome positioning assay, we demonstrate
that active, but not inactive, non-CpG island promoters display a
nucleosome-depleted region (NDR) immediately upstream of the transcription start
site (TSS). Furthermore, using NOMe-Seq and clonal analysis, we show that in
RUNX3 expressing 623 melanoma cells, RUNX3 P1 has two distinct chromatin
configurations: one is unmethylated with an NDR upstream of the TSS; another is
methylated and nucleosome occupied, indicating that RUNX3 P1 is monoallelically
methylated. Together, these results demonstrate that the epigenetic signatures
comprising DNA methylation, histone marks and nucleosome occupancy of non-CpG
island promoters are almost identical to CpG island promoters, suggesting that
aberrant methylation patterns of non-CpG island promoters may also contribute to
tumorigenesis and should therefore be included in analyses of cancer
epigenetics. Alternative splicing of pre-messenger RNA is a key feature of transcriptome
expansion in eukaryotic cells, yet its regulation is poorly understood.
Spliceosome assembly occurs co-transcriptionally, raising the possibility that
DNA structure may directly influence alternative splicing. Supporting such an
association, recent reports have identified distinct histone methylation
patterns, elevated nucleosome occupancy and enriched DNA methylation at exons
relative to introns. Moreover, the rate of transcription elongation has been
linked to alternative splicing. Here we provide the first evidence that a
DNA-binding protein, CCCTC-binding factor (CTCF), can promote inclusion of weak
upstream exons by mediating local RNA polymerase II pausing both in a mammalian
model system for alternative splicing, CD45, and genome-wide. We further show
that CTCF binding to CD45 exon 5 is inhibited by DNA methylation, leading to
reciprocal effects on exon 5 inclusion. These findings provide a mechanistic
basis for developmental regulation of splicing outcome through heritable
epigenetic marks. DNA methylation and nucleosome positioning work together to generate chromatin
structures that regulate gene expression. Nucleosomes are typically mapped using
nuclease digestion requiring significant amounts of material and varying enzyme
concentrations. We have developed a method (NOMe-seq) that uses a GpC
methyltransferase (M.CviPI) and next generation sequencing to generate a high
resolution footprint of nucleosome positioning genome-wide using less than 1
million cells while retaining endogenous DNA methylation information from the
same DNA strand. Using a novel bioinformatics pipeline, we show a striking
anti-correlation between nucleosome occupancy and DNA methylation at CTCF
regions that is not present at promoters. We further show that the extent of
nucleosome depletion at promoters is directly correlated to expression level and
can accommodate multiple nucleosomes and provide genome-wide evidence that
expressed non-CpG island promoters are nucleosome-depleted. Importantly,
NOMe-seq obtains DNA methylation and nucleosome positioning information from the
same DNA molecule, giving the first genome-wide DNA methylation and nucleosome
positioning correlation at the single molecule, and thus, single cell level,
that can be used to monitor disease progression and response to therapy. We determined genome-wide nucleosome occupancies in mouse embryonic stem cells
and their neural progenitor and embryonic fibroblast counterparts to assess
features associated with nucleosome positioning during lineage commitment.
Cell-type- and protein-specific binding preferences of transcription factors to
sites with either low (Myc, Klf4 and Zfx) or high (Nanog, Oct4 and Sox2)
nucleosome occupancy as well as complex patterns for CTCF were identified.
Nucleosome-depleted regions around transcription start and transcription
termination sites were broad and more pronounced for active genes, with distinct
patterns for promoters classified according to CpG content or histone
methylation marks. Throughout the genome, nucleosome occupancy was correlated
with certain histone methylation or acetylation modifications. In addition, the
average nucleosome repeat length increased during differentiation by 5-7 base
pairs, with local variations for specific regions. Our results reveal regulatory
mechanisms of cell differentiation that involve nucleosome repositioning. DNA methylation inhibitors such as 5-aza-2'-deoxycytidine (5-Aza-CdR) are
currently used for the treatment of myelodysplastic syndrome. Although global
DNA demethylation has been observed after treatment, it is unclear to what
extent demethylation induces changes in nucleosome occupancy, a key determit
of gene expression. We use the colorectal cancer cell line HCT116 as a model to
address this question and determine that <2% of regions demethylated by
5-Aza-CdR treatment assume an open configuration. Consolidating our findings, we
detect nucleosome retention at sites of global DNA methylation loss in DKO1, an
HCT116-derived non-tumorigenic cell-line engineered for DNA methyltransferase
disruption. Notably, regions that are open in both HCT116 cells after treatment
and in DKO1 cells include promoters belonging to tumor suppressors and genes
under-expressed in colorectal cancers. Our results indicate that only a minority
of demethylated promoters are associated with nucleosome remodeling, and these
could potentially be the epigenetic drivers causing the loss of tumorigenicity.
Furthermore, we show that the chromatin opening induced by the histone
deacetylase inhibitor suberoylanilide hydroxamic acid has strikingly distinct
targets compared with those of 5-Aza-CdR, providing a mechanistic explanation
for the importance of combinatorial therapy in eliciting maximal de-repression
of the cancer epigenome. DNA methylation is known to regulate transcription and was recently found to be
involved in exon recognition via cotranscriptional splicing. We recently
observed that exon-intron architectures can be grouped into two classes: one
with higher GC content in exons compared to the flanking introns, and the other
with similar GC content in exons and introns. The first group has higher
nucleosome occupancy on exons than introns, whereas the second group exhibits
weak nucleosome marking of exons, suggesting another type of epigenetic marker
distinguishes exons from introns when GC content is similar. We find different
and specific patterns of DNA methylation in each of the GC architectures; yet in
both groups, DNA methylation clearly marks the exons. Exons of the leveled GC
architecture exhibit a significantly stronger DNA methylation signal in relation
to their flanking introns compared to exons of the differential GC architecture.
This is accentuated by a reduction of the DNA methylation level in the intronic
sequences in proximity to the splice sites and shows that different epigenetic
modifications mark the location of exons already at the DNA level. Also, lower
levels of methylated CpGs on alternative exons can successfully distinguish
alternative exons from constitutive ones. Three positions at the splice sites
show high CpG abundance and accompany elevated nucleosome occupancy in a leveled
GC architecture. Overall, these results suggest that DNA methylation affects
exon recognition and is influenced by the GC architecture of the exon and
flanking introns. CpG islands (CGIs) are commonly used as genomic markers to study the patterns
and regulatory consequences of DNA methylation. Interestingly, recent studies
reveal a substantial diversity among CGIs: long and short CGIs, for example,
exhibit contrasting patterns of gene expression complexity and nucleosome
occupancy. Evolutionary origins of CGIs are also highly heterogeneous. In order
to systematically evaluate potential diversities among CGIs and ultimately to
illuminate the link between diversity of CGIs and their epigenetic variation, we
analyzed the nucleotide-resolution DNA methylation maps (methylomes) of multiple
cellular origins. We discover novel 'clusters' of CGIs according to their
patterns of DNA methylation; the stably hypomethylated CGI cluster (cluster I),
sperm-hypomethylated CGI cluster (cluster II), and variably methylated CGI
cluster (cluster III). These epigenomic CGI clusters are strikingly distinct at
multiple biological features including genomic, evolutionary, and functional
characteristics. At the genomic level, the stably hypomethylated CGI cluster
tends to be longer and harbors many more CpG dinucleotides than those in other
clusters. They are also frequently associated with promoters, while CGI clusters
II and III mostly reside in intragenic or intergenic regions and exhibit highly
tissue-specific DNA methylation. Functional ontology terms and transcriptional
profiles co-vary with CGI clusters, indicating that the regulatory functions of
CGIs are tightly linked to their heterogeneity. Finally, CGIs associated with
distinctive biological processes, such as diseases, aging, and imprinting, occur
disproportionately across CGI clusters. These new findings provide an effective
means to combine existing knowledge on CGIs into a genomic context while
bringing new insights that elucidate the significance of DNA methylation across
different biological conditions and demography. During differentiation of embryonic stem cells, chromatin reorganizes to
establish cell type-specific expression programs. Here, we have dissected the
linkages between DNA methylation (5mC), hydroxymethylation (5hmC), nucleosome
repositioning, and binding of the transcription factor CTCF during this process.
By integrating MNase-seq and ChIP-seq experiments in mouse embryonic stem cells
(ESC) and their differentiated counterparts with biophysical modeling, we found
that the interplay between these factors depends on their genomic context. The
mostly unmethylated CpG islands have reduced nucleosome occupancy and are
enriched in cell type-independent binding sites for CTCF. The few remaining
methylated CpG dinucleotides are preferentially associated with nucleosomes. In
contrast, outside of CpG islands most CpGs are methylated, and the average
methylation density oscillates so that it is highest in the linker region
between nucleosomes. Outside CpG islands, binding of TET1, an enzyme that
converts 5mC to 5hmC, is associated with labile, MNase-sensitive nucleosomes.
Such nucleosomes are poised for eviction in ESCs and become stably bound in
differentiated cells where the TET1 and 5hmC levels go down. This process
regulates a class of CTCF binding sites outside CpG islands that are occupied by
CTCF in ESCs but lose the protein during differentiation. We rationalize this
cell type-dependent targeting of CTCF with a quantitative biophysical model of
competitive binding with the histone octamer, depending on the TET1, 5hmC, and
5mC state. It is well established that cancer-associated epigenetic repression occurs
concomitant with CpG island hypermethylation and loss of nucleosomes at
promoters, but the role of nucleosome occupancy and epigenetic reprogramming at
distal regulatory elements in cancer is still poorly understood. Here, we
evaluate the scope of global epigenetic alterations at enhancers and insulator
elements in prostate and breast cancer cells using simultaneous genome-wide
mapping of DNA methylation and nucleosome occupancy (NOMe-seq). We find that the
genomic location of nucleosome-depleted regions (NDRs) is mostly cell type
specific and preferentially found at enhancers in normal cells. In cancer cells,
however, we observe a global reconfiguration of NDRs at distal regulatory
elements coupled with a substantial reorganization of the cancer methylome.
Aberrant acquisition of nucleosomes at enhancer-associated NDRs is associated
with hypermethylation and epigenetic silencing marks, and conversely, loss of
nucleosomes with demethylation and epigenetic activation. Remarkably, we show
that nucleosomes remain strongly organized and phased at many facultative distal
regulatory elements, even in the absence of a NDR as an anchor. Finally, we find
that key transcription factor (TF) binding sites also show extensive peripheral
nucleosome phasing, suggesting the potential for TFs to organize NDRs
genome-wide and contribute to deregulation of cancer epigenomes. Together, our
findings suggest that "decommissioning" of NDRs and TFs at distal regulatory
elements in cancer cells is accompanied by DNA hypermethylation susceptibility
of enhancers and insulator elements, which in turn may contribute to an altered
genome-wide architecture and epigenetic deregulation in maligcy. The fundamental repeating unit of eukaryotic chromatin is the nucleosome.
Besides being involved in packaging DNA, nucleosome organization plays an
important role in transcriptional regulation and cellular identity. Currently,
there is much debate about the major determits of the nucleosome architecture
of a genome and its significance with little being known about its role in stem
cells. To address these questions, we performed ultra-deep sequencing of
nucleosomal DNA in two human embryonic stem cell lines and integrated our data
with numerous epigenomic maps. Our analyses have revealed that the genome is a
determit of nucleosome organization with transcriptionally inactive regions
characterized by a "ground state" of nucleosome profiles driven by underlying
DNA sequences. DNA sequence preferences are associated with heterogeneous
chromatin organization around transcription start sites. Transcription, histone
modifications, and DNA methylation alter this "ground state" by having distinct
effects on both nucleosome positioning and occupancy. As the transcriptional
rate increases, nucleosomes become better positioned. Exons transcribed and
included in the final spliced mRNA have distinct nucleosome profiles in
comparison to exons not included at exon-exon junctions. Genes marked by the
active modification H3K4m3 are characterized by lower nucleosome occupancy
before the transcription start site compared to genes marked by the inactive
modification H3K27m3, while bivalent domains, genes associated with both marks,
lie exactly in the middle. Combinatorial patterns of epigenetic marks (chromatin
states) are associated with unique nucleosome profiles. Nucleosome organization
varies around transcription factor binding in enhancers versus promoters. DNA
methylation is associated with increasing nucleosome occupancy and different
types of methylations have distinct location preferences within the nucleosome
core particle. Finally, computational analysis of nucleosome organization alone
is sufficient to elucidate much of the circuitry of pluripotency. Our results,
suggest that nucleosome organization is associated with numerous genomic and
epigenomic processes and can be used to elucidate cellular identity. DNA methylation and nucleosome positioning are two key mechanisms that
contribute to the epigenetic control of gene expression. During carcinogenesis,
the expression of many genes is altered alongside extensive changes in the
epigenome, with repressed genes often being associated with local DNA
hypermethylation and gain of nucleosomes at their promoters. However the
spectrum of alterations that occur at distal regulatory regions has not been
extensively studied. To address this we used Nucleosome Occupancy and
Methylation sequencing (NOMe-seq) to compare the genome-wide DNA methylation and
nucleosome occupancy profiles between normal and cancer cell line models of the
breast and prostate. Here we describe the bioinformatic pipeline and methods
that we developed for the processing and analysis of the NOMe-seq data published
by (Taberlay et al., 2014 [1]) and deposited in the Gene Expression Omnibus with
accession GSE57498. |
Which R/Bioconductor package has been developed for the analysis of psychiatric disease genes? | PsyGeNET is a knowledge resource on psychiatric diseases and their genes, developed by text mining and curated by domain experts. Psygenet2r is an R package that contains a variety of functions for leveraging PsyGeNET database and facilitating its analysis and interpretation. The package offers different types of queries to the database along with variety of analysis and visualization tools, including the study of the anatomical structures in which the genes are expressed and gaining insight of gene's molecular function. Psygenet2r is especially suited for network medicine analysis of psychiatric disorders. | |
Which Janus kinase does decernotinib target? | Decernotinib (VX-509) is a potent and selective inhibitor of janus kinase 3 (JAK3). | Cytokines, growth factors, and other chemical messengers rely on a class of
intracellular nonreceptor tyrosine kinases known as Janus kinases (JAKs) to
rapidly transduce intracellular signals. A number of these cytokines are
critical for lymphocyte development and mediating immune responses. JAK3 is of
particular interest due to its importance in immune function and its expression,
which is largely confined to lymphocytes, thus limiting the potential impact of
JAK3 inhibition on nonimmune physiology. The aim of this study was to evaluate
the potency and selectivity of the investigational JAK3 inhibitor VX-509
(decernotinib)
[(R)-2-((2-(1H-pyrrolo[2,3-b]pyridin-3-yl)pyrimidin-4-yl)amino)-2-methyl-N-(2,2,2-trifluoroethyl)butanamide]
against JAK3 kinase activity and inhibition of JAK3-mediated signaling in vitro
and JAK3-dependent physiologic processes in vivo. These results demonstrate that
VX-509 potently inhibits JAK3 in enzyme assays (Ki = 2.5 nM + 0.7 nM) and
cellular assays dependent on JAK3 activity (IC50 range, 50-170 nM), with limited
or no measurable potency against other JAK isotypes or non-JAK kinases. VX-509
also showed activity in two animal models of aberrant immune function. VX-509
treatment resulted in dose-dependent reduction in ankle swelling and paw weight
and improved paw histopathology scores in the rat collagen-induced arthritis
model. In a mouse model of oxazolone-induced delayed-type hypersensitivity,
VX-509 reduced the T cell-mediated inflammatory response in skin. These findings
demonstrate that VX-509 is a selective and potent inhibitor of JAK3 in vitro and
modulates proinflammatory response in models of immune-mediated diseases, such
as collagen-induced arthritis and delayed-type hypersensitivity. The data
support evaluation of VX-509 for treatment of patients with autoimmune and
inflammatory diseases such as rheumatoid arthritis. |
What is the genetic basis of the Delayed Sleep-Phase Syndrome (DSPS)? | Circadian gene mutations are also associated with circadian rhythm disorders such as familial advanced sleep phase syndrome, delayed sleep phase syndrome, and non-24-hour sleep-wake syndrome. Possible association of human leucocyte antigen DR1 with delayed sleep phase syndrome. The study investigated the human leucocyte antigen (HLA), types A, B and DR, of 42 patients with delayed sleep phase syndrome (DSPS) and compared the frequencies of the antigens with those in 117 healthy controls. The comparison revealed that the gene frequencies and positivities of HLA-A, -B and -DR, except for DR1, had no significant differences between the patients and controls. The frequency of HLA-DR1 was increased in the DSPS patients as compared with that in the healthy controls (P = 0.0069 in positivity). In human leukocyte antigen (HLA) typing, the incidence of DR1 positivity alone was significantly higher in DSPS patients than in healthy subjects. | The study investigated the human leucocyte antigen (HLA), types A, B and DR, of
42 patients with delayed sleep phase syndrome (DSPS) and compared the
frequencies of the antigens with those in 117 healthy controls. The comparison
revealed that the gene frequencies and positivities of HLA-A, -B and -DR, except
for DR1, had no significant differences between the patients and controls. The
frequency of HLA-DR1 was increased in the DSPS patients as compared with that in
the healthy controls (P = 0.0069 in positivity). Although the corrected P-value
(0.069) for multiple comparisons almost reached the significance level, the
results indicated a possible association of the HLA-DR1 antigen with DSPS. This
study suggests that there are genetic predispositions to DSPS. We classified 64 patients with chronic delayed sleep phase syndrome (DSPS) into
the primary (n = 53) and secondary (n = 11) group according to presence or
absence of such signs as difficulty in waking up which appeared much earlier
than the onset of DSPS. The age at the onset of the early signs concentrated in
adolescence. The familial occurrence of DSPS was noted in 11 patients of the
primary group. In human leukocyte antigen (HLA) typing, the incidence of DR1
positivity alone was significantly higher in DSPS patients than in healthy
subjects. Minnesota Multiphasic Personality Inventory revealed high scores on
depression, psychoasthenia and hypochondriasis. We suggest that a predisposition
to DSPS includes biological, genetic, social and psychological factors, various
combinations of which may lead to DSPS. Recent progress in biological clock research has facilitated genetic analysis of
circadian rhythm sleep disorders, such as delayed sleep phase syndrome (DSPS)
and non-24-h sleep-wake syndrome (N-24). We analyzed the human period3 (hPer3)
gene, one of the human homologs of the Drosophila clock-gene period (Per), as a
possible candidate for rhythm disorder susceptibility. All of the coding exons
in the hPer3 gene were screened for polymorphisms by a PCR-based strategy using
genomic DNA samples from sleep disorder patients and control subjects. We
identified six sequence variations with amino acid changes, of which five were
common and predicted four haplotypes of the hPer3 gene. One of the haplotypes
was significantly associated with DSPS (Bonferroni's corrected P = 0.037; odds
ratio = 7.79; 95% CI 1.59-38.3) in our study population. Our results suggest
that structural polymorphisms in the hPer3 gene may be implicated in the
pathogenesis of DSPS. Arylalkylamine N-acetyltransferase (AA-NAT) is a rate-limiting enzyme in
melatonin hormone synthesis and participates in daily oscillations of the
melatonin level. We studied the association between the AA-NAT gene and delayed
sleep phase syndrome (DSPS). Results indicate that there is a significant
difference in allele positivity at the single nucleotide polymorphism involved
in an amino acid substitution from alanine to threonine at position 129 between
patients with DSPS and healthy controls. The frequency of the 129 threonine
allele is significantly higher in the patients than in the controls ( P=0.0029).
The data suggest that AA-NAT could be a susceptibility gene for DSPS. Depression is one of the leading causes of morbidity worldwide and represents a
huge burden to society. As with many other psychiatric disorders, a genetic
basis for depression has been identified. Evidence for the role of circadian
genes in depression is particularly compelling. Circadian gene mutations are
also associated with circadian rhythm disorders such as familial advanced sleep
phase syndrome, delayed sleep phase syndrome, and non-24-hour sleep-wake
syndrome. Such disorders, plus the other manifestations of a disrupted circadian
system such as hormone dysregulation, are often observed among those with
depression. This suggests a shared aetiology between circadian disruption and
depression, although the exact mechanisms underlying the association are
unclear. This paper reviews the molecular mechanisms involved in depression,
with an emphasis on circadian genes. Twin studies in depression have reported
probandwise concordance rates of 40% and 70% using narrow and broad diagnostic
criteria, respectively, and heritability of over 85% for bipolar disorder. In
association studies, increased susceptibility to depression has been noted in
those with polymorphisms in the following: D-amino-acid-oxidase activator/G30
gene complex, glucocorticoid receptor gene, serotonin transporter gene,
tryptophan hydroxylase 2 gene, dopamine transporter gene and G protein-coupled
receptor 50 gene. Polymorphisms in these genes have also been linked to a better
or worse response to antidepressant therapy, an increased likelihood of
responding poorly to adversity and increased suicide ideation. Polymorphisms in
the CLOCK, BMAL1, Per3 and TIMELESS genes have been associated with
susceptibility to mood disorder, and single nucleotide polymorphisms and
haplotypes in several circadian genes have been observed among those displaying
certain circadian phenotypes, including worse mood in the evening, insomnia in
mania and early, middle or late insomnia in depression. Manipulation of the
circadian timing system via sleep deprivation, bright light or pharmacological
therapy has also been shown to alleviate depressive symptoms, providing further
evidence for the role of circadian dysfunction in depression pathophysiology.
The new antidepressant agomelatine is the first melatonergic antidepressant with
an innovative mode of action: it is a melatonergic MT(1), MT(2) receptor agonist
and 5-HT(2c) antagonist, and is able to restore the internal clock, which is
profoundly disturbed in depression, thus being efficacious in major depressive
disorders. In conclusion, a wealth of evidence is now available supporting a
genetic basis for depression. The apparent importance of mutations in the
circadian genes in determining disease susceptibility, disease recurrence and
response to treatment suggests that the circadian pathway represents an
attractive target for pharmacological manipulation to improve management of this
debilitating disorder. |
What are jakinibs? | Jakinibs are Janus kinase (JAK) inhibitors. They are considered a new class of kinase inhibitors in cancer and autoimmune disease. Jakinibs can differ substantially in their selectivity against JAKs. | Cytokines are critical for normal cell growth and immunoregulation but also
contribute to growth of maligt cells and drive immune-mediated disease. A
large subset of immunoregulatory cytokines uses the type I and type II cytokine
receptors and pharmacological targeting of these cytokines/cytokines receptors
has proven to be efficacious in treating immune and inflammatory diseases. These
receptors rely on Janus family of kinases (Jaks) for signal transduction.
Recently the first Jak inhibitor (jakinib) has been approved by the FDA and a
second has been recommended for approval. Many other Jakinibs are likely to
follow and in this brief review, we will discuss the state-of-the art of this
new class of pharmacological agents. |
What is the function of the gene MDA5? | Melanoma differentiation-associated gene 5 (MDA5) is a pattern recognition receptor that recognizes cytoplasmic viral double-stranded RNA (dsRNA) and initiates rapid innate antiviral responses. MDA5 forms a filament-like multimer along the dsRNA leading to oligomerization, which in turn activates the adaptor protein mitochondrial antiviral signaling protein (MAVS) to provide a signal platform for the induction of type I interferon (IFN) and proinflammatory cytokines. The conformational switch of MDA5 causes antiviral defense, but excessive activation of the MDA5-MAVS pathway may result in autoimmune diseases. | Sensing of viral RNA by the cytosolic receptors RIG-I and melanoma
differentiation-associated gene 5 (MDA5) leads to innate antiviral response. How
RIG-I and MDA5 are dynamically regulated in innate antiviral response is not
well understood. Here, we show that TRIM38 positively regulates MDA5- and
RIG-I-mediated induction of downstream genes and acts as a SUMO E3 ligase for
their dynamic sumoylation at K43/K865 and K96/K888, respectively, before and
after viral infection. The sumoylation of MDA5 and RIG-I suppresses their
K48-linked polyubiquitination and degradation in uninfected or early-infected
cells. Sumoylation of the caspase recruitment domains of MDA5 and RIG-I is also
required for their dephosphorylation by PP1 and activation upon viral infection.
At the late phase of viral infection, both MDA5 and RIG-I are desumoylated by
SENP2, resulting in their K48-linked polyubiquitination and degradation. These
findings suggest that dynamic sumoylation and desumoylation of MDA5 and RIG-I
modulate efficient innate immunity to RNA virus and its timely termination. The innate immune system plays a critical role in pathogen recognition and
initiation of protective immune response through the recognition of pathogen
associated molecular patterns (PAMPs) by its pattern recognition receptors
(PRRs). Nucleic acids including RNA and DNA have been recognized as very
important PAMPs of pathogens especially for viruses. RNA are the major PAMPs of
RNA viruses, to which most severe disease causing viruses belong thus posing a
tougher challenge to human and animal health. Therefore, the understanding of
the immune biology of RNA PRRs is critical for control of pathogen infections
especially for RNA virus infections. RNA PRRs are comprised of TLR3, TLR7, TLR8,
RIG-I, MDA5, NLRP3, NOD2, and some other minorities. This review introduces
these RNA PRRs by describing the cellular localizations, ligand recognitions,
activation mechanisms, cell signaling pathways, and recognition of pathogens;
the cross-talks between various RNA PRRs are also reviewed. The deep insights of
these RNA PRRs can be utilized to improve anti-viral immune response. © 2017
IUBMB Life, 69(5):297-304, 2017. Human noroviruses are a major cause of acute gastroenteritis worldwide, but the
lack of a robust cell culture system or small animal model have hampered a
better understanding of innate immunity against these viruses. Tulane virus (TV)
is the prototype virus of a tentative new genus, Recovirus, in the family
Caliciviridae. Its epidemiology and biological properties most closely resemble
human norovirus. The host innate immune response to RNA virus infection
primarily involves pathogen-sensing toll-like receptors (TLRs) TLR3 and TLR7 and
retinoic acid-inducible gene I-like receptor RIG-I and melanoma differentiation
associated gene 5 (MDA5). In this study, by using siRNA knockdown, we report
that TV infection in LLC-MK2 cells results in an early [3 h post infection (h
p.i.), P<0.05] RIG-I-dependent and type I interferon-mediated antiviral
response, whereas an MDA5-mediated antiviral effect was observed at later (12 h
p.i.; P<0.05) stages of TV replication. Induction of RIG-I and MDA5 was critical
for inhibition of TV replication. Furthermore, pre-activation of the RIG-I/MDA5
pathway prevented TV replication (>900-fold decrease; P<0.05), suggesting that
RIG-I and MDA5 ligands could be used to develop novel preventive and therapeutic
measures against norovirus. Author information:
(1)Department of Immunology & Center for Immunotherapy, Institute of Basic
Medical Sciences, Peking Union Medical College, Chinese Academy of Medical
Sciences, Beijing 100005, China.
(2)Department of Immunology, School of Basic Medical Science, Shandong
University, Ji, Shandong 250012, China; State Key Laboratory of Microbial
Technology, Shandong University, Ji, Shandong 250012, China.
(3)Department of Immunology, School of Basic Medical Science, Shandong
University, Ji, Shandong 250012, China.
(4)Department of Immunology, School of Basic Medical Science, Shandong
University, Ji, Shandong 250012, China; State Key Laboratory of Microbial
Technology, Shandong University, Ji, Shandong 250012, China. Electronic
address: [email protected].
(5)Department of Immunology & Center for Immunotherapy, Institute of Basic
Medical Sciences, Peking Union Medical College, Chinese Academy of Medical
Sciences, Beijing 100005, China; National Key Laboratory of Medical Immunology &
Institute of Immunology, Second Military Medical University, Shanghai, China.
Electronic address: [email protected]. |
Which is the conserved motif of DEAD box proteins? | The conserved motif is: Asp(D)-Glu-(E)-Ala(A)-Asp(D) | |
Which individuals show preferential colonization of the Prevotellaceae bacteria in their guts? | Although the distinction of enterotypes as either discrete clusters or a continuum will require additional investigation, numerous studies have demonstrated the co-exclusion of the closely related Prevotellaceae and Bacteroides genera in the gut microbiota of healthy human subjects where Prevotella appears to be a discriminatory taxon for residence in more agrarian societies. | Recent evidence suggests that the microbial community in the human intestine may
play an important role in the pathogenesis of obesity. We examined 184,094
sequences of microbial 16S rRNA genes from PCR amplicons by using the 454
pyrosequencing technology to compare the microbial community structures of 9
individuals, 3 in each of the categories of normal weight, morbidly obese, and
post-gastric-bypass surgery. Phylogenetic analysis demonstrated that although
the Bacteria in the human intestinal community were highly diverse, they fell
mainly into 6 bacterial divisions that had distinct differences in the 3 study
groups. Specifically, Firmicutes were domit in normal-weight and obese
individuals but significantly decreased in post-gastric-bypass individuals, who
had a proportional increase of Gammaproteobacteria. Numbers of the
H(2)-producing Prevotellaceae were highly enriched in the obese individuals.
Unlike the highly diverse Bacteria, the Archaea comprised mainly members of the
order Methanobacteriales, which are H(2)-oxidizing methanogens. Using real-time
PCR, we detected significantly higher numbers of H(2)-utilizing methanogenic
Archaea in obese individuals than in normal-weight or post-gastric-bypass
individuals. The coexistence of H(2)-producing bacteria with relatively high
numbers of H(2)-utilizing methanogenic Archaea in the gastrointestinal tract of
obese individuals leads to the hypothesis that interspecies H(2) transfer
between bacterial and archaeal species is an important mechanism for increasing
energy uptake by the human large intestine in obese persons. The large bacterial
population shift seen in the post-gastric-bypass individuals may reflect the
double impact of the gut alteration caused by the surgical procedure and the
consequent changes in food ingestion and digestion. Two anaerobic, non-spore-forming, pleomorphic, Gram-negative rods, designated
YIT 11840T and YIT 11841T, were isolated from human faeces. The organisms were
catalase-negative, produced succinic and acetic acids as end products of glucose
metabolism and had DNA G+C contents of approximately 48-49 mol%. Although the
phenotypic characteristics of these two strains were very similar, analysis of
their 16S rRNA gene sequences showed that they are only distantly related
(93.8%), indicating that they represent two different species. A comparative
sequence analysis revealed that these two species are members of the family
'Prevotellaceae' but are phylogenetically distant (<88% sequence similarity)
from the known genera belonging to this family, including Prevotella, Hallela
and Xylanibacter. On the basis of the phylogenetic analysis and physiological
tests, strains YIT 11840T and YIT 11841T represent two novel species of a new
genus, for which the names Paraprevotella clara gen. nov., sp. nov. (type strain
YIT 11840T=JCM 14859T=DSM 19731T), the type species, and Paraprevotella
xylaniphila sp. nov. (type strain YIT 11841T=JCM 14860T=DSM 19681T) are
proposed. The human gut is the natural habitat for a large and dynamic bacterial community
that has a great relevance for health. Metagenomics is increasing our knowledge
of gene content as well as of functional and genetic variability in this
microbiome. However, little is known about the active bacteria and their
function(s) in the gastrointestinal tract. We performed a metatranscriptomic
study on ten healthy volunteers to elucidate the active members of the gut
microbiome and their functionality under conditions of health. First, the
microbial cDNAs obtained from each sample were sequenced using 454 technology.
The analysis of 16S transcripts showed the phylogenetic structure of the active
microbial community. Lachnospiraceae, Ruminococcaceae, Bacteroidaceae,
Prevotellaceae, and Rickenellaceae were the predomit families detected in the
active microbiota. The characterization of mRNAs revealed a uniform functional
pattern in healthy individuals. The main functional roles of the gut microbiota
were carbohydrate metabolism, energy production and synthesis of cellular
components. In contrast, housekeeping activities such as amino acid and lipid
metabolism were underrepresented in the metatranscriptome. Our results provide
new insights into the functionality of the complex gut microbiota in healthy
individuals. In this RNA-based survey, we also detected small RNAs, which are
important regulatory elements in prokaryotic physiology and pathogenicity. The human gut contains a vast number of microorganisms known collectively as the
"gut microbiota". Despite its importance in maintaining the health of the host,
growing evidence suggests the gut microbiota may also be an important factor in
the pathogenesis of various diseases, a number of which have shown a rapid
increase in incidence over the past few decades. Factors including age,
genetics, and diet may influence microbiota composition. We used diet
inventories and 16S rDNA sequencing to characterize fecal samples from 98
individuals. Fecal communities clustered into previously described enterotypes
distinguished primarily by levels of Bacteroides and Prevotella. Enterotypes
were associated with long-term diets, particularly protein and animal fat
(Bacteroides) vs. simple carbohydrates (Prevotella). Although the distinction of
enterotypes as either discrete clusters or a continuum will require additional
investigation, numerous studies have demonstrated the co-exclusion of the
closely related Prevotellaceae and Bacteroides genera in the gut microbiota of
healthy human subjects where Prevotella appears to be a discriminatory taxon for
residence in more agrarian societies. Ultimately, the impact of diet on the
human gut microbiota may be an important environmental factor involved in the
pathogenesis of disease states that show a rapidly increasing incidence in
industrialized nations such as the inflammatory bowel diseases (IBD). We investigated the effect of consuming probiotic fermented milk (PFM) on the
microbial community structure in the human intestinal tract by using
high-throughput barcoded pyrosequencing. Six healthy adults ingested 2 servings
of PFM daily for 3 wk, and their fecal microbiota were analyzed before and after
3 wk of PFM ingestion period and for another 3 wk following the termination of
PFM ingestion (the noningestion period). Fecal microbial communities were
characterized by sequencing of the V1-V3 hypervariable regions of the 16S rRNA
gene. All subjects showed a similar pattern of microbiota at the phylum level,
where the relative abundance of Bacteriodetes species increased during the PFM
ingestion period and decreased during the noningestion period. The increase in
Bacteroidetes was found to be due to an increase in members of the families
Bacteroidaceae or Prevotellaceae. In contrast to PFM-induced adaptation at the
phylum level, the taxonomic composition at the genus level showed a considerable
alteration in fecal microbiota induced by PFM ingestion. As revealed by analysis
of operational taxonomic units (OTU), the numbers of shared OTU were low among
the 3 different treatments (before, during, and after PFM ingestion), but the
abundance of the shared OTU was relatively high, indicating that the majority
(>77.8%) of total microbiota was maintained by shared OTU during PFM ingestion
and after its termination. Our results suggest that PFM consumption could alter
microbial community structure in the gastrointestinal tract of adult humans
while maintaining the stability of microbiota. BACKGROUND: Oxygen is generally considered essential for lethal
photosensitisation by photodynamic processes. The oral anaerobes, Prevotella
intermedia and P. nigrescens are known to be photosensitive, but are also
extremely sensitive to the cytotoxic effects of oxygen.
METHODS: The Prevotellaceae were exposed to two 405 nm light sources for
different exposure times in an anaerobic chamber. Viable counts of the light
exposed samples were compared to light-free controls to determine the proportion
of bacteria killed.
RESULTS: Lethal photosensitivity was demonstrated against P. intermedia and P.
nigrescens. The proportions of bacteria killed by either the light-emitting
diode or laser pointer were similar at a given energy density (J/cm(2)).
CONCLUSIONS: Lethal photosensitivity was demonstrated in two species of
Prevotella under anaerobic conditions. Author information:
(1)Aix Marseille Université, URMITE, UM63, CNRS 7278, IRD 198, INSERM 1095,
Institut Hospitalo-Universitaire Méditerranée-Infection, Faculté de médecine,
Marseille, France.
(2)Aix Marseille Université, URMITE, UM63, CNRS 7278, IRD 198, INSERM 1095,
Institut Hospitalo-Universitaire Méditerranée-Infection, Faculté de médecine,
Marseille, France; Special Infectious Agents Unit, King Fahd Medical Research
Center, King Abdulaziz University, Jeddah, Saudi Arabia. |
Describe GARLIC (GWAS-based Prediction Toolkit for Connecting Diseases and Cell Types) | GARLIC (GWAS-based Prediction Toolkit for Connecting Diseases and Cell Types) is a bioinformatic toolkit for aetiologically connecting diseases and cell type-specific regulatory maps. GARLIC can be used to retrieve potential disease-causative genetic variants overlapping regulatory sequences of interest. Overall, GARLIC can satisfy several important needs within the field of medical genetics, thus potentially assisting in the ultimate goal of uncovering the elusive and complex genetic basis of common human disorders. | Genome-wide association studies (GWAS) have emerged as a powerful tool to
uncover the genetic basis of human common diseases, which often show a complex,
polygenic and multi-factorial aetiology. These studies have revealed that 70-90%
of all single nucleotide polymorphisms (SNPs) associated with common complex
diseases do not occur within genes (i.e. they are non-coding), making the
discovery of disease-causative genetic variants and the elucidation of the
underlying pathological mechanisms far from straightforward. Based on emerging
evidences suggesting that disease-associated SNPs are frequently found within
cell type-specific regulatory sequences, here we present GARLIC (GWAS-based
Prediction Toolkit for Connecting Diseases and Cell Types), a user-friendly,
multi-purpose software with an associated database and online viewer that, using
global maps of cis-regulatory elements, can aetiologically connect human
diseases with relevant cell types. Additionally, GARLIC can be used to retrieve
potential disease-causative genetic variants overlapping regulatory sequences of
interest. Overall, GARLIC can satisfy several important needs within the field
of medical genetics, thus potentially assisting in the ultimate goal of
uncovering the elusive and complex genetic basis of common human disorders. |
How does Dst1 knock-out affect transcription in yeast? | While TFIIS has a pronounced effect on transcription elongation in vitro, the deletion of DST1 has no major effect on cell viability. We showed that: dst1 and rpb9 mutants have a synthetic growth defective phenotype when combined with fyv4, gim5, htz1, yal011w, ybr231c, soh1, vps71, and vps72 mutants that is exacerbated during germination or at high salt concentrations. In vivo, some dst1 mutants were partly defective in tRNA synthesis and showed a reduced Pol III occupancy at the restrictive temperature. | Deletions of three yeast genes, SET2, CDC73, and DST1, involved in
transcriptional elongation and/or chromatin metabolism were used in conjunction
with genetic array technology to screen approximately 4700 yeast deletions and
identify double deletion mutants that produce synthetic growth defects. Of the
five deletions interacting genetically with all three starting mutations, one
encoded the histone H2A variant Htz1 and three encoded components of a novel 13
protein complex, SWR-C, containing the Snf2 family ATPase, Swr1. The SWR-C also
copurified with Htz1 and Bdf1, a TFIID-interacting protein that recognizes
acetylated histone tails. Deletions of the genes encoding Htz1 and seven
nonessential SWR-C components caused a similar spectrum of synthetic growth
defects when combined with deletions of 384 genes involved in transcription,
suggesting that Htz1 and SWR-C belong to the same pathway. We show that
recruitment of Htz1 to chromatin requires the SWR-C. Moreover, like Htz1 and
Bdf1, the SWR-C promotes gene expression near silent heterochromatin. Transcription by RNA polymerase II (pol II) requires the ordered binding of
distinct protein complexes to catalyze initiation, elongation, termination, and
coupled mRNA processing events. One or more proteins from each complex are known
to bind pol II via the carboxy-terminal domain (CTD) of the largest subunit,
Rpb1. How binding is coordinated is not known, but it might involve
conformational changes in the CTD induced by the Ess1 peptidyl-prolyl cis/trans
isomerase. Here, we examined the role of ESS1 in transcription by studying one
of its multicopy suppressors, BYE1. We found that Bye1 is a negative regulator
of transcription elongation. This led to the finding that Ess1 also inhibits
elongation; Ess1 opposes elongation factors Dst1 and Spt4/5, and overexpression
of ESS1 makes cells more sensitive to the elongation inhibitor 6-AU. In reporter
gene assays, ess1 mutations reduce the ability of elongation-arrest sites to
stall polymerase. We also show that Ess1 acts positively in transcription
termination, independent of its role in elongation. We propose that Ess1-induced
conformational changes attenuate pol II elongation and help coordinate the
ordered assembly of protein complexes on the CTD. In this way, Ess1 might
regulate the transition between multiple steps of transcription. TFIIS promotes the intrinsic ability of RNA polymerase II to cleave the 3'-end
of the newly synthesized RNA. This stimulatory activity of TFIIS, which is
dependent upon Rpb9, facilitates the resumption of transcription elongation when
the polymerase stalls or arrests. While TFIIS has a pronounced effect on
transcription elongation in vitro, the deletion of DST1 has no major effect on
cell viability. In this work we used a genetic approach to increase our
knowledge of the role of TFIIS in vivo. We showed that: (1) dst1 and rpb9
mutants have a synthetic growth defective phenotype when combined with fyv4,
gim5, htz1, yal011w, ybr231c, soh1, vps71, and vps72 mutants that is exacerbated
during germination or at high salt concentrations; (2) TFIIS and Rpb9 are
essential when the cells are challenged with microtubule-destabilizing drugs;
(3) among the SDO (synthetic with Dst one), SOH1 shows the strongest genetic
interaction with DST1; (4) the presence of multiple copies of TAF14, SUA7,
GAL11, RTS1, and TYS1 alleviate the growth phenotype of dst1 soh1 mutants; and
(5) SRB5 and SIN4 genetically interact with DST1. We propose that TFIIS is
required under stress conditions and that TFIIS is important for the transition
between initiation and elongation in vivo. TFIIS, an elongation factor encoded by DST1 in Saccharomyces cerevisiae,
stimulates transcript cleavage in arrested RNA polymerase II. Two components of
the RNA polymerase II machinery, Med13 (Srb9) and Spt8, were isolated as
two-hybrid partners of the conserved TFIIS N-terminal domain. They belong to the
Cdk8 module of the Mediator and to a subform of the SAGA co-activator,
respectively. Co-immunoprecipitation experiments showed that TFIIS can bind the
Cdk8 module and SAGA in cell-free extracts. spt8Delta and dst1Delta mutants were
sensitive to nucleotide-depleting drugs and epistatic to null mutants of the RNA
polymerase II subunit Rpb9, suggesting that their elongation defects are
mediated by Rpb9. rpb9Delta, spt8Delta and dst1Delta were lethal in cells
lacking the Rpb4 subunit. The TFIIS N-terminal domain is also strictly required
for viability in rpb4Delta, although it is not needed for binding to RNA
polymerase II or for transcript cleavage. It is proposed that TFIIS and the
Spt8-containing form of SAGA co-operate to rescue RNA polymerase II from
unproductive elongation complexes, and that the Cdk8 module temporarily blocks
transcription during transcript cleavage. TFIIS is a transcription elongation factor that stimulates transcript cleavage
activity of arrested RNA polymerase II (Pol II). Recent studies revealed that
TFIIS has also a role in Pol II transcription initiation. To improve our
understanding of TFIIS function in vivo, we performed genome-wide location
analysis of this factor. Under normal growth conditions, TFIIS was detected on
Pol II-transcribed genes, and TFIIS occupancy was well correlated with that of
Pol II, indicating that TFIIS recruitment is not restricted to NTP-depleted
cells. Unexpectedly, TFIIS was also detected on almost all Pol III-transcribed
genes. TFIIS and Pol III occupancies correlated well genome-wide on this novel
class of targets. In vivo, some dst1 mutants were partly defective in tRNA
synthesis and showed a reduced Pol III occupancy at the restrictive temperature.
In vitro transcription assays suggested that TFIIS may affect Pol III start site
selection. These data provide strong in vivo and in vitro evidence in favor of a
role of TFIIS as a general Pol III transcription factor. Yeast Rpb4, a subunit of RNA pol II is not essential for viability but is
involved in multiple cellular phenotypes such as temperature sensitivity,
enhanced pseudohyphal morphology, and decreased sporulation. Both in vivo and in
vitro studies strongly support involvement of Rpb4 in transcription initiation,
while its role in transcription elongation is not entirely consistent. Here we
show that Rpb4 is not required for recruitment of RNA pol II on the coding
region of YLR454w, a representative long gene. Yet we find strong genetic
interaction of rpb4∆ with mutants in many transcription elongation factors such
as Paf1, Spt4, Dst1, Elp3 and Rpb9. We demonstrate that, Rpb4 interacts
functionally with Paf1 to affect the transcription elongation of the FKS1 gene.
Our results suggest that while Rpb4 is not required for general transcription
elongation, it could support transcription elongation for specific of class of
genes by interaction with other elongation factors. Abasic or AP sites generated by spontaneous DNA damage accumulate at a higher
rate in actively transcribed regions of the genome in S. cerevisiae and are
primarily repaired by base excision repair (BER) pathway. We have demonstrated
that transcription-coupled nucleotide excision repair (NER) pathway can
functionally replace BER to repair those AP sites located on the transcribed
strand much like the strand specific repair of UV-induced pyrimidine dimers.
Previous reports indicate that Rad26, a yeast homolog of transcription-repair
coupling factor CSB, partly mediates strand-specific repair of UV-dimers as well
as AP lesions. Here, we report that Def1, known to promote ubiquitination and
degradation of stalled RNA polymerase complex, also directs NER to AP lesions on
the transcribed strand of an actively transcribed gene but that its function is
dependent on metabolic state of the yeast cells. We additionally show that Dst1,
a homolog of mammalian transcription elongation factor TFIIS, interferes with
NER-dependent repair of AP lesions while suppressing homologous recombination
pathway. Overall, Def1 and Dst1 mediate very different outcomes in response to
AP-induced transcription arrest. |
Which drug can be reversed with idarucizumab? | Idarucizumab is an antibody fragment that specifically reverses dabigatran mediated anticoagulation. | Urgent surgery or life-threatening bleeding requires prompt reversal of the
anticoagulant effects of dabigatran. This study assessed the ability of three-
and four-factor prothrombin complex concentrate (PCC) and idarucizumab (specific
antidote for dabigatran) to reverse the anticoagulant effects of dabigatran in a
porcine model of trauma. Twelve animals were given dabigatran etexilate (DE)
orally and dabigatran intravenously, before infliction of trauma. Six animals
received tranexamic acid plus fibrinogen concentrate 12 minutes post-injury. Six
PCCs (each 30 and 60 U/kg) and idarucizumab (30 and 60 mg/kg) were added to
blood samples ex vivo. Coagulation was assessed by several coagulation assays.
All coagulation parameters were altered after dabigatran infusion (plasma level:
442 ± 138 ng/ml). Both three- and four-factor PCCs mostly or completely reversed
the effects of dabigatran on thromboelastometry variables and PT but not on
aPTT. Idarucizumab neutralised plasma concentrations of dabigatran, and reversed
the effects of the drug on coagulation variables. Thrombin generation showed
dose-dependent over-correction following the addition of PCC, implying that
elevated levels of thrombin are required to overcome dabigatran-induced
coagulopathy. In contrast, treatment with idarucizumab returned thrombin
generation to baseline levels. Following trauma, therapy with tranexamic acid
plus fibrinogen improved correction of coagulation parameters by PCC, and
thromboelastometry parameters by idarucizumab. All investigated PCCs improved
dabigatran- and trauma-induced coagulopathy to a similar degree. In conclusion,
this study shows that three- and four-factor PCCs are similarly effective for
dabigatran reversal. Idarucizumab also reversed the effects of dabigatran and,
unlike PCCs, was not associated with over-correction of thrombin generation. Lack of specific antidotes is a major concern in intracerebral hemorrhage (ICH)
related to direct anticoagulants including dabigatran (OAC-ICH). We examined the
efficacy of idarucizumab, an antibody fragment binding to dabigatran, in a mouse
model of OAC-ICH. Dabigatran etexilate (DE) dose-dependently prolonged diluted
thrombin time and tail-vein bleeding time, which were reversed by idarucizumab.
Pretreatment with DE increased intracerebral hematoma volume and cerebral
hemoglobin content. Idarucizumab in equimolar dose prevented excess hematoma
expansion for both DE doses. In more extensive ICH, idarucizumab significantly
reduced mortality. Thus, idarucizumab prevents excess intracerebral hematoma
formation in mice anticoagulated with dabigatran and reduces mortality. BACKGROUND: Idarucizumab is a monoclonal antibody fragment that binds dabigatran
with high affinity in a 1:1 molar ratio. We investigated the safety,
tolerability, and efficacy of increasing doses of idarucizumab for the reversal
of anticoagulant effects of dabigatran in a two-part phase 1 study (rising-dose
assessment and dose-finding, proof-of-concept investigation). Here we present
the results of the proof-of-concept part of the study.
METHODS: In this randomised, placebo-controlled, double-blind, proof-of-concept
phase 1 study, we enrolled healthy volunteers (aged 18-45 years) with a
body-mass index of 18·5-29·9 kg/m(2) into one of four dose groups at SGS Life
Sciences Clinical Research Services, Belgium. Participants were randomly
assigned within groups in a 3:1 ratio to idarucizumab or placebo using a
pseudorandom number generator and a supplied seed number. Participants and care
providers were masked to treatment assignment. All participants received oral
dabigatran etexilate 220 mg twice daily for 3 days and a final dose on day 4.
Idarucizumab (1 g, 2 g, or 4 g 5-min infusion, or 5 g plus 2·5 g in two 5-min
infusions given 1 h apart) was administered about 2 h after the final dabigatran
etexilate dose. The primary endpoint was incidence of drug-related adverse
events, analysed in all randomly assigned participants who received at least one
dose of dabigatran etexilate. Reversal of diluted thrombin time (dTT), ecarin
clotting time (ECT), activated partial thromboplastin time (aPTT), and thrombin
time (TT) were secondary endpoints assessed by measuring the area under the
effect curve from 2 h to 12 h (AUEC2-12) after dabigatran etexilate ingestion on
days 3 and 4. This trial is registered with ClinicalTrials.gov, number
NCT01688830.
FINDINGS: Between Feb 23, and Nov 29, 2013, 47 men completed this part of the
study. 12 were enrolled into each of the 1 g, 2 g, or 5 g plus 2·5 g
idarucizumab groups (nine to idarucizumab and three to placebo in each group),
and 11 were enrolled into the 4 g idarucizumab group (eight to idarucizumab and
three to placebo). Drug-related adverse events were all of mild intensity and
reported in seven participants: one in the 1 g idarucizumab group (infusion site
erythema and hot flushes), one in the 5 g plus 2·5 g idarucizumab group
(epistaxis); one receiving placebo (infusion site haematoma), and four during
dabigatran etexilate pretreatment (three haematuria and one epistaxis).
Idarucizumab immediately and completely reversed dabigatran-induced
anticoagulation in a dose-dependent manner; the mean ratio of day 4 AUEC2-12 to
day 3 AUEC2-12 for dTT was 1·01 with placebo, 0·26 with 1 g idarucizumab (74%
reduction), 0·06 with 2 g idarucizumab (94% reduction), 0·02 with 4 g
idarucizumab (98% reduction), and 0·01 with 5 g plus 2·5 g idarucizumab (99%
reduction). No serious or severe adverse events were reported, no adverse event
led to discontinuation of treatment, and no clinically relevant difference in
incidence of adverse events was noted between treatment groups.
INTERPRETATION: These phase 1 results show that idarucizumab was associated with
immediate, complete, and sustained reversal of dabigatran-induced
anticoagulation in healthy men, and was well tolerated with no unexpected or
clinically relevant safety concerns, supporting further testing. Further
clinical studies are in progress.
FUNDING: Boehringer Ingelheim Pharma GmbH & Co KG. BACKGROUND: Specific reversal agents for non-vitamin K antagonist oral
anticoagulants are lacking. Idarucizumab, an antibody fragment, was developed to
reverse the anticoagulant effects of dabigatran.
METHODS: We undertook this prospective cohort study to determine the safety of 5
g of intravenous idarucizumab and its capacity to reverse the anticoagulant
effects of dabigatran in patients who had serious bleeding (group A) or required
an urgent procedure (group B). The primary end point was the maximum percentage
reversal of the anticoagulant effect of dabigatran within 4 hours after the
administration of idarucizumab, on the basis of the determination at a central
laboratory of the dilute thrombin time or ecarin clotting time. A key secondary
end point was the restoration of hemostasis.
RESULTS: This interim analysis included 90 patients who received idarucizumab
(51 patients in group A and 39 in group B). Among 68 patients with an elevated
dilute thrombin time and 81 with an elevated ecarin clotting time at baseline,
the median maximum percentage reversal was 100% (95% confidence interval, 100 to
100). Idarucizumab normalized the test results in 88 to 98% of the patients, an
effect that was evident within minutes. Concentrations of unbound dabigatran
remained below 20 ng per milliliter at 24 hours in 79% of the patients. Among 35
patients in group A who could be assessed, hemostasis, as determined by local
investigators, was restored at a median of 11.4 hours. Among 36 patients in
group B who underwent a procedure, normal intraoperative hemostasis was reported
in 33, and mildly or moderately abnormal hemostasis was reported in 2 patients
and 1 patient, respectively. One thrombotic event occurred within 72 hours after
idarucizumab administration in a patient in whom anticoagulants had not been
reinitiated.
CONCLUSIONS: Idarucizumab completely reversed the anticoagulant effect of
dabigatran within minutes. (Funded by Boehringer Ingelheim; RE-VERSE AD
ClinicalTrials.gov number, NCT02104947.). The vitamin K antagonists (VKAs) have been the standard (and only) oral
anticoagulants used for the long-term treatment or prevention of venous
thromboembolism or stroke in patients with atrial fibrillation. The coagulopathy
induced by VKAs can be reversed with vitamin K, and in urgent situations, the
vitamin K-dependent coagulation factors can be replaced by transfusion. In the
last decade, a new class of oral anticoagulants has been developed, direct oral
anticoagulants that bind to a specific coagulation factor and neutralize it.
These compounds were shown to be effective and safe compared with the VKAs and
were licensed for specific indications, but without a specific reversal agent.
The absence of a reversal agent is a barrier to more widespread use of these
agents. Currently, for the management of major life-threatening bleeding with
the direct oral anticoagulants, most authorities recommend the use of four
factor prothrombin complex concentrates. There are now three reversal agents in
development and poised to enter the market. Idarucizumab is a specific antidote
targeted to reverse the direct thrombin inhibitor, dabigatran, which was
recently approved for use in the USA. Andexanet alfa is an antidote targeted to
reverse the oral direct factor Xa inhibitors as well as the indirect inhibitor
enoxaparin. Ciraparantag is an antidote targeted to reverse the direct thrombin
and factor Xa inhibitors as well as the indirect inhibitor enoxaparin. OBJECTIVE: To evaluate the role of idarucizumab, a humanized monoclonal antibody
fragment, as a specific reversal agent for the anticoagulant activity of
dabigatran and to review the pharmacology, pharmacokinetic properties, efficacy,
and safety of this agent.
METHODS: A literature search was conducted consisting of a PubMed database using
the MeSH term idarucizumab and the key word dabigatran antidote. Studies
evaluating the pharmacology, pharmacokinetics, safety, and efficacy of
idarucizumab for the reversal of the anticoagulant activity of dabigatran were
included.
RESULTS: Idarucizumab represents a novel treatment option as it is the only
humanized, monoclonal antibody fragment that specifically binds to dabigatran.
Studies evaluating reversal of dabigatran-induced anticoagulation have
demonstrated immediate, complete, and sustained effects with idarucizumab.
Idarucizumab did not overcorrect thrombin generation. Additionally, evaluations
have shown that dabigatran can be safely reinitiated 24 hours after the
administration of idarucizumab. The United States Food and Drug Administration
granted priority review for the biologic license application and accelerated
approval for idarucizumab.
CONCLUSION: Idarucizumab represents an encouraging development in the reversal
of dabigatran. Its novel mechanism of action, pharmacokinetics, tolerability,
and lack of thrombotic events contribute positively to its use in patients who
experience bleeding or for those who require emergent surgery or procedures. BACKGROUND: The approval of the oral direct thrombin inhibitor, dabigatran
etexilate, gave patients an alternative to oral anticoagulation with warfarin.
Like all anticoagulants, the primary adverse event (AE) associated with
dabigatran is bleeding. Until the FDA approval of idarucizumab, there had been
no reversal agent for dabigatran-induced anticoagulation in patients with
life-threatening or uncontrollable bleeding, or those requiring emergent
procedures.
AREAS OF UNCERTAINTY: The primary purpose of this review is to summarize the
safety and efficacy of idarucizumab, a monoclonal antibody fragment, and its use
as a reversal agent for dabigatran.
DATA SOURCES: A literature search was conducted through MEDLINE (1946 to
November week 1 2015) and Embase (1980-2015 week 46) using the search term
idarucizumab. Clinicaltrials.gov was consulted for a comprehensive list of
ongoing and completed studies. Additional studies were identified through
bibliographical citations. Clinical trials in animals and humans published in
English evaluating the safety and efficacy of idarucizumab for reversal of
anticoagulant treatment with dabigatran were included for review.
RESULTS: Idarucizumab has been shown to significantly reverse the anticoagulant
effects of dabigatran in both healthy volunteers and patients requiring a
reversal agent because of either overt bleeding or an emergency surgery or
invasive procedure. The most common AEs were headache, nasopharyngitis, back
pain, skin irritation, hypokalemia, delirium, constipation, pyrexia, and
pneumonia. Deaths reported in idarucizumab studies were attributed to either the
index event or a preexisting comorbidity. Most adverse effects were minor, but
21 serious AEs have been reported in the published data including thrombotic
events.
CONCLUSIONS: Given the increased use of direct oral anticoagulants, such as
dabigatran, a need for specific reversal agents exists. Idarucizumab has been
shown to be safe and effective in the reversal of dabigatran-induced
anticoagulation in patients requiring emergent or urgent surgery or in patients
with severe bleeding. Anticoagulation therapy is indicated for management of various clinical
conditions to prevent adverse events and introduction of direct oral
anticoagulants (DOACs) has ushered in a new era in anticoagulation therapy.
Major advantages of DOACS include fewer drug interactions and that they do not
need periodic monitoring. Several patients who were not on anticoagulation
before due to older age, polypharmacy/drug interaction concerns, and logistics
of periodic monitoring are now on anticoagulation with DOACs. Despite their many
advantages, a challenge while prescribing DOACs is very limited availability of
specific reversal agents and lack of understanding or guidance about the
treatment strategy in case of major life threatening bleeding or need for urgent
surgery. So far only one reversal agent has been approved by the Food and Drug
Administration (FDA), idarucizumab for one of the DOACs i.e., dabigatran.
Several other reversal agents are under final phases of development such as
andexanet alfa and PER977 (ciraparantag) and will help in developing specific
strategies for reversal of these agents. In this article, we review current
strategies to manage bleeding with DOACs and provide guidance to clinicians of
inhibiting LF activity in vitro and in cells, as well as in animal models of
anthrax infection. Venous thromboembolism (VTE) is associated with significant morbidity and
mortality. Factors such as the presence of transient risk factors for VTE, risk
of bleeding, and location of deep vein thrombosis (DVT) determine the duration
of anticoagulation. Extended anticoagulation is offered to patients with
unprovoked pulmonary embolism (PE) or proximal DVT and a low risk of bleeding.
Anticoagulation for 3 months is advised in patients with provoked DVT or PE,
high risk of bleeding, and isolated distal or upper extremity DVT. In patients
with unprovoked PE or proximal DVT and a low risk of bleeding, who want to stop
anticoagulation after 3 months, further risk stratification is necessary.
Clinical scoring system, and thrombophilia testing otherwise not routinely
performed, may be considered to measure risk of annual recurrence in such cases.
Short-term anticoagulation may be considered in subsegmental PE and superficial
vein thrombosis, particularly if patients are at low risk of bleeding and have
persistent risk factors for recurrent VTE. In cases of catheter-associated
thrombosis, the catheter need not be removed routinely, and the patient may be
anticoagulated for 3 months or longer if the catheter is maintained in patients
with cancer. Extensive screening for occult cancer in cases of unprovoked VTE is
not beneficial. New oral anticoagulants such as apixaban, rivaroxaban, or
dabigatran may be preferred to vitamin K antagonists in patients without cancer
or renal failure, more so after the development of reversal agents such as
idarucizumab and andexanet alfa. BACKGROUND AND OBJECTIVES: Idarucizumab is an antibody fragment that
specifically reverses dabigatran-mediated anticoagulation. Safety,
pharmacokinetics and pharmacodynamics of idarucizumab were investigated in
dabigatran-treated, middle-aged, elderly and renally impaired volunteers with
characteristics similar to patients receiving anticoagulant therapy.
METHODS: In this randomized, double-blind, crossover study, 46 subjects (12
middle-aged, 45-64 years; 16 elderly, 65-80 years; and 18 with mild or moderate
renal impairment) received dabigatran etexilate (DE; 220 or 150 mg twice daily)
for 4 days. Idarucizumab doses of 1, 2.5 and 5 g or 2 × 2.5 g 1 h apart, or
placebo, were administered as a rapid (5 min) infusion ~2 h after DE at steady
state.
RESULTS: Dabigatran-prolonged diluted thrombin time, ecarin clotting time and
activated partial thromboplastin time were reversed to baseline immediately
after idarucizumab infusion in all groups. Reversal was sustained with doses
≥2.5 g. Idarucizumab was well tolerated under all conditions. No impact of age
on idarucizumab pharmacokinetics was observed; however, subjects with mild or
moderate renal impairment demonstrated increased exposure (up to 84 %),
decreased clearance and prolonged (by up to 49 %) initial half-life of
idarucizumab compared with healthy middle-aged subjects.
CONCLUSIONS: Impaired renal function was associated with increased exposure and
decreased clearance of idarucizumab. Idarucizumab resulted in immediate,
complete and sustained reversal of dabigatran anticoagulant activity, and was
safe and well tolerated in middle-aged, elderly and renally impaired volunteers.
The results support the clinical use of a 5 g dose of idarucizumab.
CLINICAL TRIAL REGISTRATION: http://www.clinicaltrials.gov . Unique identifier:
NCT01955720. Atrial fibrillation (AF), a common cardiac arrhythmia associated with increased
risk of heart failure, thromboembolic phenomena and death, is a leading cause of
hospitalization of adults. A major complication of AF is an increased risk of
ischemic stroke leading to long-term disability and in severe cases, death.
Historically, Coumadin has been the drug of choice for chronic anticoagulation
and stroke prevention in AF patients however, given the need for constant
monitoring and multiple drug interactions, newer anticoagulants have been
developed. One such drug is dabigatran, with the promise of less frequent
monitoring and decreased bleeding tendencies as compared to Coumadin. The main
disadvantage of dabigatran has been the lack of a reversal agent in case of
severe bleeding or emergent surgical intervention. This was until the recent The
Food and Drug Administration approval of idarucizumab, a potential reversal
agent for dabigatran. In this article, we discuss the evidence addressing
idarucizumab safety, tolerability and its efficacy for reversing effect of
dabigatran. Idarucizumab is a monoclonal antibody fragment specifically targeted to
dabigatran. It has demonstrated prompt and durable reversal of the anticoagulant
effects of dabigatran in animal studies and phase 1 studies of young, elderly,
and renally impaired volunteers. Although elective invasive procedures and most
bleeding complications in dabigatran-treated patients can be managed by
temporarily stopping dabigatran therapy and using supportive measures, there are
rare clinical situations that require urgent reversal of the anticoagulant
effect of dabigatran. The effectiveness and safety of 5 g of intravenous
idarucizumab is being investigated in a prospective, open-label, single-cohort
study in patients with serious bleeding or in those requiring an urgent
procedure. In an interim analysis of the first 90 participants, idarucizumab
rapidly and completely reversed the anticoagulant activity of dabigatran in
88%-98% of participants, and there were no safety concerns, with no deaths or
serious adverse events being attributable to idarucizumab. Supported by these
interim results, idarucizumab has been approved in the United States and the
European Union for use when reversal of the anticoagulant effects of dabigatran
is needed for emergency surgery/urgent procedures or in patients with
life-threatening or uncontrolled bleeding. Clinical use of idarucizumab should
follow the same processes as patient enrollment in this study, which is
projected to be completed in 2016. The outcomes achieved with this specific
reversal agent are likely to be of continued interest to treating physicians. Author information:
(1)At Presbyterian College School of Pharmacy in Clinton, S.C., Aida Rebecca
Bickley is an assistant professor of pharmacy practice and critical care and
Caleb Wallace is a doctor of pharmacy student. The authors have disclosed no
potential conflicts of interest, ficial or otherwise. Idarucizumab is a monoclonal antibody fragment specifically targeted to
dabigatran. It has demonstrated prompt and durable reversal of the anticoagulant
effects of dabigatran in animal studies and phase 1 studies of young, elderly,
and renally impaired volunteers. Although elective invasive procedures and most
bleeding complications in dabigatran-treated patients can be managed by
temporarily stopping dabigatran therapy and using supportive measures, there are
rare clinical situations that require urgent reversal of the anticoagulant
effect of dabigatran. The effectiveness and safety of 5 g of intravenous
idarucizumab is being investigated in a prospective, open-label, single-cohort
study in patients with serious bleeding or in those requiring an urgent
procedure. In an interim analysis of the first 90 participants, idarucizumab
rapidly and completely reversed the anticoagulant activity of dabigatran in
88%-98% of participants, and there were no safety concerns, with no deaths or
serious adverse events being attributable to idarucizumab. Supported by these
interim results, idarucizumab has been approved in the United States and the
European Union for use when reversal of the anticoagulant effects of dabigatran
is needed for emergency surgery/urgent procedures or in patients with
life-threatening or uncontrolled bleeding. Clinical use of idarucizumab should
follow the same processes as patient enrollment in this study, which is
projected to be completed in 2016. The outcomes achieved with this specific
reversal agent are likely to be of continued interest to treating physicians. BACKGROUND: The development of novel oral anticoagulants (NOACs) has
revolutionized oral anticoagulation. Rapid incorporation of NOACs into general
practice has heightened the demand for directed reversal agents. Idarucizumab is
a targeted reversal agent that is approved for the urgent reversal of the
anticoagulant effects of dabigatran. While it is a welcome addition to reversal
strategies of dabigatran, a number of clinical questions exist regarding its
place in therapy.
OBJECTIVE: We describe controversies regarding the use of idarucizumab therapy
in patients with dabigatran-associated bleeding.
DISCUSSION: Although existing clinical studies show a rapid reversal of
coagulation assays, these studies did not describe a corresponding improvement
in mortality or rapid cessation of hemorrhage. It is questionable how heavily
clinicians can rely upon the use of the surrogate endpoints in clinical studies,
such as ecarin clotting time and dilute thrombin time. Another issue is whether
patients exhibiting re-elevation of coagulation assays would benefit from an
additional dose of idarucizumab, because this has not been studied. It is
currently unclear if blood products must be given in addition to idarucizumab
can be used as monotherapy.
CONCLUSIONS: The initial data suggest a definite role for idarucizumab in
treatment of bleeding associated with dabigatran. As more clinical practice
experience is gained with the agent and the remaining data on its use are
released, clinicians can better guide the clinical use of idarucizumab. At
present, there is currently not enough evidence for idarucizumab to be used as
monotherapy. BACKGROUND AND PURPOSE: Non-vitamin K anticoagulants (NOAC) such as dabigatran
have become important therapeutic options for the prevention of stroke. Until
recently, there were only nonspecific agents to reverse their anticoagulant
effects in a case of emergency. Idarucizumab, an antibody fragment targeting
dabigatran, is the first specific antidote for a NOAC to be approved, but
real-world experience is limited.
METHODS: We report two cases of patients on dabigatran with acute intracerebral
hemorrhage who received idarucizumab.
RESULTS: In both cases, idarucizumab promptly reversed the anticoagulant effect
of dabigatran and there was no hematoma expansion in follow-up imaging.
CONCLUSIONS: In addition to clinical and preclinical studies, our cases add to
the experience regarding the safety and efficacy of idarucizumab. They show that
idarucizumab may be an important safety option for patients on dabigatran in
emergency situations. INTRODUCTION: Idarucizumab is a reversal agent for dabigatran etexilate. By
reversing the anticoagulating effect of dabigatran etexilate with idarucizumab
(Praxbind), patients presenting with an acute ischemic stroke can now be
eligible for thrombolysis.
PATIENT: We describe our experience with idarucizumab in a 71-year-old male
patient pretreated with dabigatran etexilate. The patient arrived with a
hemiparesis, central facial palsy, and dysarthria.
METHOD: Dabigatran etexilate was antagonized with idarucizumab, approximately
2.5 hours after the patient's last dose. Immediately after the infusion of
idarucizumab, the patient received thrombolytic therapy.
RESULTS: The hemiparesis and the central facial palsy were fully remitted 3 days
after the onset of symptoms, and the dysarthria was remitted 2 days afterwards.
DISCUSSION: Non-vitamin K oral anticoagulants (NOACs) are widely used for the
prevention of embolic stroke in patients with atrial fibrillation. Dabigatran
etexilate is an oral thrombin inhibitor that can be reversed by idarucizumab.
Idarucizumab, a monoclonal antibody fragment, directly binds dabigatran
etexilate and neutralizes its activity.
CONCLUSION: Reversal of dabigatran etexilate using idarucizumab was safe and
successful with no recombit tissue plasminogen activator interactions. PURPOSE OF REVIEW: We review the current evidence for medical and surgical
treatments of spontaneous intracerebral hemorrhage (ICH).
RECENT FINDINGS: Therapy with hemostatic agents (e.g. factor VIIa and tranexamic
acid) if started early after bleeding onset may reduce hematoma expansion, but
their clinical effectiveness has not been shown. Rapid anticoagulation reversal
with prothrombin concentrates (PCC) plus vitamin K is the first choice in
vitamin K antagonist-related ICH. In ICH related to dabigatran, anticoagulation
can be rapidly reversed with idarucizumab. PCC are recommended for ICH related
to FXa inhibitors, whereas specific reversal agents are not yet approved. While
awaiting ongoing trials studying minimally invasive approaches or
hemicraniectomy, the role of surgery in ICH remains to be defined. Therapies
targeting downstream molecular cascades in order to prevent secondary neuronal
damage are promising, but the complexity and multi-phased nature of ICH
pathophysiology is challenging. Finally, in addition to blood pressure control,
antithrombotic prevention after ICH has to consider the risk of recurrent
bleeding as well as the risk of ischemic events. Treatment of acute ICH remains
challenging, and many promising interventions for acute ICH await further
evidence from trials. |
What is the function of the TMEM132 genes? | The extra-cellular portions of TMEM132 proteins contain five conserved domains including three tandem immunoglobulin domains, and a cohesin domain homologue, the first such domain found in animals. These findings strongly predict a cellular adhesion function for TMEM132 family, connecting the extracellular medium with the intracellular actin cytoskeleton. | |
Does DDX54 play a role in DNA damage response? | Yes. DDX54, a candidate genotoxic stress responsive RNA helicase, regulates transcriptome dynamics during DNA damage response. | The cellular response to genotoxic stress is mediated by a well-characterized
network of DNA surveillance pathways. The contribution of post-transcriptional
gene regulatory networks to the DNA damage response (DDR) has not been
extensively studied. Here, we systematically identified RNA-binding proteins
differentially interacting with polyadenylated transcripts upon exposure of
human breast carcinoma cells to ionizing radiation (IR). Interestingly, more
than 260 proteins, including many nucleolar proteins, showed increased binding
to poly(A)+ RNA in IR-exposed cells. The functional analysis of DDX54, a
candidate genotoxic stress responsive RNA helicase, revealed that this protein
is an immediate-to-early DDR regulator required for the splicing efficacy of its
target IR-induced pre-mRNAs. Upon IR exposure, DDX54 acts by increased
interaction with a well-defined class of pre-mRNAs that harbor introns with weak
acceptor splice sites, as well as by protein-protein contacts within components
of U2 snRNP and spliceosomal B complex, resulting in lower intron retention and
higher processing rates of its target transcripts. Because DDX54 promotes
survival after exposure to IR, its expression and/or mutation rate may impact
DDR-related pathologies. Our work indicates the relevance of many
uncharacterized RBPs potentially involved in the DDR. |
List main clinical features of the POEMS syndrome. | POEMS is an acronym for the main clinical features of the syndrome, namely, Polyneuropathy, Organomegaly, Endocrinopathy, M protein, and Skin abnormalities. Other features include papilledema, extravascular volume overload, sclerotic bone lesions, thrombocytosis, and Castleman disease. It is a multisystemc disorder with a good long-term prognosis. | POEMS syndrome, a rare multisystem disease, is a variant of osteosclerotic
myeloma and is characterized by polyneuropathy, organomegaly, endocrinopathy,
monoclonal proteins, and skin changes. Presented herein is a case of POEMS
syndrome with flushing. The flushing was intermittent, involving the face and
upper third of the trunk, and was associated with hypotension and bronchospasm.
Final diagnosis was made by biopsy examination of an axillary lymph node, which
showed angiofollicular hyperplasia that stained strongly and selectively for
lambda light chains. The patient had most of the typical features of POEMS
syndrome but was unique in that her most striking finding was carcinoid-like
flushing. The flushing improved with steroid therapy, as did some of the other
clinical features of her disease. This case suggests that idiopathic flushing
can be added to the skin changes observed in POEMS syndrome. The POEMS (polyneuropathy, organomegaly, endocrinopathy, monoclonal gammapathy,
and skin changes) syndrome is a rare variant of plasma cell dyscrasia with
multisystemic manifestations. We present 4 cases with arterial symptoms typical
of acute arterial obliteration (AAO) and review 9 similar cases in the
literature. The clinical course of AAO was unusual and particularly severe when
affecting the lower limbs; recurrent events required amputations. As
demonstrated by angiographic and histologic studies, thrombotic and atheromatous
lesions were the main pathologic features of AAO. Atherosclerotic risk factors
were absent or moderate in 3 of our cases, and no cause of thrombosis other than
the POEMS syndrome was found. A high production of cytokines was found in all
cases, with elevated serum levels of interleukin-1 beta (9/9 samples),
interleukin-6 (7/9 samples), and tumor necrosis factor-alpha (6/9 samples). We
suggest that arterial manifestations should be added to the spectrum of
manifestations of the POEMS syndrome. Cytokines may mediate the POEMS
syndrome-associated AAO, as previously proposed for the other systemic
manifestations of this disorder. The POEMS syndrome is a rare multisystemic disorder with polyneuropathy,
organomegaly, endocrinopathy of various forms, production of monoclonal (M)
component, and skin changes. We describe a 46-year-old man who developed ascites
one year after the onset of peripheral neuropathy with accompanying muscle
atrophies and increasing weakness. Extensive evaluation revealed that the
patient had no underlying liver disease, maligcy, infection, or cardiac or
renal disease. The ascites initially responded to high-dose corticosteroid
therapy. The patient had many clinical features of the described POEMS syndrome
including sclerotic bone lesions, a persistent lambda-paraprotein and refractory
ascites. In this case ascites was a main presenting feature. Thus, the POEMS
syndrome must be added to the list of rare causes of refractory ascites. A 48-year-old man with polyneuropathy, organomegaly, endocrinopathy, monoclonal
gammopathy, and skin changes (POEMS) syndrome had bilateral optic disc edema
(ODE), bilateral cystoid macular edema (CME), anasarca, and elevated serum
vascular endothelial growth factor (VEGF). This is the first reported example of
ophthalmoscopic, angiographic, and optical coherence tomographic evidence of the
combination of ODE and CME in this syndrome. This combination of features
suggests that the ODE in this condition may be due to increased vascular
permeability. OBJECTIVE: To describe the first reported case of a patient with POEMS syndrome
(polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy, and skin
changes) in conjunction with the endocrinologic manifestation of
panhypopituitarism due to a large clinically nonfunctioning pituitary adenoma.
METHODS: We present the clinical, laboratory, and radiologic details of the case
and review the relevant updated literature.
RESULTS: A 48-year-old man with hypopituitarism and progressive polyneuropathy
presented to an outside hospital with confusion and diaphoresis. He also had
diffuse lymphadenopathy, monoclonal gammopathy, and skin lesions consistent with
a diagnosis of POEMS syndrome. Cytopathologic study of a lymph node showed
findings consistent with Castleman disease. A large suprasellar mass was found
to be the cause of the hypopituitarism.
CONCLUSION: POEMS syndrome is a rare paraneoplastic condition, commonly
associated with Castleman disease, that manifests with progressive distal
polyneuropathy and a monoclonal plasma cell disorder, often accompanied by
endocrinopathy, organomegaly, skin changes, sclerotic bone lesions, ascites,
erythrocytosis, and thrombocytosis. Our current patient had all 5 classic
features of POEMS syndrome along with some diagnostic elements of Castleman
disease, sclerotic bone lesions, and thrombocytosis. To our knowledge, this is
the first known reported case of a patient whose endocrinologic manifestation of
POEMS syndrome was panhypopituitarism attributable to a large clinically
nonfunctioning pituitary adenoma. POEMS syndrome, also known as Crow-Fukase syndrome, represents a rare
multisystem syndrome characterized by polyneuropathy, organomegaly,
endocrinopathy, M protein, and skin changes. Hypothyroidism is one of the common
endocrine abnormalities which are central features of POEMS syndrome. The
clinical data associated with the measurement of thyroid function and its
clinical significance in POEMS syndrome is still rare. Herein, we report 24
cases with POEMS syndrome which were studied thyroid function and clinical
manifestations and performed an associated analysis between hypothyroidism and
edema/effusions. Of the 24 patients with POEMS syndrome, 17(70.8%) had a
recognized hypothyroidism (including 11 clinical hypothyroidism and 6
subclinical hypothyroidism). Fourteen patients (58.3%) had some form of
extravascular volume overload. In 14 patients with edema/effusions, 12 were
diagnosed as having hypothyroidism. Hypothyroidism may be one of causes of
edema/effusions. After thyroid hormone treatment and chemotherapy, symptoms of
hypothyroidism and edema /effusions were improved greatly. OBJECTIVE: Polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy,
and skin changes (POEMS) syndrome is a rare disorder. This study investigated
the types of ocular signs and symptoms in patients with POEMS and any systemic
factors that may be associated with development of such ocular findings.
DESIGN: Case series from tertiary referral center.
PARTICIPANTS: A total of 33 patients with POEMS syndrome underwent at least 1
ophthalmologic examination and were included in the study.
METHODS: A 10-year retrospective chart review of patients diagnosed with POEMS
syndrome was performed.
MAIN OUTCOME MEASURES: Visual symptoms, visual acuity, presence of optic disc
edema (ODE), and levels of systemic factors (including plasma vascular
endothelial growth factor [VEGF], plasma interlukin-6 [IL-6], and raised
intracranial pressure) and their relationship to ODE.
RESULTS: Five of the patients (15%) reported diplopia, 15 patients (45%) had
blurred vision, and 3 patients (9%) had ocular pain. The most common ocular
finding was bilateral ODE in 17 patients (52%). Of the patients with ODE, 5
(29%) were asymptomatic at the first ocular examination. Among patients with
ODE, there was a significant difference (P = 0.03) between the mean plasma VEGF
level at the time of diagnosis of the ODE compared with when the ODE resolved.
There was no difference in plasma IL-6 levels between people with and without
ODE. Patients with ODE had a higher mean lumbar puncture opening pressure
(276±14 mm H(2)O; normal range, 100-250 mm H(2)0) than patients without ODE,
although the difference was not statistically significant (P = 0.08).
CONCLUSIONS: Optic disc edema is a common finding in patients with POEMS.
Because patients can be asymptomatic, eye examinations should be performed in
all patients with POEMS. There may be an association between elevated VEGF and
intracranial pressure and ODE; further studies are required. We report a case of a 64-year-old man with POEMS (polyneuropathy, organomegaly,
endocrinopathy, monoclonal gammopathy and skin changes) syndrome that had been
previously misdiagnosed as systemic sclerosis. He had typical symptoms of POEMS
syndrome, however, the existence of skin sclerosis, contracture of fingers and
pigmentation were similar to that of systemic sclerosis. Ten patients, including
the patient discussed in this case, visited our department between 1990 and
2011. Among them, five patients had skin sclerosis. Therefore, we compared skin
lesions and clinical/laboratory features of POEMS syndrome and systemic
sclerosis in an attempt to distinguish these disorders. Regarding the cutaneous
and laboratory findings, the existence of hemangioma or hypertrichosis is
indicative of POEMS syndrome. By contrast, the existence of systemic
sclerosis-specific autoantibodies, nail fold bleeding, digital ulcer/digital
pitting scar or telangiectasia is highly suggestive of systemic sclerosis. To
our knowledge, this is the first report to discuss in detail the differentiation
between POEMS syndrome and systemic sclerosis. Polyneuropathy is often an initial manifestation of polyneuropathy,
organomegaly, endocrinopathy, M protein and skin changes (POEMS) syndrome and
therefore this disorder is frequently misdiagnosed as chronic inflammatory
demyelinating polyneuropathy (CIDP). We reviewed electrophysiological data in 20
patients with POEMS syndrome and 36 matched patients with CIDP to compare the
electrophysiological features of POEMS syndrome and CIDP. Compared with CIDP
controls, POEMS patients demonstrated (1) less prolonged distal motor latency
and less reduced motor nerve and sensory nerve conduction velocities, (2)
greater reduction of amplitudes of compound motor action potentials (CMAP) in
distal stimulation, and similar reduction of amplitudes of CMAP in proximal
stimulation, (3) similar reduction of amplitudes of sensory nerve action
potentials (SNAP) in median and ulnar nerves, and a greater reduction of
amplitudes of SNAP in tibial and peroneal nerves, (4) less temporal dispersion,
(5) less frequent conduction block, (6) more frequent neurogenic injury in the
muscles of the upper and lower limbs, and more frequent neurogenic injury in the
muscles of the lower than upper limbs, (7) similar F wave and H reflex
abnormalities, and (8) less frequent skin sympathetic response abnormalities. We
concluded that before development of typical clinical manifestations, POEMS
neuropathy can be distinguished from CIDP by neural electrophysiological
examination. These electrophysiological features can be used for early diagnosis
and initiating correct treatment of POEMS syndrome. BACKGROUND: Polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy,
and skin changes (POEMS) syndrome is an uncommon condition related to a
paraneoplastic syndrome secondary to an underlying plasma cell disorder. Among
the myriad of manifestations of the disease, ocular signs and symptoms are
relatively prevalent, affecting about half of all patients with the disease.
OBJECTIVE: To report the ocular manifestations of POEMS syndrome.
CASE: A 47-year-old lady diagnosed to have POEMS syndrome presented with
painless progressive visual diminution. Her color vision was impaired. There was
bilateral papilloedema.
CONCLUSION: POEMS syndrome should be considered among the differential diagnoses
of all patients with a bilateral papilledema in which no other cause can be
readily elucidated. POEMS syndrome is a rare conglomeration of disorders associated with plasma cell
dyscrasia. The acronym POEMS is derived from main features of the syndrome
namely 'polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy and
skin lesions'. Other clinical features include presence of sclerotic bone
lesions, Castleman's disease, papilledema, pleural effusion, edema, ascites,
erythrocytosis and thrombocytosis. Myeloma is the most common plasma cell
dyscrasia associated with POEMS syndrome. Renal involvement is rare and renal
biopsy is characterized by glomerular involvement with membranoproliferative
glomerulonephritis and endothelial injury. We report a case of a 67-year-old
male who presented with clinical features satisfying the diagnostic criteria of
POEMS syndrome and had rapidly progressive renal failure. Renal biopsy showed
extensive interstitial infiltration by plasma cells and concomitant presence of
classic polyarteritis nodosa. Although association with small-vessel vasculitis
has been reported in patients with POEMS syndrome, to the best of our knowledge,
this is the first report of POEMS syndrome associated with medium-sized vessel
vasculitis. BACKGROUND: Polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy
and skin changes (POEMS) syndrome is a multisystem disorder arising from
underlying plasma cell dyscrasia. Renal impairment and related pathological
changes have been reported, but data on its prevalence, response to therapy and
impact on survival are still lacking.
METHODS: We retrospectively reviewed 299 patients diagnosed with POEMS syndrome
in a tertiary-care university hospital from 2000 until 2014. The estimated
glomerular filtration rate (eGFR) was used to define renal impairment and
response, according to International Myeloma Working Group criteria. We examined
the impact of renal impairment and response on patient survival.
RESULTS: Sixty-seven patients (22.4%) had renal impairment (eGFR < 60
mL/min/1.73 m(2)) at baseline. In a multivariate analysis, ascites was
independently associated with renal impairment [odds ratio (OR) 12.366, P <
0.001]. Renal impairment was reversible in 66.0% of patients receiving therapy
and was associated with a shorter time interval between symptom onset and
treatment (OR 0.059, P = 0.043) and a vascular endothelial growth factor
remission (OR 15.958, P = 0.050) in a multivariate analysis. In terms of
therapy, patients with a renal response more commonly received a novel
agent-based regimen (P = 0.037), which also led to a shorter response time (P =
0.001). With a median follow-up of 27.4 months, inferior survival was observed
in patients with severe renal impairment (eGFR < 30 mL/min/1.73 m(2)), but not
in those with moderate dysfunction (eGFR 30-59 mL/min/1.73 m(2)), compared with
patients without renal impairment. A renal response, if achieved, predicted
improved survival.
CONCLUSIONS: Renal impairment is a common complication of POEMS syndrome, but
can be reversed with effective therapy in most cases. POEMS syndrome is a paraneoplastic manifestation associated with hematopoietic
disorders such as multiple myeloma and Castleman disease. POEMS is an acronym
for the main clinical features of the syndrome, namely, Polyneuropathy,
Organomegaly, Endocrinopathy, M protein, and Skin abnormalities. Glomeruloid
hemangiomas are considered to be a specific clinical marker of POEMS syndrome.
However, while they are not pathognomonic, their presence should raise suspicion
of this syndrome or alert clinicians to its possible future development, as
these lesions can appear years before the onset of the syndrome. We report the
cases of 2 women with plasma cell dyscrasias and sudden onset of lesions with a
vascular appearance and histologic findings consistent with glomeruloid
hemangioma. Recognition of this vascular tumor is important for the early
diagnosis of POEMS syndrome. POEMS syndrome (polyneuropathy, organomegaly, endocrinopathy, monoclonal
gammopathy and skin changes) is a multisystem disorder with a good long-term
prognosis. In its dozens of clinical features, those with independent prognostic
value are still not well characterized. We retrospectively included 362 patients
with newly diagnosed POEMS syndrome at our institute from 2000 to 2015. On the
basis of a randomized sample splitting, we first identified four baseline
clinical variables, including age >50 years (hazards ratio (HR) 4.07, 95%
confidence interval (CI) 1.41-11.76, P=0.009), pulmonary hypertension (HR 3.99,
95% CI 1.44-11.04, P=0.008), pleural effusion (HR 3.81, 95% CI 1.23-11.79,
P=0.02) and estimated glomerular filtration rate <30 ml/min/1.73 m2 (HR 8.25,
95% CI 2.18-31.25, P=0.002), associated with inferior overall survival in the
derivation cohort, with the use of multivariate Cox regression model. These
factors were incorporated together to develop a prognostic nomogram. Concordance
index calculation (0.727, 95% CI 0.601-0.853, P=0.018) and calibration curve
plotting demonstrated its significant predictive and discriminatory capacity in
the validation cohort. This nomogram could be a useful and convenient tool in
clinical practice to evaluate individualized prognosis in patients with newly
diagnosed POEMS syndrome. Pulmonary hypertension is one of the well-known clinical manifestations of
polyneuropathy, organomegaly, endocrinopathy, monoclonal gammopathy, and skin
changes (POEMS) syndrome, occurring in approximately 25-30% of the affected
individuals. However, the histopathologic spectrum of pulmonary hypertension
associated with POEMS syndrome has not been fully documented in the literature.
Herein, we report an autopsy case of POEMS syndrome in a patient whose lung
tissues showed histopathology indistinguishable from that of idiopathic
pulmonary arterial hypertension with abundant plexiform lesions in the small
pulmonary arteries. POEMS syndrome is a rare paraneoplastic syndrome secondary to a plasma cell
dyscrasia. Recognition of a combination of peripheral neuropathy, organomegaly,
endocrinopathy, monoclonal plasmaproliferative disorder, skin changes,
papilledema, extravascular volume overload, sclerotic bone lesions,
thrombocytosis, and Castleman disease is the first step in managing the disease.
Increased blood levels of vascular endothelial growth factor are usually
confirmatory. This rare disorder should not be missed, especially if the patient
has a putative diagnosis of chronic inflammatory polyradiculoneuropathy, a
lambda restricted monoclonal gammopathy, and thrombocytosis, and is not
responding as expected to immunomodulatory therapy commonly used for chronic
inflammatory polyradiculoneuropathy. Treatment of polyneuropathy, organomegaly, endocrinopathy, monoclonal
gammopathy, and skin changes (POEMS) syndrome should be directed at the
underlying plasma cell clone with risk-adapted therapy based on the extent of
the plasma cell disorder. Radiation therapy is effective for patients with a
localized presentation, without bone marrow involvement, and 1 to 3 bone
lesions. Patients with disseminated disease should receive, preferably,
high-dose chemotherapy with peripheral blood transplantation. Low-dose melphalan
and dexamethasone or new agents used in myeloma are also effective. The most
promising agent is lenalidomide, which could be given before high-dose therapy
or radiation to get rapid neurologic responses. Polyneuropathy, organomegaly, endocrinopathy, monoclonal plasma cell disorder,
skin changes (POEMS) syndrome is a rare paraneoplastic disorder. The
polyneuropathy can be the presenting symptom and is typically a painful,
motor-predomit polyradiculoneuropathy often mimicking chronic inflammatory
demyelinating polyradiculoneuropathy. The presence of a lambda monoclonal
protein, elevated vascular endothelial growth factor, systemic features, and
treatment resistance are clues to the diagnosis. Castleman disease (CD) is seen
in a subset of these patients, and when present the neuropathy is similar but
less severe. In contrast, in those patients with purely CD, the neuropathy is
often a mild, painless distal sensory neuropathy. |
Which retinal dystrophy related gene is targeted by the AAV2-hRPE65v2 drug? | AAV2-hRPE65v2, also called voretigene neparvovec, targets the RPE65 gene, whose mutations lead to retinal dystrophy. | Conflict of interest statement: Declaration of interests SR’s institution has
received grants from Spark Therapeutics and he has provided presentations on
behalf of Spark Therapeutics. DC and AD have received grants from Spark
Therapeutics. JB has received grants from the Foundation Fighting Blindness, the
National Institutes of Health, and Spark Therapeutics; non-ficial support
from the Center for Advanced Retinal and Ocular Therapeutics, University of
Pennsylvania, and the FM Kirby Foundation; has a provisional patent pending and
a US patent licensed to Spark Therapeutics for which she has waived ficial
interest; and is a coauthor on a copyrighted visual function questionnaire used
in the present study. JAW is an employee of and has equity/options in Spark
Therapeutics; has received grants from the National Institutes of Health
(specifically for Clinical and Translational Research Centre services); and has
a patent pending pertaining to the primary endpoint measure licensed to Spark
Therapeutics. DCC is an employee of Spark Therapeutics and has a patent pending
pertaining to the primary endpoint measure licensed to Spark Therapeutics. Z-FY,
AT, and JWi are employees of Statistics Collaborative, which provides
statistical and regulatory consulting to Spark Therapeutics. JP is an employee
of Westat, which was contracted by Spark Therapeutics. SM is a clinical
coordinator of a study sponsored by Spark Therapeutics, receives salary support
from the Center for Cellular and Molecular Therapeutics (CCMT) at The Children’s
Hospital of Philadelphia, and has a patent pending licensed to Spark
Therapeutics. KAM is a clinical coordinator of a study sponsored by Spark
Therapeutics and received salary support from CCMT at Children’s Hospital of
Philadelphia. JWa, TLK, and MD have received salary support from Spark
Therapeutics and grants from the Carver Center for Macular Degeneration and
Children’s Hospital of Philadelphia; TLK has received travel support from Spark
Therapeutics. JAH has received consulting fees from Merck and Kalvista, grants
from ThromboGenics, and serves on boards for Janssen and Celgene. ES’s
institution has received grants from Children’s Hospital of Philadelphia and
Spark Therapeutics. EHS has received grants from Oxford Biomedica. KW is an
employee of and has equity/options with Spark Therapeutics and has received
grants from the National Institutes of Health. FS has received grants from
Regione Campania and serves on boards for Sanofi, Dompe Farmaceutici, and Spark
Therapeutics. JFW has a patent and a patent pending, both licensed to Spark
Therapeutics. KAH is an employee of Spark Therapeutics and has a patent pending
pertaining to the primary endpoint measure. AMM has received grants from Spark
Therapeutics, the National Institutes of Health, and the Foundation Fighting
Blindness; non-ficial support from the Center for Advanced Retinal and Ocular
Therapeutics, University of Pennsylvania, and the FM Kirby Foundation; and has a
provisional patent pending and a US patent licensed to Spark Therapeutics for
which he has waived ficial interest. OE, HR, LR, LAG, FPH, LD, XZ, VBM, WP,
MW, CJ, DG, and BPL declare no competing interests. |
What is the function of yeast TERRA RNAs? | The ends of linear eukaryotic chromosomes are transcribed into different species of non-coding transcripts (the telomeric transcriptome), including TERRA (telomeric repeat-containing RNA) molecules. TERRA are part of the DNA damage response triggered by dysfunctional telomeres. In addition to its role as a template-encoding telomeric DNA synthesis, telomerase RNA has been shown to function as a flexible scaffold for protein subunits of the RNP holoenzyme. | Telomere-repeat-encoding RNA (referred to as TERRA) has been identified as a
potential component of yeast and mammalian telomeres. We show here that TERRA
RNA interacts with several telomere-associated proteins, including telomere
repeat factors 1 (TRF1) and 2 (TRF2), subunits of the origin recognition complex
(ORC), heterochromatin protein 1 (HP1), histone H3 trimethyl K9 (H3 K9me3), and
members of the DNA-damage-sensing pathway. siRNA depletion of TERRA caused an
increase in telomere dysfunction-induced foci, aberrations in metaphase
telomeres, and a loss of histone H3 K9me3 and ORC at telomere repeat DNA.
Previous studies found that TRF2 amino-terminal GAR domain recruited ORC to
telomeres. We now show that TERRA RNA can interact directly with the TRF2 GAR
and ORC1 to form a stable ternary complex. We conclude that TERRA facilitates
TRF2 interaction with ORC and plays a central role in telomere structural
maintece and heterochromatin formation. Telomeric repeat-containing RNA (TERRA) has been implicated in the control of
heterochromatin and telomerase. We demonstrate that yeast TERRA is regulated by
telomere-binding proteins in a chromosome-end-specific manner that is dependent
on subtelomeric repetitive DNA elements. At telomeres that contain only
X-elements, the Rap1 carboxy-terminal domain recruits the Sir2/3/4 and Rif1/2
complexes to repress transcription in addition to promoting
Rat1-nuclease-dependent TERRA degradation. At telomeres that contain Y'
elements, however, Rap1 represses TERRA through recruitment of Rif1 and Rif2.
Our work emphasizes the importance of subtelomeric DNA in the control of
telomeric protein composition and telomere transcription. The ends of linear eukaryotic chromosomes are transcribed into different species
of non-coding transcripts (the telomeric transcriptome), including TERRA
(telomeric repeat-containing RNA) molecules; however, the functions associated
with the telomeric transcriptome remain elusive. Experimental evidence
accumulated during the past few years indicates that the transcriptional
activity of telomeres is changed in cells in which the integrity of the
telomeres or the heterochromatic state of chromosome ends is altered. On the
contrary transcription of a telomere appears not to be influenced by its length.
In this paper we briefly review the current state of knowledge on the
composition, biogenesis, and regulation of the telomeric transcriptome from
yeasts to humans. We also suggest a model in which TERRA is part of the DNA
damage response triggered by dysfunctional telomeres and discuss the potential
involvement of telomere transcription in the development of human pathologies. In most eukaryotes, the ribonucleoprotein complex telomerase is responsible for
maintaining telomere length. In recent years, single-cell microscopy techniques
such as fluorescent in situ hybridization and live-cell imaging have been
developed to image the RNA subunit of the telomerase holoenzyme. These
techniques are now becoming important tools for the study of telomerase
biogenesis, its association with telomeres and its regulation. Here, we present
detailed protocols for live-cell imaging of the Saccharomyces cerevisiae
telomerase RNA subunit, called TLC1, and also of the non-coding telomeric
repeat-containing RNA TERRA. We describe the approach used for genomic
integration of MS2 stem-loops in these transcripts, and provide information for
optimal live-cell imaging of these non-coding RNAs. While the mechanisms of telomere maintece has been investigated in dividing
cells, little is known about the stability of telomeres in quiescent cells and
how dysfunctional telomeres are processed in non-proliferating cells. Here we
examine the stability of telomeres in quiescent cells using fission yeast. While
wild type telomeres are stable in quiescence, we observe that eroded telomeres
were highly rearranged during quiescence in telomerase minus cells. These
rearrangements depend on homologous recombination (HR) and correspond to
duplications of subtelomeric regions. HR is initiated at newly identified
subtelomeric homologous repeated sequences (HRS). We further show that TERRA
(Telomeric Repeat-containing RNA) is increased in post-mitotic cells with short
telomeres and correlates with telomere rearrangements. Finally, we demonstrate
that rearranged telomeres prevent cells to exit properly from quiescence. Taken
together, we describe in fission yeast a mode of telomere repair mechanism
specific to post-mitotic cells that is likely promoted by transcription. |
What are the 4 cardinal signs of inflammation according to Celsus? | redness or rubor , heat or calor, swelling or tumor, and pain or dolor | John Hunter's A Treatise on the Blood, Inflammation and Gunshot Wounds was
published in 1794. Throughout the nineteenth century this was considered the
most important study of inflammation and has been widely quoted since. After a
section on the nature of blood and the circulatory system, in which he describes
the vascular supply in detail, he passes on to an extensive survey of
inflammation. This is based mainly on his wide clinical experience, including
that as a military surgeon. He, however, supplements this with a number of
experiments, some of which are classic. He bases his observations on the four
cardinal signs of Celsus (redness, heat, swelling and pain). Inflammation is
then divided into three main groups: adhesive, suppurative and ulcerative. He
discusses the nature of pus and the formation and treatment of abscesses. He
describes his experiments on the transplantation of tissues under the general
heading of adhesive inflammation. This, he states, underlies the union of wounds
and thus the union of tissues after transplantation. Although unaware of the
role of infecting organisms as a cause of inflammation, he makes observations on
inflammation in smallpox, venereal infections and tuberculosis. He relates these
to his observations on inflammatory aspects of wound healing. Lister was
particularly influenced by Hunter's observations in the development of
antisepsis. As well as the local effect of inflammation, Hunter was concerned
with the constitutional effects such as fever. 1. Knowledge of slit lamp illumination techniques along with solid concepts of
the disease processes can allow an understanding of the visible morphologic
structures seen in the disease process during examination with the
biomicroscope. 2. Inflammation can be defined as the interaction between a
stimulus and a host, frequently resulting in some degree of structural change
within the host. 3. The cellular responses include a predictably sequential
order of events. The four cardinal signs of inflammation are redness, swelling,
heat, and pain, phenomena explained at the cellular level. The concept of the four cardinal signs of acute inflammation comes from
antiquity as rubor et tumor cum calore et dolore, (redness and swelling with
heat and pain) extended later by functio laesa (loss of function). The
contemporary understanding of this process we owe to 19th-century milestone
discoveries by Rudolph Virchow, Julius Cohnheim and Elie Metchnikoff. In the
20th century, the development of potent technological tools allowed the rapid
expansion of knowledge of the cells and mediators of inflammatory processes, as
well as the molecular mechanisms of their interactions. It turned out that some
mediators of inflammation have both local and distant targets, among them the
liver (responding by the production of several acute phase reactants) and
neurohormonal centers. In the last decades it has become clear that the immune
system shares mediators and their receptors with the neurohormonal system of the
body; thus, they form a common homeostatic entity. Such an integrative view,
introduced by J. Edwin Blalock, when combined with Hans Selye's concept of
stress, led to the contemporary understanding of sickness behavior, defined by
Robert Dantzer as a highly organized strategy of the organism to fight
infections and to respond to other environmental stressors. According to semiotics, which may be defined as the doctrine of the essential
nature and fundamental varieties of signs, objects, and interpretants, pain is
considered to be a sign (significant) with very different meanings
(significance) either as a naturalistic symptom (of disease) or as a symbol used
in a metaphorical context. When following this methodological perspective it is
possible to interpret medical as well as poetic writings on equal terms. In
Graeco-Roman medical texts pain was mostly understood as a result and an
indicator of disease, but nonetheless as a symptom which seemed to be actively
produced by the affected body. Especially in the Corpus Hippocraticum dating
from the 5th and 4th century B. C. this materialistic and at the same time
psychosomatic attitude can be noticed. Aristotle (4th century B. C.), Celsus
(1st century A. D.), and the famous experimental physiologist Galen (2nd century
A. D.) agreed that pain was a sign of evil which should be fought without
exception. It was Galen who added the disturbance of function (functio laesa) as
the fifth cardinal sign of inflammation to the four well-known cardinal signs of
Celsus (rubor, calor, tumor, dolor). He also coined the term [see text] to
characterize an attack of migraine. In algotherapy, Galen used a complex
pharmacological system which was based upon the four cardinal qualities of
humoral pathology. On the other hand, pain was designed as a multi-dimensional
symbol by the famous Graeco-Roman epic poets. In Homer's Odyssey (8th century B.
C.), pain appears transformed into the shape of a scar which is visible and
palpable on the hero's leg like an identification tag, whereas in Virgil's
Aeneids (1st century B. C.) pain symbolizes weakness and defencelessness which
can only be alleviated by the goddess Venus. Inflammation is a protective response essential for maintaining human health and
for fighting disease. As an active innate immune reaction to challenge,
inflammation gives rise to clinical cardinal signs: rubor, calor, dolor, tumor
and functio laesa. Termination of acute inflammation was previously recognized
as a passive process; a natural decay of pro-inflammatory signals. We now
understand that the natural resolution of inflammation involves well-integrated,
active, biochemical programs that return tissues to homeostasis. This review
focuses on recent advances in the understanding of the role of endogenous lipid
mediators that modulate cellular fate and inflammation. Biosynthesis of
eicosanoids and other lipids in exudates coincides with changes in the types of
inflammatory cells. Resolution of inflammation is initiated by an active class
switch in lipid mediators, such as classic prostaglandins and leukotrienes, to
the production of proresolution mediators. Endogenous pro-resolving lipid
mediators, including arachidonic acid-derived lipoxins, aspirin-triggered
lipoxins, ω3-eicosapentaenoic acid-derived resolvins of the E-series,
docosahexaenoic acid-derived resolvins of the D-series, protectins and maresins,
are biosynthesized during the resolution phase of acute inflammation. Depending
on the type of injury and the type of tissue, the initial cells that respond are
polymorphonuclear leukocytes, monocytes/macrophages, epithelial cells or
endothelial cells. The selective interaction of specific lipid mediators with G
protein-coupled receptors expressed on innate immune cells (e.g. G
protein-coupled receptor 32, lipoxin A4 receptor/formyl peptide receptor2,
chemokine-like receptor 1, leukotriene B4 receptor type 1 and cabannoid receptor
2) induces cessation of leukocyte infiltration; vascular permeability/edema
returns to normal with polymorphonuclear neutrophil death (mostly via
apoptosis), the nonphlogistic infiltration of monocyte/macrophages and the
removal (by macrophages) of apoptotic polymorphonuclear neutrophils, foreign
agents (bacteria) and necrotic debris from the site. While an acute inflammatory
response that is resolved in a timely manner prevents tissue injury, inadequate
resolution and failure to return tissue to homeostasis results in
neutrophil-mediated destruction and chronic inflammation. A better understanding
of the complex mechanisms of lipid agonist mediators, cell targets and actions
allows us to exploit and develop novel therapeutic strategies to treat human
inflammatory diseases, including periodontal diseases. PURPOSE: The aim of this study was to examine the literature with respect to
inflammation of the ocular surface and the presence of inflammatory mediators in
the tear film during contact lens wear.
METHODS: The literature on contact lens discomfort that relates to signs of
inflammation was searched. Reference was paid to the cardinal signs of
inflammation (pain, heat, redness, and swelling) as well as the appearance of
inflammatory mediators in the tear film during contact lens wear.
RESULTS: Contact lens wear does induce discomfort, which is a mild form of pain,
and wearing of lenses can induce increases in limbal and conjunctival redness.
However, there is little evidence for a direct relationship between limbal or
conjunctival redness and contact lens discomfort. Contact lenses may increase
the temperature of the ocular surface by a small amount (≤2.1°C). Corneal or
conjunctival swelling has not been associated with contact lens discomfort.
Complement or kinin breakdown products or histamine in tears are not associated
with contact lens discomfort. Cytokines are generally not related to discomfort,
although nerve growth factor levels were increased in tears of symptomatics in
one report. The presence of degraded lipids, leukotriene B4 and peroxidation
products, and the enzyme secretory phospholipase A2 in tears has been associated
with contact lens discomfort, and this area is worthy of further research.
CONCLUSIONS: There have been only a few studies that have investigated a role
for inflammation in contact lens-induced discomfort, and the strongest
associations have been found with lipid degradation processes. Inflammation is the body's response to injury or infection. As early as 2000
years ago, the Roman encyclopaedist Aulus Cornelius Celsus recognised four
cardinal signs of this response-redness, heat, swelling and pain; a fifth sign
is loss of function.[...]. |
Do bacteria from the genus Morexella cause respiratory infections? | Bacteria from the genus Morexella can cause respiratory infections | The efficacy and safety of oral ofloxacin, 400 mg once daily, for the treatment
of patients with lower respiratory tract infections were studied. The most
common species recovered from the sputum specimens of these patients were
Haemophilus influenzae, followed by Streptococcus pneumoniae (S. pneumoniae),
Staphylococcus aureus (S. aureus), Gram positive cocci unidentified, Pseudomonas
aeruginosa (P. aeruginosa), Morexella catarrhalis, Streptococcus epidermidis,
and another Haemophilus species in this order. All these bacteria were
susceptible to ofloxacin except for one strain of methicillin resistant S.
aureus. A satisfactory clinical outcome was achieved in 34 of 40 patients (85%).
It is concluded that ofloxacin, 400 mg once daily, is useful for patients with
respiratory tract infections. ETHNOPHARMACOLOGICAL RELEVANCE: The antibacterial activities of 18 plants from
10 different families were investigated for their antimicrobial efficacy, based
on the traditional uses of these species by Bakola pygmies living in Central
Africa, especially along the Ngoyang area in Cameroon for the treatment of
respiratory and tuberculosis-related symptoms. The aim of the study is to test
the antimicrobial efficacy of these plants against some pathogens associated
with respiratory disease and to determine if there is any validation for the
traditional use against Mycobacterium species.
MATERIALS AND METHODS: Medium polar extracts were prepared in MeOH/DCM (1:1,
v/v) from the plant parts of each species used traditionally and were assayed
against pathogens associated with respiratory tract ailments [Staphylococcus
aureus (ATCC 25923), Klebsiella pneumoniae (ATCC 13883) and Morexella
cattarhalis (ATCC 14468)] using the minimum inhibitory concentration (MIC)
method. Two additional faster growing Mycobacterium strains [Mycobacterium
smegmatis (ATCC 23246) and Mycobacterium aurum (NCTC 10437)] were included in
the assay as predictive test organisms for the more pathogenic strain
Mycobacterium tuberculosis.
RESULTS: Some plant species, such as Alchornea floribunda, Musanga cecropioides
(both leaves and stem bark), Tetracera potatoria and Xylopia aethiopica (stem
bark), were effective in inhibiting Morexella cattarhalis, having MIC values
between 65 and 250 μg/mL. Some noteworthy antimycobacterial inhibition (MIC≤200
μg/mL and as low as MIC 6.5 µg/mL) for 54% of the extracts were observed.
CONCLUSION: While moderate activity was shown for pathogens causing respiratory
tract infections, these plant species seems to be selectively targeting
Mycobacteria spp. suggesting that the traditional use for treating tuberculosis
related symptoms may be indeed be accurate. |
Which two genes are implicated in Juvenile polyposis syndrome? | Juvenile polyposis syndrome (JPS) is a rare autosomal dominant disorder predisposing to gastrointestinal hamartomatous polyps and cancer with a pathogenic SMAD4 or BMPR1A germline mutation being identified in about 40-50% of patients. | BACKGROUND: In patients with juvenile polyposis syndrome (JPS) the frequency of
large genomic deletions in the SMAD4 and BMPR1A genes was unknown.
METHODS: Mutation and phenotype analysis was used in 80 unrelated patients of
whom 65 met the clinical criteria for JPS (typical JPS) and 15 were suspected to
have JPS.
RESULTS: By direct sequencing of the two genes, point mutations were identified
in 30 patients (46% of typical JPS). Using MLPA, large genomic deletions were
found in 14% of all patients with typical JPS (six deletions in SMAD4 and three
deletions in BMPR1A). Mutation analysis of the PTEN gene in the remaining 41
mutation negative cases uncovered a point mutation in two patients (5%). SMAD4
mutation carriers had a significantly higher frequency of gastric polyposis
(73%) than did patients with BMPR1A mutations (8%) (p<0.001); all seven cases of
gastric cancer occurred in families with SMAD4 mutations. SMAD4 mutation
carriers with gastric polyps were significantly older at gastroscopy than those
without (p<0.001). In 22% of the 23 unrelated SMAD4 mutation carriers,
hereditary hemorrhagic telangiectasia (HHT) was also diagnosed clinically. The
documented histologic findings encompassed a wide distribution of different
polyp types, comparable with that described in hereditary mixed polyposis
syndromes (HMPS).
CONCLUSIONS: Screening for large deletions raised the mutation detection rate to
60% in the 65 patients with typical JPS. A strong genotype-phenotype correlation
for gastric polyposis, gastric cancer, and HHT was identified, which should have
implications for counselling and surveillance. Histopathological results in
hamartomatous polyposis syndromes must be critically interpreted. Juvenile polyposis syndrome is a rare autosomal domit syndrome characterized
by multiple distinct juvenile polyps in the gastrointestinal tract and an
increased risk of colorectal cancer. The cumulative life-time risk of colorectal
cancer is 39% and the relative risk is 34. Juvenile polyps have a distinctive
histology characterized by an abundance of edematous lamina propria with
inflammatory cells and cystically dilated glands lined by cuboidal to columnar
epithelium with reactive changes. Clinically, juvenile polyposis syndrome is
defined by the presence of 5 or more juvenile polyps in the colorectum, juvenile
polyps throughout the gastrointestinal tract or any number of juvenile polyps
and a positive family history of juvenile polyposis. In about 50%-60% of
patients diagnosed with juvenile polyposis syndrome a germline mutation in the
SMAD4 or BMPR1A gene is found. Both genes play a role in the BMP/TGF-beta
signalling pathway. It has been suggested that cancer in juvenile polyposis may
develop through the so-called "landscaper mechanism" where an abnormal stromal
environment leads to neoplastic transformation of the adjacent epithelium and in
the end invasive carcinoma. Recognition of this rare disorder is important for
patients and their families with regard to treatment, follow-up and screening of
at risk individuals. Each clinician confronted with the diagnosis of a juvenile
polyp should therefore consider the possibility of juvenile polyposis syndrome.
In addition, juvenile polyposis syndrome provides a unique model to study
colorectal cancer pathogenesis in general and gives insight in the molecular
genetic basis of cancer. This review discusses clinical manifestations,
genetics, pathogenesis and management of juvenile polyposis syndrome. BACKGROUND: Juvenile polyposis syndrome is a domit GI polyposis syndrome
defined by ≥ 5 GI juvenile polyps or ≥ 1 juvenile polyps with a family history
of juvenile polyposis. Mutations in BMPR1A or SMAD4 are found in 50% of
individuals. Hereditary hemorrhagic telangiectasia is a domit disorder
characterized by epistaxis, visceral arteriovenous malformations, and
telangiectasias. Hereditary hemorrhagic telangiectasia is diagnosed when ≥ 3
criteria including clinical manifestations or a family history, are present. A
juvenile polyposis-hereditary hemorrhagic telangiectasia overlap syndrome has
previously been reported in 22% of patients with juvenile polyposis due to a
SMAD4 mutation.
OBJECTIVE: Our objective was to determine the prevalence and clinical
manifestations of hereditary hemorrhagic telangiectasia by Curacao criteria in
our juvenile polyposis SMAD4 patients.
DESIGN, PATIENTS, AND SETTING: This was a cohort study of juvenile polyposis
patients in our inherited colon cancer registries. Hereditary hemorrhagic
telangiectasia manifestations were obtained from medical records, patient
contact, and/or prospective hereditary hemorrhagic telangiectasia screening. The
Curacao criteria was used for diagnosis of hereditary hemorrhagic telangiectasia
(≥ 3 criteria diagnostic; 2 criteria suspect of).
MAIN OUTCOME MEASURES: Prevalence and clinical manifestations of hereditary
hemorrhagic telangiectasia in juvenile polyposis SMAD4 patients.
RESULTS: Forty-one juvenile polyposis families were identified. Genetic testing
was available for individuals within 18 families. SMAD4 mutations were found in
21 relatives in 9 families. Eighty-one percent of SMAD4 patients had hereditary
hemorrhagic telangiectasia and 14% were suspected of having hereditary
hemorrhagic telangiectasia. Epistaxis and asthma are the most common symptoms in
our overlap patients. Symptomatic and subclinical arteriovenous malformations
were noted near universally.
LIMITATIONS: There was a single, tertiary referral center.
CONCLUSIONS: Nearly all juvenile polyposis SMAD4 patients have the overlap
syndrome. The clinical implications and need for hereditary hemorrhagic
telangiectasia screening are important factors for genetic testing in juvenile
polyposis. Health care providers must be cognizant of the juvenile
polyposis-hereditary hemorrhagic telangiectasia overlap syndrome and the
implications for management of these patients. Juvenile polyposis syndrome (JPS) is an autosomal domit predisposition to the
occurrence of hamartomatous polyps in the gastrointestinal tract. Diagnosis of
JPS is based on the occurrence of numerous colon and rectum polyps or any number
of polyps with family history and, in the case of juvenile polyps, their
occurrence also outside the large intestine. The JPS is caused by mutations in
SMAD4 and BMPR1A. Products of the SMAD4 gene are involved in signal transduction
in the transforming growth factor β pathway and BMPR1A protein is a receptor
belonging to the family of transmembrane serine/threonine kinases. Both proteins
are responsible for processes determining appropriate development of colonic
mucosa. The JPS belongs to the group of hamartomatous polyposes. The
hamartomatous polyposis syndromes constitute a group of diseases in which
manifestations differ slightly and only molecular diagnostics gives the
possibility of verifying the clinical diagnosis. We describe a patient with a severe juvenile polyposis phenotype, due to a de
novo deletion of chromosome 10q22.3-q24.1. He was initially diagnosed with
Juvenile polyposis syndrome (JPS) at age four after presenting with hematochezia
due to multiple colonic juvenile polyps. He then re-presented at 23 years with
recurrent hematochezia from juvenile polyps in his ileoanal pouch. He is one of
the earliest reported cases of JPS associated with a large deletion of
chromosome 10. Since his initial diagnosis of JPS further studies have confirmed
an association between JPS and mutations in BMPR1A in chromosome band 10q23.2,
which is in close proximity to PTEN. Mutations in PTEN cause Cowden syndrome
(CS) and other PTEN hamartoma tumor syndromes. Due to the chromosome 10 deletion
involving contiguous portions of BMPR1A and PTEN in our patient, he may be at
risk for CS associated cancers and features, in addition to the polyps
associated with JPS. This case presents new challenges in developing appropriate
surveillance algorithms to account for the risks associated with each syndrome
and highlights the importance of longitudinal follow-up and transitional care
between pediatric and adult gastroenterology for patients with hereditary
polyposis syndromes. Author information:
(1)Genomic Medicine Institute, Cleveland Clinic, Cleveland, Ohio; Lerner
Research Institute, Cleveland Clinic, Cleveland, Ohio; Division of Medical
Oncology, National Cancer Centre, Singapore; Oncology Academic Clinical Program,
Duke-NUS Graduate Medical School, Singapore.
(2)Genomic Medicine Institute, Cleveland Clinic, Cleveland, Ohio; Lerner
Research Institute, Cleveland Clinic, Cleveland, Ohio.
(3)Institute of Molecular and Cell Biology, A∗STAR, Singapore.
(4)Genomic Medicine Institute, Cleveland Clinic, Cleveland, Ohio; Lerner
Research Institute, Cleveland Clinic, Cleveland, Ohio; Taussig Cancer Institute,
Cleveland Clinic, Cleveland, Ohio.
(5)Centre for Computational Biology, Duke-NUS Graduate Medical School,
Singapore.
(6)Department of Pathology, Singapore General Hospital, Singapore.
(7)Genomic Medicine Institute, Cleveland Clinic, Cleveland, Ohio; Lerner
Research Institute, Cleveland Clinic, Cleveland, Ohio; Taussig Cancer Institute,
Cleveland Clinic, Cleveland, Ohio; CASE Comprehensive Cancer Center, Case
Western Reserve University, Cleveland, Ohio; Department of Genetics and Genome
Sciences, Case Western Reserve University, Cleveland, Ohio. Electronic address:
[email protected]. Juvenile polyposis syndrome (JPS) is a rare autosomal domit disorder
characterized by the development of multiple hamartomatous polyps in the
gastrointestinal tract. Polyps are most common in the colorectum (98% of
patients) and the stomach (14%). Causative mutations for JPS have been
identified in two genes to date, SMAD4 and BMPR1A. SMAD4 mutations are
associated with a higher incidence of gastric polyposis. In this case report, we
describe two patients with massive gastric polyposis associated with a SMAD4
mutation. Both presented with anaemia and both had colonic polyps. Initial
endoscopic findings revealed giant rugal folds suggestive of Ménétrier disease.
However, as other possible gastropathies could not be differentiated on the
basis of histology, a definitive diagnosis of JPS required additional mutation
analysis. In patients with polyposis predomit in or limited to the stomach,
establishing a diagnosis based solely on the pathological features of polyps can
be challenging due to difficulties in differentiating JPS from other
hypertrophic gastropathies. Mutation analysis should be considered early in the
diagnostic process in cases of suspected juvenile polyposis, thus facilitating
rapid diagnosis and adequate follow-up. Juvenile polyposis syndrome (JPS) is a rare autosomal domit disorder
predisposing to gastrointestinal hamartomatous polyps and cancer with a
pathogenic SMAD4 or BMPR1A germline mutation (1st-hit) being identified in about
40-50% of patients. Little is known, however, about the occurrence and nature of
somatic alterations (2nd-hit) in SMAD4-/BMPR1A-related juvenile polyps. In this
study, we screened 25 polyps from three patients carrying either a pathogenic
SMAD4 (c.1244-1247delACAG) or BMPR1A (c.583C>T; p.Gln195*) germline mutation for
somatic alterations. The SMAD4-related polyps were also analyzed for SMAD4
protein expression by immunohistochemistry. Despite comprehensive screening for
loss of heterozygosity (LOH), mutations in the coding sequence, chromosomal
rearrangements, and promoter methylation, no somatic alterations could be
identified in 14 SMAD4-related polyps. SMAD4 protein expression, however, was
lost in 8 (57%) of 14 juvenile polyps with 6 showing concomitant loss in both,
the epithelial and stromal, compartments. In the BMPR1A-related polyps, five out
of nine (56%) displayed LOH. Further analysis of selected polyps revealed that
LOH was gene copy number neutral and had occurred in the epithelial compartment.
The heterogeneity of genetic mutations and protein expression levels indicates
that different modes of gene inactivation can be operational in SMAD4- and
BMPR1A-related polyp formation. Our observation, that about half of
BMPR1A-related polyps displayed LOH, predomitly in the epithelial
compartment, is compatible with BMPR1A acting as a tumour suppressor gene.
Still, it remains to be determined whether juvenile polyp development generally
requires loss of BMPR1A expression or, as observed in some SMAD4-related polyps,
can occur despite normal protein expression. |
Do chromatin features predict genes associated with eQTLs? | Yes, genomic proximity plus five TF and chromatin features are able to predict>90% of target genes within 1 megabase of eQTLs | Cell type-specific gene expression in humans involves complex interactions
between regulatory factors and DNA at enhancers and promoters. Mapping studies
for expression quantitative trait loci (eQTLs), transcription factors (TFs) and
chromatin markers have become widely used tools for identifying gene regulatory
elements, but prediction of target genes remains a major challenge. Here, we
integrate genome-wide data on TF-binding sites, chromatin markers and functional
annotations to predict genes associated with human eQTLs. Using the random
forest classifier, we found that genomic proximity plus five TF and chromatin
features are able to predict >90% of target genes within 1 megabase of eQTLs.
Despite being regularly used to map target genes, proximity is not a good
indicator of eQTL targets for genes 150 kilobases away, but insulators, TF
co-occurrence, open chromatin and functional similarities between TFs and genes
are better indicators. Using all six features in the classifier achieved an area
under the specificity and sensitivity curve of 0.91, much better compared with
at most 0.75 for using any single feature. We hope this study will not only
provide validation of eQTL-mapping studies, but also provide insight into the
molecular mechanisms explaining how genetic variation can influence gene
expression. |
List the 6 genes associated with the autosomal recessive form of Osteogenesis imperfecta | There are at least 6 genes associated with osteogenesis imperfecta, Sp7/Osx, FK506-binding protein, Hsp47/SERPINH1, WNT1, CRTAP, P3H1, and PPIB, LEPRE1,PLOD2, TMEM38B | Classic osteogenesis imperfecta, an autosomal domit disorder associated with
osteoporosis and bone fragility, is caused by mutations in the genes for type I
collagen. A recessive form of the disorder has long been suspected. Since the
loss of cartilage-associated protein (CRTAP), which is required for
post-translational prolyl 3-hydroxylation of collagen, causes severe
osteoporosis in mice, we investigated whether CRTAP deficiency is associated
with recessive osteogenesis imperfecta. Three of 10 children with lethal or
severe osteogenesis imperfecta, who did not have a primary collagen defect yet
had excess post-translational modification of collagen, were found to have a
recessive condition resulting in CRTAP deficiency, suggesting that prolyl
3-hydroxylation of type I collagen is important for bone formation. Autosomal domit osteogenesis imperfecta (OI) is caused by mutations in the
genes (COL1A1 or COL1A2) encoding the chains of type I collagen. Recently,
dysregulation of hydroxylation of a single proline residue at position 986 of
both the triple-helical domains of type I collagen alpha1(I) and type II
collagen alpha1(II) chains has been implicated in the pathogenesis of recessive
forms of OI. Two proteins, cartilage-associated protein (CRTAP) and
prolyl-3-hydroxylase-1 (P3H1, encoded by the LEPRE1 gene) form a complex that
performs the hydroxylation and brings the prolyl cis-trans isomerase
cyclophilin-B (CYPB) to the unfolded collagen. In our screen of 78 subjects
diagnosed with OI type II or III, we identified three probands with mutations in
CRTAP and 16 with mutations in LEPRE1. The latter group includes a mutation in
patients from the Irish Traveller population, a genetically isolated community
with increased incidence of OI. The clinical features resulting from CRTAP or
LEPRE1 loss of function mutations were difficult to distinguish at birth.
Infants in both groups had multiple fractures, decreased bone modeling
(affecting especially the femurs), and extremely low bone mineral density.
Interestingly, "popcorn" epiphyses may reflect underlying cartilaginous and bone
dysplasia in this form of OI. These results expand the range of CRTAP/LEPRE1
mutations that result in recessive OI and emphasize the importance of
distinguishing recurrence of severe OI of recessive inheritance from those that
result from parental germline mosaicism for COL1A1 or COL1A2 mutations. Osteogenesis imperfecta is a systemic heritable disorder of connective tissue
whose cardinal manifestation is bone fragility. In approximately 90% of
individuals with osteogenesis imperfecta, mutations in either of the genes
encoding the pro-alpha1 or pro-alpha2 chains of type I collagen (COL1A1 or
COL1A2) can be identified. Of those without collagen mutations, a number of them
will have mutations involving the enzyme complex responsible for
posttranslational hydroxylation of the position 3 proline residue of COL1A1. Two
of the genes encoding proteins involved in that enzyme complex, LEPRE1 and
cartilage-associated protein, when mutated have been shown to cause autosomal
recessive osteogenesis imperfecta, which has a moderate to severe clinical
phenotype, often indistinguishable from osteogenesis imperfecta types II or III.
Mutations in COL1A1 or COL1A2 which result in an abnormal protein still capable
of forming a triple helix cause a more severe phenotype than mutations that lead
to decreased collagen production as a result of the domit negative effect
mediated by continuous protein turnover. The current standard of care includes a
multidisciplinary approach with surgical intervention when necessary, proactive
physiotherapy, and consideration for the use of bisphosphonates all in attempts
to improve quality of life. Osteogenesis imperfecta (OI) is a heterogeneous group of inherited disorders of
bone formation, resulting in low bone mass and an increased propensity to
fracture. It exhibits a broad spectrum of clinical severity, ranging from
multiple fractures in utero and perinatal death, to normal adult stature and low
fracture incidence. Extra-skeletal features of OI include blue sclera, hearing
loss, skin hyperlaxity, joint hyperextensibility, and dentinogenesis imperfecta.
The proα1(I) and proα2(I) chains of collagen 1 are encoded by the COL1A1 and
COL1A2 genes, respectively; quantitative or qualitative defects in type I
collagen synthesis usually manifest as types of OI or some sub-types of EDS. The
majority of patients (about 90%) with a clinical diagnosis of OI have a mutation
in the COL1A1 or COL1A2 genes, which shows an autosomal domit pattern of
inheritance. Six other genes, CRTAP, LEPRE1, FKBP10, PP1B, SP7/Osterix (OSX),
and SERPINH1, are associated with autosomal recessive forms of OI. However,
other, rare phenotypes have also been described. There are many differential
diagnoses of the short, syndromic child, including chromosomal, single gene, and
multifactorial causes. However, one condition of particular relevance in the
context of this report is the Russell-Silver syndrome (RSS). As originally
described, the RSS is a very specific condition. However, it has subsequently
become an umbrella term for a heterogeneous group of conditions presenting with
short stature and triangular shape to the face. A significant proportion of
these are now believed to be due to imprinting defects at 11p15. However, the
cause in many cases remains unknown. We describe two cases with a phenotypic
overlap between OI and RSS who both have COL1A1 mutations. Thus, a type 1
collagenopathy should be considered in the differential diagnosis of syndromic
short stature. Bruck syndrome (BS) is an autosomal recessive syndromic form of osteogenesis
imperfecta (OI) that is characterized by the additional presence of pterygium
formation. We have recently shown that FKBP10 previously reported as a novel
autosomal recessive OI gene also defines a novel Bruck syndrome locus (BKS3). In
this manuscript, we extend our analysis to describe a mutation previously
described in isolated OI patients and show that it results in BS phenotype in a
Saudi family. More interestingly, we describe a novel FKBP10 mutation that
results in isolated OI as well as BS phenotype in the same family. These
results, combined with recently published work, confirm that FKBP10 is a
bonafide BS locus and lay the foundation for future research into modifiers that
underlie the phenotypic heterogeneity of FKBP10 mutations. A new paradigm has emerged for osteogenesis imperfecta as a collagen-related
disorder. The more prevalent autosomal domit forms of osteogenesis imperfecta
are caused by primary defects in type I collagen, whereas autosomal recessive
forms are caused by deficiency of proteins which interact with type I
procollagen for post-translational modification and/or folding. Factors that
contribute to the mechanism of domit osteogenesis imperfecta include
intracellular stress, disruption of interactions between collagen and
noncollagenous proteins, compromised matrix structure, abnormal cell-cell and
cell-matrix interactions and tissue mineralization. Recessive osteogenesis
imperfecta is caused by deficiency of any of the three components of the
collagen prolyl 3-hydroxylation complex. Absence of 3-hydroxylation is
associated with increased modification of the collagen helix, consistent with
delayed collagen folding. Other causes of recessive osteogenesis imperfecta
include deficiency of the collagen chaperones FKBP10 or Serpin H1. Murine models
are crucial to uncovering the common pathways in domit and recessive
osteogenesis imperfecta bone dysplasia. Clinical management of osteogenesis
imperfecta is multidisciplinary, encompassing substantial progress in physical
rehabilitation and surgical procedures, management of hearing, dental and
pulmonary abnormalities, as well as drugs, such as bisphosphonates and
recombit human growth hormone. Novel treatments using cell therapy or new
drug regimens hold promise for the future. Autosomal recessive osteogenesis imperfecta (AR-OI) is an inherited condition
which in recent years has been shown with increasing genetic and clinical
heterogeneity. In this article, we performed clinical assessment and sought
mutations in patients from 10 unrelated families with AR-OI, one of whom was
presented with the additional features of Bruck syndrome (BS). Pathogenic
changes were identified in five different genes: three families had mutations in
FKBP10, three in SERPINF1, two in LEPRE1, one in CRTAP, and one in PPIB. With
the exception of a FKBP10 mutation in the BS case, all changes are novel. Of
note, insertion of an AluYb8 repetitive element was detected in exon 6 of
SERPINF1. Since the studied patients had variable manifestations and some
distinctive features, genotype/phenotype correlations are suggested. Autosomal recessive osteogenesis imperfecta (OI) accounts for 10% of all OI
cases, and, currently, mutations in 10 genes (CRTAP, LEPRE1, PPIB, SERPINH1,
FKBP10, SERPINF1, SP7, BMP1, TMEM38B, and WNT1) are known to be responsible for
this form of the disease. PEDF is a secreted glycoprotein of the serpin
superfamily that maintains bone homeostasis and regulates osteoid
mineralization, and it is encoded by SERPINF1, currently associated with OI type
VI (MIM 172860). Here, we report a consanguineous Brazilian family in which
multiple individuals from at least 4 generations are affected with a severe form
of OI, and we also report an unrelated individual from the same small city in
Brazil with a similar but more severe phenotype. In both families the same
homozygous SERPINF1 19-bp deletion was identified which is not known in the
literature yet. We described intra- and interfamilial clinical and radiological
phenotypic variability of OI type VI caused by the same homozygous SERPINF1
19-bp deletion and suggest a founder effect. Furthermore, the SERPINF1
genotypes/phenotypes reported so far in the literature are reviewed. Osteogenesis Imperfecta (OI) is an inherited bone fragility disorder most
commonly associated with autosomal domit mutations in the type I collagen
genes. Autosomal recessive mutations in a number of genes have also been
described, including the BMP1 gene that encodes the mammalian Tolloid (mTLD) and
its shorter isoform bone morphogenic protein-1 (BMP1). To date, less than 20
individuals with OI have been identified with BMP1 mutations, with skeletal
phenotypes ranging from mild to severe and progressively deforming. In the
majority of patients, bone fragility was associated with increased bone mineral
density (BMD); however, the full range of phenotypes associated with BMP1
remains unclear. Here, we describe three children with mutations in BMP1
associated with a highly variable phenotype: a sibship homozygous for the
c.2188delC mutation that affects only the shorter BMP1 isoform and a further
patient who is compound heterozygous for a c.1293C>G nonsense mutation and a
c.1148G>A missense mutation in the CUB1 domain. These individuals had recurrent
fractures from early childhood, are hypermobile and have no evidence of
dentinogenesis imperfecta. The homozygous siblings with OI had normal areal BMD
by dual energy X-ray absorptiometry whereas the third patient presented with a
high bone mass phenotype. Intravenous bisphosphonate therapy was started in all
patients, but discontinued in two patients and reduced in another due to
concerns about increasing bone stiffness leading to chalk-stick fractures. Given
the association of BMP1-related OI with very high bone material density,
concerns remain whether anti-resorptive therapy is indicated in this ultra-rare
form of OI.© 2016 Wiley Periodicals, Inc. Osteogenesis imperfecta (OI) is a genetic disorder characterised by low bone
mineral density resulting in fractures. 85-90% of patients with OI carry a
variant in the type 1 collagen genes, COL1A1 and COL1A2, which follows an
autosomal domit pattern of inheritance. However, within the last two decades,
there have been growing number of variants identified in genes that follow an
autosomal recessive pattern of inheritance. Our proband is a child born in
Mexico with multiple fractures of ribs, minimal calvarial mineralisation,
platyspondyly, marked compression and deformed long bones. He also presented
with significant hydranencephaly, requiring ventilatory support from birth, and
died at 8days of age. A homozygous c.338_357delins22 variant in exon 2 of
SERPINH1 was identified. This gene encodes heat shock protein 47, a
collagen-specific chaperone which binds to the procollagen triple helix and is
responsible for collagen stabilisation in the endoplasmic reticulum. There is
minimal literature on the mechanism of action for variants in SERPINH1 resulting
in osteogenesis imperfecta. Here we discuss this rare, previously unreported
variant, and expand on the phenotypic presentation of this novel variant
resulting in a severe, lethal phenotype of OI in association with
hydranencephaly. We sought to characterize the phenotypes and identify the SEC24D gene mutations
associated with Chinese families of osteogenesis imperfecta (OI). Using
whole-exome sequencing, we discovered two novel compound SEC24D mutations of OI
patients. Our study extended both the phenotypic and the genotype of the OI
patients with SEC24D mutations.
INTRODUCTION: To date, only three compound heterozygous mutations in the SEC24D
gene have been found to cause recessively inherited forms of OI. We sought to
characterize the phenotypes and to identify the SEC24D gene mutations associated
with Chinese families with OI.
METHODS: Using whole-exome sequencing in two probands, we identified two novel
compound heterozygous mutations in SEC24D. In family 1, the proband was a
23-year-old male; he had recurrent fractures and dentinogenesis imperfecta. His
anterior fontanel was not closed, and he showed facial dysmorphism. A computed
tomography three-dimensional imaging of the cranium showed skull deformities
associated with a broad ossification defect in the frontoapical area, a widened
sagittal suture, and Wormian bones. In family 2, the proband was a 7-year-old
boy, who also had recurrent fractures and dentinogenesis imperfecta. His
anterior fontanel was not closed, and he did not have obvious facial
dysmorphism.
RESULTS: We identified one novel compound heterozygous missense substitution in
the proband in family 1, including c.2723G>A (p. Cys908Tyr) and c.2842T>C (p.
Ser948Pro). In the proband in family 2, we identified another novel compound
heterozygous missense substitution, including c.938G>A (p. Arg313His) and
c.875C>T (p. Pro292Leu).
CONCLUSIONS: We discovered two novel compound SEC24D mutations of autosomal
recessive OI patients. Our study extended both the phenotypic and the genotypic
spectrum of the autosomal recessive OI patients with SEC24D mutations. BACKGROUND: Osteogenesis imperfecta (OI) is a heterogeneous bone disorder
characterized by recurrent fractures. Although most cases of OI have
heterozygous mutations in COL1A1 or COL1A2 and show autosomal domit
inheritance, during the last years there has been an explosion in the number of
genes responsible for both recessive and domit forms of this condition.
Herein, we have analyzed a cohort of patients with OI, all offspring of
unaffected parents, to determine the spectrum of variants accounting for these
cases. Twenty patients had nonrelated parents and were sporadic, and 21 were
born to consanguineous relationships.
METHODS: Mutation analysis was performed using a next-generation sequencing gene
panel, homozygosity mapping, and whole exome sequencing (WES).
RESULTS: Patients offspring of nonconsanguineous parents were mostly identified
with COL1A1 or COL1A2 heterozygous changes, although there were also a few cases
with IFITM5 and WNT1 heterozygous mutations. Only one sporadic patient was a
compound heterozygote for two recessive mutations. Patients offspring of
consanguineous parents showed homozygous changes in a variety of genes including
CRTAP,FKBP10,LEPRE1,PLOD2,PPIB,SERPINF1,TMEM38B, and WNT1. In addition, two
patients born to consanguineous parents were found to have de novo COL1A1
heterozygous mutations demonstrating that causative variants in the collagen I
structural genes cannot be overlooked in affected children from consanguineous
couples. Further to this, WES analysis in probands lacking mutations in OI genes
revealed deleterious variants in SCN9A,NTRK1, and SLC2A2, which are associated
with congenital indifference to pain (CIP) and Fanconi-Bickel syndrome (FBS).
CONCLUSION: This work provides useful information for clinical and genetic
diagnosis of OI patients with no positive family history of this disease. Our
data also indicate that CIP and FBS are conditions to be considered in the
differential diagnosis of OI and suggest a positive role of SCN9A and NTRK1 in
bone development. |
What is masitinib an inhibitor of? | Masitinib is an inhibitor of mast cell-glia axis and a Fyn kinase blocker. It is an oral tyrosine kinase inhibitor with activity against c-Kit and platelet-derived growth factor receptors (PDGFR). | This study evaluated the therapeutic potential of masitinib, an oral tyrosine
kinase inhibitor with activity against c-Kit and platelet-derived growth factor
receptors (PDGFR), to reduce ischemic brain area and neurological deficit. Using
a well-established filament model of ischemic stroke in rats, the responses to
oral treatment with masitinib alone or in combination with recombit tissue
plasminogen activator (rt-PA) were compared to those after rt-PA (10 mg/kg
intravenously (i.v.)) monotherapy. In both cases, two doses of masitinib were
used--25 or 100 mg/kg, twice per day. Ischemic brain area and the neurological
deficit were assessed using the triphenyltetrazolium chloride (TTC) method and
behavioral neurological tests, respectively. Masitinib, as a single agent,
reduced significantly the infarct size, as compared with the stroke control
group. Brain ischemic area decreased from 9.14 to 4.36 % (25 mg/kg) or 2.60 %
(100 mg/kg). Moreover, a combined treatment of masitinib with rt-PA produced a
stronger effect than the one observed after each of the compound alone. The size
of the brain ischemic area (rt-PA 1.67 %) was further reduced to 0.83 or 0.7 %
at masitinib doses of 25 and 100 mg/kg, respectively. Masitinib reduced
significantly brain ischemia induced by experimental stroke and potentiated the
therapeutic effect of rt-PA. Alzheimer's disease (AD) is a degenerative neurological disorder that is the
most common cause of dementia and disability in older patients. Available
treatments are symptomatic in nature and are only sufficient to improve the
quality of life of AD patients temporarily. A potential strategy, currently
under investigation, is to target cell-signaling pathways associated with
neurodegeneration, in order to decrease neuroinflammation, excitotoxicity, and
to improve cognitive functions. Current review centers on the role of
neuroinflammation and the specific contribution of mast cells to AD
pathophysiology. The authors look at masitinib therapy and the evidence
presented through preclinical and clinical trials. Dual actions of masitinib as
an inhibitor of mast cell-glia axis and a Fyn kinase blocker are discussed in
the context of AD pathology. Masitinib is in Phase III clinical trials for the
treatment of maligt melanoma, mastocytosis, multiple myeloma,
gastrointestinal cancer and pancreatic cancer. It is also in Phase II/III
clinical trials for the treatment of multiple sclerosis, rheumatoid arthritis
and AD. Additional research is warranted to better investigate the potential
effects of masitinib in combination with other drugs employed in AD treatment. |
What is the name of the RNAi investigational drug being developed against hereditary amyloidosis? | Patisiran. | BACKGROUND: Patisiran is an investigational RNA interference (RNAi) therapeutic
in development for the treatment of hereditary ATTR (hATTR) amyloidosis, a
progressive disease associated with significant disability, morbidity, and
mortality.
METHODS: Here we describe the rationale and design of the Phase 3 APOLLO study,
a randomized, double-blind, placebo-controlled, global study to evaluate the
efficacy and safety of patisiran in patients with hATTR amyloidosis with
polyneuropathy. Eligible patients are 18-85 years old with hATTR amyloidosis,
investigator-estimated survival of ≥2 years, Neuropathy Impairment Score (NIS)
of 5-130, and polyneuropathy disability score ≤IIIb. Patients are randomized 2:1
to receive either intravenous patisiran 0.3 mg/kg or placebo once every 3 weeks.
The primary objective is to determine the efficacy of patisiran at 18 months
based on the difference in the change in modified NIS+7 (a composite measure of
motor strength, sensation, reflexes, nerve conduction, and autonomic function)
between the patisiran and placebo groups. Secondary objectives are to evaluate
the effect of patisiran on Norfolk-Diabetic Neuropathy quality of life
questionnaire score, nutritional status (as evaluated by modified body mass
index), motor function (as measured by NIS-weakness and timed 10-m walk test),
and autonomic symptoms (as measured by the Composite Autonomic Symptom Score-31
questionnaire). Exploratory objectives include assessment of cardiac function
and pathologic evaluation to assess nerve fiber innervation and amyloid burden.
Safety of patisiran will be assessed throughout the study.
DISCUSSION: APOLLO represents the largest randomized, Phase 3 study to date in
patients with hATTR amyloidosis, with endpoints that capture the multisystemic
nature of this disease.
TRIAL REGISTRATION: This trial is registered at clinicaltrials.gov ( NCT01960348
); October 9, 2013. |
What is the function of the H19 (ICR) locus in the human genome? | We found that localized DNA demethylation at the H19 imprinting control region (ICR) induced by 5-AzaCdR, reduced IGF2, increased H19 expression, increased CTCF and cohesin recruitment and changed histone modifications. | We have identified a region with characteristics of a paternal-specific
methylation imprint at the human H19 locus. This region, extending from -2.0 kb
upstream to the start of transcription, is heavily methylated in sperm and on
the paternal allele in somatic cells. This methylation was preserved during
pre-implantation. Structural analysis revealed the presence of CpG islands and a
large direct repeat with a 400 bp sequence reiterated several times, but no
significant sequence homology to the corresponding region of the mouse H19 gene.
These findings could suggest a role for secondary DNA structure in genomic
imprinting across the species, and they also present a puzzling aspect of the
evolution of the H19 regulatory region in human and mouse. A subset of genes in mammals are subject to genomic imprinting. The mouse H19
gene, for example, is active only when maternally inherited and the neighboring
Igf2 gene is paternally expressed. This imprinted expression pattern is
regulated by the imprinting control region (ICR) upstream of the H19 gene. A
maternally inherited H19 ICR inhibits Igf2 gene activation by the downstream
enhancer due to its insulator function while it suppresses H19 gene
transcription by promoter DNA methylation when paternally inherited. These
parent-of-origin specific functions depend on the allele-specific methylation of
the ICR DNA, which is established during gametogenesis. Therefore, the ICR may
also function as a landmark for epigenetic modifications. To examine whether the
ICR confers these activities autonomously, we introduced a 2.9-kbp
ICR-containing DNA fragment into a human beta-globin yeast artificial chromosome
at the 3' end of the locus control region and established transgenic mouse
lines. Expression of all of the beta-like globin genes was higher when the
transgene was paternally inherited. In accord with this result, transgenic ICR
DNA from nucleated erythrocytes was more heavily methylated when paternally
transmitted. Chromatin immunoprecipitation assays confirmed that CCCTC binding
factor is preferentially recruited to the maternal transgenic ICR in vivo.
Surprisingly however, the parent-of-origin specific methylation pattern was not
observed in germ cell DNA in testis, demonstrating that methylation was
established after fertilization. Thus, the ICR autonomously recapitulated
imprinting within the normally nonimprinted transgenic beta-globin gene locus,
but the temporal establishment of imprinting methylation differs from that at
the endogenous Igf2/H19 locus. Genomic imprinting at the H19/Igf2 locus is governed by a cis-acting
Imprinting-Control Region (ICR), located 2 kb upstream of the H19 gene. This
region possesses an insulator function which is activated on the unmethylated
maternal allele through the binding of the CTCF factor. It has been previously
reported that paternal transmission of the H19(SilK) deletion, which removes the
3' portion of H19 ICR, leads to the loss of H19 imprinting. Here we show that,
in the liver, this reactivation of the paternal H19 gene is concomitant to a
dramatic decrease in Igf2 mRNA levels. This deletion alters higher-order
chromatin architecture, Igf2 promoter usage and tissue-specific expression.
Therefore, when methylated, the 3' portion of the H19 ICR is a bi-functional
regulatory element involved not only in H19 imprinting but also in 'formatting'
the higher-order chromatin structure for proper tissue-specific expression of
both H19 and Igf2 genes. It is thought that the H19 imprinting control region (ICR) directs the silencing
of the maternally inherited Igf2 allele through a CTCF-dependent chromatin
insulator. The ICR has been shown to interact physically with a silencer region
in Igf2, differentially methylated region (DMR)1, but the role of CTCF in this
chromatin loop and whether it restricts the physical access of distal enhancers
to Igf2 is not known. We performed systematic chromosome conformation capture
analyses in the Igf2/H19 region over >160 kb, identifying sequences that
interact physically with the distal enhancers and the ICR. We found that, on the
paternal chromosome, enhancers interact with the Igf2 promoters but that, on the
maternal allele, this is prevented by CTCF binding within the H19 ICR. CTCF
binding in the maternal ICR regulates its interaction with matrix attachment
region (MAR)3 and DMR1 at Igf2, thus forming a tight loop around the maternal
Igf2 locus, which may contribute to its silencing. Mutation of CTCF binding
sites in the H19 ICR leads to loss of CTCF binding and de novo methylation of a
CTCF target site within Igf2 DMR1, showing that CTCF can coordinate regional
epigenetic marks. This systematic chromosome conformation capture analysis of an
imprinting cluster reveals that CTCF has a critical role in the epigenetic
regulation of higher-order chromatin structure and gene silencing over
considerable distances in the genome. The parent-of-origin-dependent expression of IGF2 and H19 is controlled by the
imprinting center 1 (IC1) consisting of a methylation-sensitive chromatin
insulator. IC1 is normally methylated on the paternal chromosome and
nonmethylated on the maternal chromosome. We found that 22 cases in a large
cohort of patients affected by Beckwith-Wiedemann syndrome (BWS) had IC1
methylated on both parental chromosomes, resulting in biallelic activation of
IGF2 and biallelic silencing of H19. These individuals had marked macrosomia and
high incidence of Wilms' tumor. A subset of these patients had 1.4- to 1.8-kb
deletions with hypermethylation of the remaining IC1 region and fully penetrant
BWS phenotype when transmitted maternally. Another subset of individuals with
IC1 hypermethylation had a similar clinical phenotype but no mutation in the
local vicinity. All these cases were sporadic and in at least two families
affected and unaffected members shared the same maternal IC1 allele but not the
abnormal maternal epigenotype. Similarly, no IC1 deletion was detected in 10
nonsyndromic Wilms' tumors with IC1 hypermethylation. In conclusion, methylation
defects at the IGF2-H19 locus can result from inherited mutations of the
imprinting center and have high recurrence risk or arise independently from the
sequence context and not transmitted to the progeny. The imprinted expression of the mouse Igf2/H19 locus is governed by the
differential methylation of the imprinting control region (ICR), which is
established initially in germ cells and subsequently maintained in somatic
cells, depending on its parental origin. By grafting a 2.9-kbp H19 ICR fragment
into a human beta-globin yeast artificial chromosome in transgenic mice, we
previously showed that the ICR could recapitulate imprinted methylation and
expression at a heterologous locus, suggesting that the H19 ICR in the
beta-globin locus contained sufficient information to maintain the methylation
mark (K. Tanimoto, M. Shimotsuma, H. Matsuzaki, A. Omori, J. Bungert, J. D.
Engel, and A. Fukamizu, Proc. Natl. Acad. Sci. USA 102:10250-10255, 2005).
Curiously, however, the transgenic H19 ICR was not methylated in sperm, which
was distinct from that seen in the endogenous locus. Here, we reevaluated the
ability of the H19 ICR to mark the parental origin using more rigid criteria. In
the testis, the methylation levels of the solitary 2.9-kbp transgenic ICR
fragment varied significantly between six transgenic mouse lines. However, in
somatic cells, the paternally inherited ICR fragment exhibited consistently
higher methylation levels at five out of six randomly integrated sites in the
mouse genome. These results clearly demonstrated that the H19 ICR could acquire
parent-of-origin-dependent methylation after fertilization independently of the
chromosomal integration site or the prerequisite methylation acquisition in male
germ cells. DNA methylation marks, a key modification of imprinting, are erased in
primordial germ cells and sex specifically re-established during gametogenesis.
Abnormal epigenetic programming has been proposed as a possible mechanism
compromising male fertility. We analysed by pyrosequencing the DNA methylation
status of 47 CpGs located in differentially methylated regions (DMRs), the DMR0
and DMR2 of the IGF2 gene and in the 3rd and 6th CTCF-binding sites of the H19
DMR in human sperm from men with normal semen and patients with teratozoospermia
(T) and/or oligo-astheno-teratozoospermia (OAT). All normal semen samples
presented the expected high global methylation level for all CpGs analysed. In
the teratozoospermia group, 11 of 19 patients presented a loss of methylation at
variable CpG positions either in the IGF2 DMR2 or in both the IGF2 DMR2 and the
6th CTCF of the H19 DMR. In the OAT group, 16 of 22 patients presented a severe
loss of methylation of the 6th CTCF, closely correlated with sperm
concentration. The methylation state of DMR0 and of the 3rd CTCF was never
affected by the pathological status of sperm samples. This study demonstrates
that epigenetic perturbations of the 6th CTCF site of the H19 DMR might be a
relevant biomarker for quantitative defects of spermatogenesis in humans.
Moreover, we defined a methylation threshold sustaining the classification of
patients in two groups, unmethylated and methylated. Using this new
classification of patients, the observed intrinsic imprinting defects of
spermatozoa appear not to impair significantly the outcome of assisted
reproductive technologies. Hyper- and hypomethylation at the IGF2-H19 imprinting control region (ICR)
result in reciprocal changes in IGF2-H19 expression and the two contrasting
growth disorders, Beckwith-Wiedemann syndrome (BWS) and Silver-Russell syndrome
(SRS). DNA methylation of the ICR controls the reciprocal imprinting of IGF2 and
H19 by preventing the binding of the insulator protein, CTCF. We here show that
local changes in histone modifications and CTCF--cohesin binding at the ICR in
BWS and SRS together with DNA methylation correlate with the higher order
chromatin structure at the locus. In lymphoblastoid cells from control
individuals, we found the repressive histone H3K9me3 and H4K20me3 marks
associated with the methylated paternal ICR allele and the bivalent
H3K4me2/H3K27me3 mark together with H3K9ac and CTCF--cohesin associated with the
non-methylated maternal allele. In patient-derived cell lines, the mat/pat
asymmetric distribution of these epigenetic marks was lost with H3K9me3 and
H4K20me3 becoming biallelic in the BWS and H3K4me2, H3K27me3 and H3K9ac together
with CTCF-cohesin becoming biallelic in the SRS. We further show that in BWS and
SRS cells, there is opposing chromatin looping conformation mediated by
CTCF--cohesin binding sites surrounding the locus. In normal cells, lack of
CTCF--cohesin binding at the paternal ICR is associated with monoallelic
interaction between two CTCF sites flanking the locus. CTCF--cohesin binding at
the maternal ICR blocks this interaction by associating with the CTCF site
downstream of the enhancers. The two alternative chromatin conformations are
differently favoured in BWS and SRS likely predisposing the locus to the
activation of IGF2 or H19, respectively. In the mouse Igf2/H19 imprinted locus, differential methylation of the
imprinting control region (H19 ICR) is established during spermatogenesis and is
maintained in offspring throughout development. Previously, however, we observed
that the paternal H19 ICR, when analyzed in yeast artificial chromosome
transgenic mice (YAC-TgM), was preferentially methylated only after
fertilization. To identify the DNA sequences that confer methylation imprinting,
we divided the H19 ICR into two fragments (1.7 and 1.2 kb), ligated them to both
ends of a λ DNA fragment into which CTCF binding sites had been inserted, and
analyzed this in YAC-TgM. The maternally inherited λ sequence, normally
methylated after implantation in the absence of H19 ICR sequences, became
hypomethylated, demonstrating protective activity against methylation within the
ICR. Meanwhile, the paternally inherited λ sequence was hypermethylated before
implantation only when a 1.7-kb fragment was ligated. Consistently, when two
subfragments of the H19 ICR were individually investigated for their activities
in YAC-TgM, only the 1.7-kb fragment was capable of introducing paternal
allele-specific DNA methylation. These results show that postfertilization
methylation imprinting is conferred by a paternal allele-specific methylation
activity present in a 1.7-kb DNA fragment of the H19 ICR, while maternal
allele-specific activities protect the allele from de novo DNA methylation. Mono-allelic expression at the mouse IGF2/H19 locus is controlled by
differential allelic DNA methylation of the imprinting control region (ICR).
Because a randomly integrated H19 ICR fragment, when incorporated into the
genome of transgenic mice (TgM), was allele-specifically methylated in somatic,
but not in germ cells, it was suggested that allele-discriminating epigenetic
signature, set within or somewhere outside of the Tg H19 ICR fragment in germ
cells, was later translated into a differential DNA methylation pattern. To test
if the chicken β-globin HS4 (cHS4) chromatin insulator might interfere with
methylation imprinting establishment at the H19 ICR, we inserted the H19 ICR
fragment, flanked by a set of floxed cHS4 core sequences, into a human β-globin
locus YAC and generated TgM (insulated ICR' TgM). As controls, the cHS4
sequences were removed from one side (5'HS4-deleted ICR') or both sides
(pseudo-WT ICR') of the insulated ICR' by in vivo cre-loxP recombination. The
data show that while maternally inherited transgenic H19 ICR was not methylated
in insulated ICR' TgM, it was significantly methylated upon paternal
transmission, though the level was lower than in the pseudo-WT ICR' control.
Because this reduced level of methylation was also observed in the 5'HS4-deleted
ICR' TgM, we speculate that the phenotype is due to VEZF1-dependent
demethylation activity, rather than the insulator function, borne in cHS4.
Collectively, although we cannot rule out the possibility that cHS4 is incapable
of blocking an allele-discriminating signal from outside of the transgene, the
epigenetic signature appears to be marked intrinsically within the H19 ICR. Parent-of-origin-specific expression at imprinted genes is regulated by
allele-specific DNA methylation at imprinting control regions (ICRs). This
mechanism of gene regulation, where one element controls allelic expression of
multiple genes, is not fully understood. Furthermore, the mechanism of gene
dysregulation through ICR epimutations, such as loss or gain of DNA methylation,
remains a mystery. We have used genetic mouse models to dissect ICR-mediated
genetic and epigenetic regulation of imprinted gene expression. The
H19/insulin-like growth factor 2 (Igf2) ICR has a multifunctional role including
insulation, activation and repression. Microdeletions at the human H19/IGF2 ICR
(IC1) are proposed to be responsible for IC1 epimutations associated with
imprinting disorders such as Beckwith-Wiedemann syndrome (BWS). Here, we have
generated and characterized a mouse model that mimics BWS microdeletions to
define the role of the deleted sequence in establishing and maintaining
epigenetic marks and imprinted expression at the H19/IGF2 locus. These mice
carry a 1.3 kb deletion at the H19/Igf2 ICR [Δ2,3] removing two of four
CCCTC-binding factor (CTCF) sites and the intervening sequence, ∼75% of the ICR.
Surprisingly, the Δ2,3 deletion does not perturb DNA methylation at the ICR;
however, it does disrupt imprinted expression. While repressive functions of the
ICR are compromised by the deletion regardless of tissue type, insulator
function is only disrupted in tissues of mesodermal origin where a significant
amount of CTCF is poly(ADP-ribosyl)ated. These findings suggest that insulator
activity of the H19/Igf2 ICR varies by cell type and may depend on cell-specific
enhancers as well as posttranslational modifications of the insulator protein
CTCF. |
What is the mode of action of teriparatide? | Teripartide is is an effective anabolic (i.e., bone growing) agent used in the treatment of some forms of osteoporosis. | An 18-month randomized double-blind study was conducted in postmenopausal women
with osteoporosis to compare the effects of once-daily teriparatide 20 microg
with alendronate 10 mg on bone histomorphometry. Biopsies were obtained from 42
patients. Indices of bone formation were significantly higher after 6 or 18
months of teriparatide compared with alendronate treatment.
INTRODUCTION: Alendronate and teriparatide increased BMD, assessed by DXA, by
different mechanisms of action, supported by changes in biochemical markers of
bone turnover. The purpose of this cross-sectional study was to explore the
differential effects of these two osteoporosis treatments at the bone tissue
level by examining bone histomorphometric parameters of bone turnover after
either 6 or 18 months of treatment.
MATERIALS AND METHODS: Patients were a cohort from a randomized parallel
double-blind study conducted to compare the effects of once-daily teriparatide
20 microg and alendronate 10 mg in postmenopausal women with osteoporosis.
Transiliac crest bone biopsies were obtained after tetracycline double labeling
from 42 patients treated for 6 months (n = 23) or 18 months (n = 14); 5
additional patients were biopsied from contralateral sides at 6 and 18 months.
Biopsy specimens adequate for quantitative analysis were analyzed by 2D
histomorphometry from 17 patients at 6 months (teriparatide, n = 8; alendronate,
n = 9) and 15 patients at 18 months (teriparatide, n = 8; alendronate, n = 7).
Data were analyzed by two-sample tests.
RESULTS: Histomorphometric indices of bone formation were significantly and
markedly greater in the teriparatide group than in the alendronate group at 6
and 18 months, whereas indices of bone resorption were only significantly
greater in the teriparatide group than in the alendronate group at 6 months.
Bone formation and activation frequency were significantly lower at 18 months
compared with 6 months in the teriparatide group, returning to levels comparable
with untreated postmenopausal women. In the teriparatide group, the peak in
histomorphometric bone formation indices coincided with peak levels for
N-terminal propeptide of type I collagen, a biochemical marker of bone
formation. The degree of mineralization was lower at 18 months than at 6 months
with treatment in both groups but was not different between groups.
CONCLUSIONS: These results confirm the opposite mechanisms of action of
teriparatide and alendronate on bone remodeling and confirm the bone formation
effect of teriparatide. The management of osteoporosis has been dominated by the use of antiresorptive
agents, such as bisphosphonates and raloxifene, which have been shown to
effectively reduce the risk of osteoporotic fractures. Teriparatide, a
recombit human parathyroid hormone, has recently become available. It has an
altogether different action. Teriparatide is an anabolic agent that primarily
stimulates bone formation. This leads to an increase in bone volume and the bone
structure and microarchitecture is restored by an increase in trabecular
thickness and an increase in the number of connections between trabeculae.
Clinically relevant studies on teriparatide in postmenopausal women and in men
have shown that it significantly lowers the risk of fracture. Teriparatide
should not be combined with bisphosphonates. There are no clear recommendations
on the order of treatment with teriparatide and bisphosphonates. The high costs
ofteriparatide have slowed the trajectory from registration to reimbursement in
the Netherlands. Since 1 February 2005 teriparatide may be prescribed for
postmenopausal women with serious osteoporosis i.e. with a minimum of 1
osteoporotic fracture who (a) despite treatment with bisphosphonates or
raloxifene after 2 prolapsed vertebrae once again have 1 or more fractures
(inadequate response); (b) can tolerate neither bisphosphonates nor raloxifene;
(c) are prescribed the drug by their treating medical specialist. Teriparatide (recombit human parathyroid hormone ) is an anabolic agent
approved for the treatment of patients at high risk for fracture. The Fracture
Prevention Trial administered teriparatide to treatment-naïve patients, leading
to its US Food and Drug Administration approval in 2002. Clinical trial data
using antiresorptive agents administered before, during, and after any
parathyroid hormone (PTH) therapy, as well as alternative PTH dosing, have
provided additional insight yet raise fundamental questions about the most
appropriate use of teriparatide. This article provides an update on
teriparatide, focusing on its mechanism of action compared with other
antiresorptive agents, indications, adverse effects, therapy duration,
combination therapy, contraindications, and cost effectiveness. Anabolic therapy for osteoporosis has become the most desirable therapeutic
option for menopausal osteoporosis. The anabolic agents currently in clinical
use are reviewed. Teriparatide (recombit human 1-34 parathyroid hormone) is
used to treat women with menopausal osteoporosis and men at high risk for
fractures. Despite PTH's clinical use, the mechanism underlying its anabolic
action requires greater elucidation. Proteol (strontium ranelate) acts by
inhibiting bone resorption and presumably promoting bone formation. Though
clinical trials have shown that strontium ranelate reduces the frequency of both
vertebral and non-vertebral fractures, its molecular target remains
controversial. Lately, with the discontinuation of estrogen replacement therapy,
phytoestrogens are gaining much attention, chiefly as prophylactic agents.
Though ipriflavone stimulates osteoblast function in vitro and favorably
influences bone turnover and spinal bone mineral density in peri- and
postmenopausal women, its clinical use is currently rather limited. As with PTH
and strontium ranelate, the mode of action of ipriflavone requires much greater
elucidation. Since osteoporosis therapies are long-term, safety is a major
consideration. PTH has been reported to be associated with incidence of
osteosarcoma and strontium ranelate with DRESS syndrome. Therefore, target-based
(and osteoblast-specific) development of molecules is expected to improve the
safety profile of anabolics. Calcium-sensing receptor, insulin-like growth
factor-1, members of wingless tail signaling family, and sclerostin are emerging
concepts in bone anabolic therapy. We will cover the preclinical development of
some bone anabolic agents under active investigation. The objective of this systematic review was to examine the influence of
treatments for postmenopausal osteoporosis (parathyroid hormone [PTH],
bisphosphonates, strontium ranelate, and denosumab) on bone quality and discuss
the clinical implications. Most bone-quality data for PTH is from teriparatide.
Teriparatide results in a rapid increase in bone-formation markers, followed by
increases in bone-resorption markers, opening an "anabolic window," a period of
time when PTH is maximally anabolic. Teriparatide reverses the structural damage
seen in osteoporosis and restores the structure of trabecular bone. It has a
positive effect on cortical bone, and any early increases in cortical porosity
appear to be offset by increases in cortical thickness and diameter.
Bisphosphonates are antiresorptive agents which reduce bone turnover, improve
trabecular microarchitecture, and mineralization. Concerns have been raised that
the prolonged antiresorptive action of bisphosphonates may lead to failure to
repair microdamage, resulting in microcracks and atypical fragility. Strontium
ranelate is thought to have a mixed mode of action, increasing bone formation
and decreasing bone resorption. Strontium ranelate improves cortical thickness,
trabecular number, and connectivity, with no change in cortical porosity.
Denosumab exerts rapid, marked, and sustained effects on bone resorption,
resulting in falls in the markers of bone turnover. Evidence from bone-quality
studies suggests that treatment-naive women, aged 60-65 years, with very low BMD
T scores may benefit from PTH as primary therapy to improve bone substrate and
build bone. Post-PTH treatment with bisphosphonates will maintain improvements
in bone quality and reduce the risk of fracture. Teriparatide, a treatment formula of parathyroid hormone (PTH) , is a powerful
anabolic agent on bone, improving its mass, geometry and microarchitecture. It
can decrease the occurrence of fracture in the subjects with increased fracture
risks. Teriparatide can provide good protection against fracture using as
monotherapy, and possibly as combination with an antiresorptive agent such as
zoledronate. Teriparatide should be considered in patients with osteoporosis who
sustained the reduction of BMD or fracture on established bisphosphonates or
SERMs. Treatment period of teriparatide is 1.5-2 years and consecutive use of
antiresorptive agents is greatly recommended to secure long-term protection
against fracture after teriparatide treatment. Teriparatide and bisphosphonates are osteoporosis medications that increase bone
mineral density (BMD) and prevent fracture, but each has a different mechanism
of action. Teriparatide promotes bone formation, while bisphosphonates suppress
bone resorption. In the clinical setting, however, drug selection is not always
tailored to the particular clinical condition of the patient or mechanism of
action of the drug. We compared the effects of teriparatide and the
bisphosphonate risedronate on bone metabolism using two ovariectomized rat
models to elucidate the optimal use of these two drugs in the clinical setting.
We first performed dose-finding experiments to determine the equivalent
effective doses of each drug (5.6 and 3.0 µg/kg for teriparatide and
risedronate, respectively). We then compared the effects of these doses on bone
metabolism after subcutaneous administration three times weekly for 4 months
starting either the day after ovariectomy (preventive study) or 12 months after
ovariectomy (therapeutic study). The increase in proximal tibial BMD under the
physical conditions that increased bone turnover at 1 to 2 months after
ovariectomy was greater in the risedronate group than in the teriparatide group.
In contrast, the increases in lumbar vertebral BMD and bone strength under the
physical conditions that significantly decreased BMD and bone strength at
12 months after ovariectomy were greater in the teriparatide group than in the
risedronate group. The present study provides important information on the
selection of antiosteoporotic drugs, including teriparatide and risedronate, in
treatment protocols tailored to the clinical conditions of patients and drug
mechanisms. Teriparatide [PTH (1-34)] is a genetically engineered analog of human
parathyroid hormone that acts as an anabolic drug by increasing activity in both
osteoblasts and osteoclasts. Intermittent (once-daily) doses of teriparatide
seem to stimulate osteoblast activity and therefore result in a net increase of
bone formation. It is recommended for use in post-menopausal women (PMW), men
with hypogonadal osteoporosis, as well as men and women with
glucocorticoid-induced osteoporosis. In vivo studies have generated important
findings regarding teriparatide's role in the enhancement of fracture healing.
The intention of this article is to review the clinical findings of teriparatide
to stimulate fracture healing. The drug was shown in a prospective randomized,
double blind study to achieve earlier radiographic cortical bridging of three of
four cortices (7.4 weeks) compared to patients who were assigned to the placebo
group (9.1 weeks). Another study compared mean time for healing and functional
outcome in two groups of elderly women who had suffered osteoporotic pelvic
fractures: one group received daily 100 μg parathyroid hormone (1-84)
injections, while the other group received no treatment. Patients who received
the PTH (1-84) injections accelerated radiographic and clinical fracture healing
(7.8 weeks) when compared to patients who received no treatment (12.6 weeks,
p<0.001). Numerous case series state the safety and potential benefits of
teriparatide use in patients recovering from fractures. In the following
scenarios, teriparatide might be considered in patients with osteoporosis and a
fracture: (1) patients with severe osteoporosis with use of bisphosphonates for
a number of years with a fracture not expected to predictably unite, e.g.
atypical femur fracture or open tibia fracture, (2) in cases where an
osteoporotic patient has failed fracture healing and is considering surgical
treatment e.g. non-union surgery. It seems prudent to reevaluate these patients
frequently and reconsider which drug class of osteoporotic drug is best for the
patient. Finally, it must be stressed that we do not recommend teriparatide in
osteoporotic patients that may be well treated with bisphosphonates and a
fracture is expected to heal uneventfully, nor when patients with metabolically
normal bone have a fracture. |
What is the Strelka workflow? | Whole genome and exome sequencing of matched tumor-normal sample pairs is becoming routine in cancer research. The consequent increased demand for somatic variant analysis of paired samples requires methods specialized to model this problem so as to sensitively call variants at any practical level of tumor impurity. Strelka is a method for somatic SNV and small indel detection from sequencing data of matched tumor-normal samples. The method uses a novel Bayesian approach which represents continuous allele frequencies for both tumor and normal samples, while leveraging the expected genotype structure of the normal. This is achieved by representing the normal sample as a mixture of germline variation with noise, and representing the tumor sample as a mixture of the normal sample with somatic variation. A natural consequence of the model structure is that sensitivity can be maintained at high tumor impurity without requiring purity estimates. Strelka has superior accuracy and sensitivity on impure samples compared with approaches based on either diploid genotype likelihoods or general allele-frequency tests. | MOTIVATION: Whole genome and exome sequencing of matched tumor-normal sample
pairs is becoming routine in cancer research. The consequent increased demand
for somatic variant analysis of paired samples requires methods specialized to
model this problem so as to sensitively call variants at any practical level of
tumor impurity.
RESULTS: We describe Strelka, a method for somatic SNV and small indel detection
from sequencing data of matched tumor-normal samples. The method uses a novel
Bayesian approach which represents continuous allele frequencies for both tumor
and normal samples, while leveraging the expected genotype structure of the
normal. This is achieved by representing the normal sample as a mixture of
germline variation with noise, and representing the tumor sample as a mixture of
the normal sample with somatic variation. A natural consequence of the model
structure is that sensitivity can be maintained at high tumor impurity without
requiring purity estimates. We demonstrate that the method has superior accuracy
and sensitivity on impure samples compared with approaches based on either
diploid genotype likelihoods or general allele-frequency tests.
AVAILABILITY: The Strelka workflow source code is available at
ftp://[email protected]/.
CONTACT: [email protected] BACKGROUND: High-throughput sequencing is rapidly becoming common practice in
clinical diagnosis and cancer research. Many algorithms have been developed for
somatic single nucleotide variant (SNV) detection in matched tumor-normal DNA
sequencing. Although numerous studies have compared the performance of various
algorithms on exome data, there has not yet been a systematic evaluation using
PCR-enriched amplicon data with a range of variant allele fractions. The
recently developed gold standard variant set for the reference individual
NA12878 by the NIST-led "Genome in a Bottle" Consortium (NIST-GIAB) provides a
good resource to evaluate admixtures with various SNV fractions.
RESULTS: Using the NIST-GIAB gold standard, we compared the performance of five
popular somatic SNV calling algorithms (GATK UnifiedGenotyper followed by simple
subtraction, MuTect, Strelka, SomaticSniper and VarScan2) for matched
tumor-normal amplicon and exome sequencing data.
CONCLUSIONS: We demonstrated that the five commonly used somatic SNV calling
methods are applicable to both targeted amplicon and exome sequencing data.
However, the sensitivities of these methods vary based on the allelic fraction
of the mutation in the tumor sample. Our analysis can assist researchers in
choosing a somatic SNV calling method suitable for their specific needs. Next generation sequencing is extensively applied to catalogue somatic mutations
in cancer, in research settings and increasingly in clinical settings for
molecular diagnostics, guiding therapy decisions. Somatic variant callers
perform paired comparisons of sequencing data from cancer tissue and matched
normal tissue in order to detect somatic mutations. The advent of many new
somatic variant callers creates a need for comparison and validation of the
tools, as no de facto standard for detection of somatic mutations exists and
only limited comparisons have been reported. We have performed a comprehensive
evaluation using exome sequencing and targeted deep sequencing data of paired
tumor-normal samples from five breast cancer patients to evaluate the
performance of nine publicly available somatic variant callers: EBCall, Mutect,
Seurat, Shimmer, Indelocator, Somatic Sniper, Strelka, VarScan 2 and Virmid for
the detection of single nucleotide mutations and small deletions and insertions.
We report a large variation in the number of calls from the nine somatic variant
callers on the same sequencing data and highly variable agreement. Sequencing
depth had markedly diverse impact on individual callers, as for some callers,
increased sequencing depth highly improved sensitivity. For SNV calling, we
report EBCall, Mutect, Virmid and Strelka to be the most reliable somatic
variant callers for both exome sequencing and targeted deep sequencing. For
indel calling, EBCall is superior due to high sensitivity and robustness to
changes in sequencing depths. Four popular somatic single nucleotide variant (SNV) calling methods (Varscan,
SomaticSniper, Strelka and MuTect2) were carefully evaluated on the real whole
exome sequencing (WES, depth of ~50X) and ultra-deep targeted sequencing
(UDT-Seq, depth of ~370X) data. The four tools returned poor consensus on
candidates (only 20% of calls were with multiple hits by the callers). For both
WES and UDT-Seq, MuTect2 and Strelka obtained the largest proportion of COSMIC
entries as well as the lowest rate of dbSNP presence and
high-alternative-alleles-in-control calls, demonstrating their superior
sensitivity and accuracy. Combining different callers does increase reliability
of candidates, but narrows the list down to very limited range of tumor read
depth and variant allele frequency. Calling SNV on UDT-Seq data, which were of
much higher read-depth, discovered additional true-positive variations, despite
an even more tremendous growth in false positive predictions. Our findings not
only provide valuable benchmark for state-of-the-art SNV calling methods, but
also shed light on the access to more accurate SNV identification in the future. Identifying genomic variants is a fundamental first step toward the
understanding of the role of inherited and acquired variation in disease. The
accelerating growth in the corpus of sequencing data that underpins such
analysis is making the data-download bottleneck more evident, placing
substantial burdens on the research community to keep pace. As a result, the
search for alternative approaches to the traditional "download and analyze"
paradigm on local computing resources has led to a rapidly growing demand for
cloud-computing solutions for genomics analysis. Here, we introduce the Genome
Variant Investigation Platform (GenomeVIP), an open-source framework for
performing genomics variant discovery and annotation using cloud- or local
high-performance computing infrastructure. GenomeVIP orchestrates the analysis
of whole-genome and exome sequence data using a set of robust and popular
task-specific tools, including VarScan, GATK, Pindel, BreakDancer, Strelka, and
Genome STRiP, through a web interface. GenomeVIP has been used for genomic
analysis in large-data projects such as the TCGA PanCanAtlas and in other
projects, such as the ICGC Pilots, CPTAC, ICGC-TCGA DREAM Challenges, and the
1000 Genomes SV Project. Here, we demonstrate GenomeVIP's ability to provide
high-confidence annotated somatic, germline, and de novo variants of potential
biological significance using publicly available data sets. Precision medicine attempts to individualize cancer therapy by matching
tumor-specific genetic changes with effective targeted therapies. A crucial
first step in this process is the reliable identification of cancer-relevant
variants, which is considerably complicated by the impurity and heterogeneity of
clinical tumor samples. We compared the impact of admixture of non-cancerous
cells and low somatic allele frequencies on the sensitivity and precision of 19
state-of-the-art SNV callers. We studied both whole exome and targeted gene
panel data and up to 13 distinct parameter configurations for each tool. We
found vast differences among callers. Based on our comprehensive analyses we
recommend joint tumor-normal calling with MuTect, EBCall or Strelka for whole
exome somatic variant calling, and HaplotypeCaller or FreeBayes for whole exome
germline calling. For targeted gene panel data on a single tumor sample,
LoFreqStar performed best. We further found that tumor impurity and admixture
had a negative impact on precision, and in particular, sensitivity in whole
exome experiments. At admixture levels of 60% to 90% sometimes seen in
pathological biopsies, sensitivity dropped significantly, even when variants
were originally present in the tumor at 100% allele frequency. Sensitivity to
low-frequency SNVs improved with targeted panel data, but whole exome data
allowed more efficient identification of germline variants. Effective somatic
variant calling requires high-quality pathological samples with minimal
admixture, a consciously selected sequencing strategy, and the appropriate
variant calling tool with settings optimized for the chosen type of data. |
Describe the Manta algorithm for detection of structural variants | Manta is a method to discover structural variants and indels from next generation sequencing data. Manta is optimized for rapid germline and somatic analysis, calling structural variants, medium-sized indels and large insertions on standard compute hardware in less than a tenth of the time that comparable methods require to identify only subsets of these variant types: for example NA12878 at 50× genomic coverage is analyzed in less than 20 min. Manta can discover and score variants based on supporting paired and split-read evidence, with scoring models optimized for germline analysis of diploid individuals and somatic analysis of tumor-normal sample pairs. Call quality is similar to or better than comparable methods, as determined by pedigree consistency of germline calls and comparison of somatic calls to COSMIC database variants. Manta consistently assembles a higher fraction of its calls to base-pair resolution, allowing for improved downstream annotation and analysis of clinical significance. | : We describe Manta, a method to discover structural variants and indels from
next generation sequencing data. Manta is optimized for rapid germline and
somatic analysis, calling structural variants, medium-sized indels and large
insertions on standard compute hardware in less than a tenth of the time that
comparable methods require to identify only subsets of these variant types: for
example NA12878 at 50× genomic coverage is analyzed in less than 20 min. Manta
can discover and score variants based on supporting paired and split-read
evidence, with scoring models optimized for germline analysis of diploid
individuals and somatic analysis of tumor-normal sample pairs. Call quality is
similar to or better than comparable methods, as determined by pedigree
consistency of germline calls and comparison of somatic calls to COSMIC database
variants. Manta consistently assembles a higher fraction of its calls to
base-pair resolution, allowing for improved downstream annotation and analysis
of clinical significance. We provide Manta as a community resource to facilitate
practical and routine structural variant analysis in clinical and research
sequencing scenarios.
AVAILABILITY AND IMPLEMENTATION: Manta is released under the open-source GPLv3
license. Source code, documentation and Linux binaries are available from
https://github.com/Illumina/manta.
CONTACT: [email protected]
SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics
online. |
Is Kummell’s disease an avascular necrosis of the vertebral body? | Yes, Kummell’s disease is an avascular necrosis of the vertebral body. | Avascular necrosis of a vertebral body, a relatively uncommon entity, is caused
by maligcy, infection, radiation, systemic steroid treatment, trauma, and the
like.1 Vertebral osteonecrosis induced by trauma is called Kvmell's disease,
because it was initially described by Hermann Kvmell of Germany in 1891.2 This
paper reported a young female with posttraumatic vertebral osteonecrosis and
analyzed the causes. She was treated by thoracoscopic surgery successfully. We report a case with classic clinical findings and imaging features of
Kummell's disease. Kummell's disease is a post-traumatic vertebral body
collapse. Initially after the trauma patients are usually asymptomatic but after
months they develop a symptomatic and progressive kyphosis of the lower thoracic
or lumbar spine. On a conventional radiograph a collapsed vertebral body with a
fracture cleft is typical and on MRIT2 weighted images the double line sign is
characteristic for Kummell's disease; an increased linear area of
hyper-intensity surrounded by an area of low signal. Percutaneous vertebroplasty
is an adequate treatment for stabilization of the fracture and pain reduction in
patients with Kummell's disease. Delayed post-traumatic osteonecrosis, also known by its eponym Kummell's
disease, is a rarely reported clinical entity that likely occurs with higher
frequency than recognized. We highlight a case of a 75-year-old female household
ambulator who presented with significant thoracolumbar pain and delayed T12
collapse after a ground-level fall. The patient had sustained a trivial fall at
home 4 months prior to this presentation and had been hospitalized in our
institution at that time for a general medical workup. Dedicated spine
radiographs were not obtained during this visit. However, lateral chest
radiograph demonstrated an intact T12 vertebral body. The patient was able to
mobilize successfully with therapy and was discharged home. During the interim
between the initial fall and subsequent presentation, she resumed physical
activity including ambulating independently and performing various housework.
Approximately 4 months following her initial injury, the patient returned to a
local emergency department with vague complaints of abdominal pain without any
history of recent fall or injury. After an unremarkable workup, the patient was
sent home. Ten days later, she represented to our institution's emergency room
with worsening pain. Radiographs and CT scan demonstrated interval collapse of
the T12 vertebral body. A linear vacuum cleft was noted on X-rays and CT. An
extensive workup to exclude other processes such as maligcy or infection,
which was negative, ensued. Delayed post-traumatic vertebral collapse was
diagnosed. A trial of medical management and therapy was attempted, but she
continued to experience significant pain. A T12 vertebroplasty was therefore
offered and performed to stabilize the injury and to relieve the pain. She was
subsequently able to be discharged from the hospital and transitioned back to
home life. At approximately 2 years following her injury, the patient was noted
to be able to ambulate with a walking aid. Her final radiograph after her
surgery demonstrated that the T12 vertebroplasty had maintained its height and
sagittal alignment. This Grand Round case highlights the clinical presentation
of Kummell's disease. Aspects of the clinical entity that will be discussed
include a historical review of the disease, hallmark radiographic findings and
treatment options. Kummell's disease is a rare, delayed posttraumatic collapse of a vertebral body
that can occur several months or even years after an osteoporotic compression
fracture. However, there are few reports of posterior element fractures
associated with Kummell's disease. A 72-year-old man who had sustained an L1
osteoporotic compression fracture 14 months prior was admitted to our
institution with incapacitating back pain. Plain radiographs showed progressive
collapse of the L1 vertebral body and severe kyphosis at the thoracolumbar
junction. Magnetic resoce imaging revealed a posterior element fracture as
well as osteonecrosis of the L1 vertebral body. An L1 percutaneous
vertebroplasty was performed, followed by bone cement-augmented screw fixation
to maintain stability and correct the kyphotic deformity. After surgery, pain
relief was immediate, and the patient was able to walk unassisted. This case
illustrates that continuous axial distraction stress caused by aggravated
kyphosis secondary to Kummell's disease may result in posterior element
fractures. Our discussion concludes with a literature review. Kummell's disease is a spinal disorder characterized by delayed post-traumatic
collapse of a vertebral body with avascular necrosis. Although definitive
treatment for Kummell's disease has not been established, it has been reported
that percutaneous vertebroplasty or kyphoplasty has shown good results. However,
these procedures are not recommended for severely collapsed vertebral bodies
because of the risk of cement leakage or technical difficulties. Authors report
a rare case of spontaneous reduction in vertebral height by the insertion of a
working cannula into the vertebral body in Kummell's disease. Kummell disease, or avascular necrosis of a vertebral body, presents as
vertebral osteonecrosis typically affecting a thoracic vertebra with compression
deformity, intravertebral vacuum cleft, and exaggerated kyphosis weeks to months
after a minor traumatic injury. This rare disease is increasing in prevalence
secondary to an aging population and the associated rise in osteoporosis.
Treatment with vertebroplasty or surgical decompression and fusion is often
required. We present a classic case of Kummell disease to illustrate the salient
features of the condition, with associated imaging findings on computed
tomography and magnetic resoce imaging. INTRODUCTION: Kummell's disease is an avascular necrosis of the vertebral body,
secondary to a vertebral compression fracture. This entity is characterised by
the gradual development in time of a vertebral body collapse following a trivial
spinal trauma, involving a worsening back pain associated with a progressive
kyphosis.
PURPOSES: The aim of this article is to carry out an international literature
review regarding Kummell's disease, addressing its physiopathology,
histopathology, clinical presentation, radiological characteristics and
treatment modalities; at the same time, the literature is updated through the
description of a new and interesting case, symbol of the pathology long-term
potential complications, if not diagnosed and therefore not suitably treated.
CASE REPORT: A patient with osteoporosis, following a slight spinal trauma,
suffered a progressive necrosis of the D11 body; although the radiological exams
showed a constant worsening of the thoracic-lumbar kyphosis and a restriction of
the spinal canal, in another medical centre he was only treated with a corset
and painkillers. A year after the injury, motor deficits concerning the lower
limbs appeared. He was then sent to us and indication for posterior internal
fixation was given. On the basis of both his medical history and radiological
and histological findings, Kummell's disease was diagnosed.
CONCLUSION: It is necessary to have a complete knowledge of the clinical,
pathological and radiological characteristics of Kummell's disease, so as to
follow a correct diagnostic course enabling to prepare the most suitable
therapy. |
What causes "Puffy hand syndrome"? | Puffy hand syndrome is a complication of intravenous drug abuse. | The incidence of vascular complications due to drug abuse is at present
increasing due to new types of drugs and to the different ways of intake of such
substances. The vascular complications related to drug abuse may affect venous,
arterious and lymphatic districts and in particular: ischemia following
intra-arterial injections, arterious and venous pseudoaneurysm, vasculitis,
aneurysms, aortic dissections, abscesses complicated by erosions of vessels,
arteriovenous fistulas, compartment syndrome, superficial and deep venous
thrombosis, septic trombophlebitis, puffy hand syndrome. The scientific
knowledge in this matter is incomplete because of the new pathological cases and
the lack of information regarding the efficacy of different treatments. The
authors report four patients affected by vascular pathologies due to drug abuse.
In one case, a heroin addict has undergone multiple fasciotomies for
compartimental syndrome arising because the patient maintained an innatural
posture for several hours during an overdose coma. In a second case, a segmental
right subclavear deep venous thrombosis has been treated by pharmacological
therapy with satisfactory functional recovery of the arm. A third patient has
been successfully submitted to intra-arterial pharmacological vasodilatation for
generalised lower limbs vasospasm caused by drug abuse. In the last case, the
voluntary swallowing of a great dose of cocaine caused the patient's death after
multiple ischemic and hemorrhagic cerebral episodes. After the description of
these cases, a review of the recent literature and some observations on this
topic are presented. A better knowledge of vascular complications due to drug
abuse should improve the therapeutical approach of these patients. Narcotic addiction may induce systemic and local complications. Intravenous
injections of drugs can cause venous thrombosis, and septic or embolic
complications. The puffy hand sign is a more uncommon complication of hard-core
injection addicts. Three long-term intravenous drug users, two males, one
female, mean age 30.6 years (26-37) presented puffy hands. These patients had
been drug addicts for four to twelve years (mean duration 7.3 years) and had
stopped heroin injections for 3-5 years (mean 4.6), participating in a
buprenorphine substitution program. The edema appeared several years after drug
cessation (1.5-5, mean 2.3). Typically the puffiness was bilateral, the hands
swollen from the proximal segments of the fingers to the wrist. In one patient,
the edema was localized both in the hands and in the feet. The edema was not
pitting and unaffected by elevation. Duplex ultrasound examination of the
extremities was normal. Lymphangiography performed in one patient was consistent
with deep lymphatic destruction. Puffy hand syndrome appears to be the end
result of lymphatic obstruction. Repeated injections of drugs in or outside the
veins destroy the lymphatics. Buprenorphine may play an important role in the
puffy hand sign. Although it is supposed to be administered orally, many drug
addicts use it as an i.v. solution. Because buprenorphine is poorly soluble, it
causes lymphatic obstruction. This type of hand for which no therapy exists must
be differentiated from deep palmar space infection with dorsal edema which
requires incision and drainage. AIM: We studied the pathogenesis of puffy hand syndrome of intravenous drug use.
We hypothesized that injections of high-dose sublingual buprenorphine, instead
of the recommended sublingual administration, could play an important role in
lymphatic obstruction and destruction.
DESIGN AND PARTICIPANTS: We set up a case-control study in substitution centres,
recruiting intravenous drug addicts with and without puffy hands, respectively.
The subjects were asked to answer anonymously a questionnaire of 40 items
comprising social and demographic status, history of illicit drugs use,
buprenorphine misuse and injection practices.
FINDINGS: We included 33 cases and 33 controls, mean age of 34 years. They were
past heroin users, mainly methadone-substituted. In multivariate analysis, sex
(women) (OR = 8.9, P = 0.03), injections in the hands (OR = 5.9, P = 0.03),
injections in the feet (OR = 6.5, P = 0.01) and the absence of tourniquet (OR =
7.0, p = 0.02) were significant risk factors for puffy hand syndrome. In 69.7%
of the cases and 59.4% of the controls, respectively, there was a high-dose
sublingual buprenorphine misuse, although it appeared not to be a significant
risk factor for puffy hand syndrome.
CONCLUSIONS: Injection practices are likely to cause puffy hands syndrome, but
buprenorphine misuse should not be considered as a significant risk factor.
However, intravenous drug users must still be warned of local and systemic
complications of intravenous drug misuse. BACKGROUND: Puffy hand syndrome is a complication of intravenous drug abuse,
which has no current available treatment. Arm and forearm edema are voluminous
and cause functional and aesthetic disturbances. We report two cases
successfully treated by low-stretch bandages.
OBSERVATIONS: A 40-year-old man and a 34-year-old woman, both intravenous drug
users, with puffy hand syndrome were hospitalized for 11 days. Treatment
included daily multilayer bandaging. Lymphedema volumes calculated by utilizing
the formula for a truncated cone decreased by 16% on the left side and 12% on
the right side for the first patient and 31 and 17% for the second. Hand
circumference decreased 4.3 cm on the left side and 3.2 cm on the right side in
case 1, and 2.5 cm and 1.9 cm respectively for case 2. The patients were taught
self-bandaging techniques during their hospital stays. Elastic gloves were
fitted at the end of treatment. Reduction of lymphedema volume remained stable
after 18 months in one patient while for the second patient further treatment
and hospitalization were required due to poor compliance.
DISCUSSION: The pathogenesis of this edema is probably multifactorial: venous,
lymphatic insufficiency and the direct toxicity of injected drugs. Lymphedema
treatment currently consists of low-stretch bandaging and wearing elastic
garments, which is effective in decreasing the volume of puffy hand syndrome. Infections generally occur in intravenous drug abuse (IVDA) patients, most
commonly affecting the spine and proximal joints. Numerous serious
musculoskeletal complications of IVDA may involve the upper extremity, however.
Soft-tissue complications in the upper extremity of IVDA patients include
cellulitis, ulceration, abscess, pyomyositis, septic bursitis, tenosynovitis,
and necrotizing fasciitis. Foreign bodies in soft tissue due to needle fragments
are common findings. Primary bone and joint IVDA complications include
osteomyelitis (acute and chronic) and septic arthritis. Other IVDA complications
in the upper extremity affecting blood vessels and lymphatics include hematoma,
arterial aneurysm and pseudoaneurysm, thrombosis, thrombophlebitis, "puffy hand"
syndrome, and lymphadenopathy. These complications usually present as urgent
issues requiring prompt and accurate evaluation in the acute setting. Diagnostic
imaging not only aids in making the correct diagnosis but also permits precise
definition of the location and extent of these abnormalities. We review the
imaging findings and illustrate a wide range of disabling and even
life-threatening complications affecting the upper extremity of IVDA patients
that require early diagnosis for optimal outcome. Puffy hand syndrome is an unrecognized complication of intravenous drug abuse.
This painless syndrome appears during or after a long period of drug addiction.
It involves the hands and sometimes the forearms, and may cause functional,
aesthetic and social disturbances when the hand volume is important.
Physiopathological mechanisms of the puffy hand syndrome are unclear and include
venous and lymphatic insufficiencies, infectious complications and direct
toxicity of injected drugs and their adulterants. Low-stretch bandage and
elastic garment, usually used in lymphedema treatment, are proposed to treat the
puffy hand syndrome. The authors report an original clinical presentation of factitious disorders of
the upper extremity in an ex-drug-addict patient with puffy hand syndrome.
Chronic self-inflicted ulcerations appeared with sequential manner. The patient
confessed deliberate self-harm and transfer of anxiety on his hands, the aspect
of which had become intolerable. Association of puffy hand syndrome with
comorbid psychosis and major depression explained immediate recurrence of
ulcerations despite fitted medication and long-term psychotherapy. Puffy hand syndrome develops after long-term intravenous drug addiction. It is
characterized by a nonpitting edema, affecting the dorsal side of fingers and
hands with puffy aspect. Frequency and severity of the complications of this
syndrome are rarely reported. Local infectious complications such as cellulitis
can be severe and can enable the diagnosis. Herein, we report the case of a
41-year-old man who went to the emergency department for abdominal pain, fever,
and bullous lesions of legs and arms with edema. Bacteriologic examination of a
closed bullous lesion evidenced a methicillin sensitive Staphylococcus aureus.
The abdomen computed tomography excluded deep infections and peritoneal
effusion. The patient was successfully treated by intravenous oxacillin and
clindamycin. He had a previous history of intravenous heroin addiction. We
retained the diagnosis of puffy hand syndrome revealed by a severe
staphylococcal infection with toxic involvement mimicking a four limbs
cellulitis. Puffy hand syndrome, apart from the chronic lymphedema treatment,
has no specific medication available. Prophylactic measures against skin
infections are essential. Intravenous drug addiction is responsible for many complications, especially
cutaneous and infectious. There is a syndrome, rarely observed in rheumatology,
resulting in "puffy hands": the puffy hand syndrome. We report two cases of this
condition from our rheumatologic consultation. Our two patients had intravenous
drug addiction. They presented with an edema of the hands, bilateral, painless,
no pitting, occurring in one of our patient during heroin intoxication, and in
the other 2 years after stopping injections. In our two patients, additional
investigations (biological, radiological, ultrasound) were unremarkable, which
helped us, in the context, to put the diagnosis of puffy hand syndrome. The
pathophysiology, still unclear, is based in part on a lymphatic toxicity of
drugs and their excipients. There is no etiological treatment but elastic
compression by night has improved edema of the hands in one of our patients. |
Describe mechanism of action of Ozanimod. | Ozanimod is a novel, selective, oral sphingosine-1-phosphate (1 and 5) receptor modulator in clinical development for the treatment of chronic immune-mediated, inflammatory diseases, such as multiple sclerosis and inflammatory bowel disease. Yet its exact mechanism of action is unknown. | Sphingosine 1-phosphate (S1P) receptor modulators possess a unique mechanism of
action as disease-modifying therapy for multiple sclerosis (MS). Subtype 1 S1P
receptors are expressed on the surfaces of lymphocytes and are important in
regulating egression from lymph nodes. The S1P receptor modulators indirectly
antagonize the receptor's function and sequester lymphocytes in lymph nodes.
Fingolimod was the first S1P agent approved in the USA in 2010 for relapsing MS
after two phase III trials (FREEDOMS and TRANSFORMS) demonstrated potent
efficacy, and good safety and tolerability. Post-marketing experience, as well
as a third phase III trial (FREEDOMS II), also showed favorable results. More
selective S1P receptor agents-ponesimod (ACT128800), siponimod (BAF312),
ozanimod (RPC1063), ceralifimod (ONO-4641), GSK2018682, and MT-1303-are still in
relatively early stages of development, but phase I and II trials showed
promising efficacy and safety. However, these observations have yet to be
reproduced in phase III clinical trials. PURPOSE OF REVIEW: We review the most recent developments regarding the
targeting of molecules involved in the traffic of leukocytes for the treatment
of inflammatory bowel diseases (IBD).
RECENT FINDINGS: We discuss the most important findings of one published phase
II trial that targeted the β7 integrin (etrolizumab), two phase II trials that
targeted the α4β7 integrin ligand: mucosal addressin cell adhesion molecule 1
(MAdCAM-1, PF-00547659), a phase II trial targeting the chemokine IP-10 (CXCL10)
in Crohn's, and a phase II trial that targeted the sphingosine-1-phosphate
receptor-1: ozanimod in patients with ulcerative colitis.
SUMMARY: Targeting molecules involved in leukocyte traffic has recently become
an effective and well tolerated strategy for the treatment of IBD. Novel
approaches now not only target the integrins on the lymphocyte surface, but also
its endothelial ligand: MAdCAM-1. As with vedolizumab, antibodies against
MAdCAM-1 appear most effective in ulcerative colitis rather than in Crohn's.
Targeting chemokines or their receptors does not appear to have the same
efficacy as those that target the most stable integrin: immunoglobulin
superfamily interactions between the lymphocyte and endothelium. Preliminary
results also suggest that the sphingosine-1-phosphate pathway might also be
targeted therapeutically in IBD, no longer with parenterally administered
antibodies but with orally administered small molecules. The sphingosine-1-phosphate receptor-1 (S1P1) agonist ozanimod ameliorates
ulcerative colitis, yet its mechanism of action is unknown. Here, we examine the
cell subsets that express S1P1 in intestine using S1P1-eGFP mice, the regulation
of S1P1 expression in lymphocytes after administration of dextran sulfate sodium
(DSS), after colitis induced by transfer of CD4+CD45RBhi cells, and by crossing
a mouse with TNF-driven ileitis with S1P1-eGFP mice. We then assayed the
expression of enzymes that regulate intestinal S1P levels, and the effect of
FTY720 on lymphocyte behavior and S1P1 expression. We found that not only T and
B cells express S1P1, but also dendritic (DC) and endothelial cells.
Furthermore, chronic but not acute inflammatory signals increased S1P1
expression, while the enzymes that control tissue S1P levels in mice and humans
with inflammatory bowel disease (IBD) were uniformly dysregulated, favoring
synthesis over degradation. Finally, we observed that FTY720 reduced T-cell
velocity and induced S1P1 degradation and retention of Naïve but not effector T
cells. Our data demonstrate that chronic inflammation modulates S1P1 expression
and tissue S1P levels and suggests that the anti-inflammatory properties of S1PR
agonists might not be solely due to their lymphopenic effects, but also due to
potential effects on DC migration and vascular barrier function. BACKGROUND: Ozanimod (RPC1063) is an oral agonist of the sphingosine-1-phosphate
receptor subtypes 1 and 5 that induces peripheral lymphocyte sequestration,
potentially decreasing the number of activated lymphocytes circulating to the
gastrointestinal tract.
METHODS: We conducted a double-blind, placebo-controlled phase 2 trial of
ozanimod in 197 adults with moderate-to-severe ulcerative colitis. Patients were
randomly assigned, in a 1:1:1 ratio, to receive ozanimod at a dose of 0.5 mg or
1 mg or placebo daily for up to 32 weeks. The Mayo Clinic score was used to
measure disease activity on a scale from 0 to 12, with higher scores indicating
more severe disease; subscores range from 0 to 3, with higher scores indicating
more severe disease. The primary outcome was clinical remission (Mayo Clinic
score ≤2, with no subscore >1) at 8 weeks.
RESULTS: The primary outcome occurred in 16% of the patients who received 1 mg
of ozanimod and in 14% of those who received 0.5 mg of ozanimod, as compared
with 6% of those who received placebo (P=0.048 and P=0.14, respectively, for the
comparison of the two doses of ozanimod with placebo). Differences in the
primary outcome between the group that received 0.5 mg of ozanimod and the
placebo group were not significant; therefore, the hierarchical testing plan
deemed the analyses of secondary outcomes exploratory. Clinical response
(decrease in Mayo Clinic score of ≥3 points and ≥30% and decrease in
rectal-bleeding subscore of ≥1 point or a subscore ≤1) at 8 weeks occurred in
57% of those receiving 1 mg of ozanimod and 54% of those receiving 0.5 mg, as
compared with 37% of those receiving placebo. At week 32, the rate of clinical
remission was 21% in the group that received 1 mg of ozanimod, 26% in the group
that received 0.5 mg of ozanimod, and 6% in the group that received placebo; the
rate of clinical response was 51%, 35%, and 20%, respectively. At week 8,
absolute lymphocyte counts declined 49% from baseline in the group that received
1 mg of ozanimod and 32% from baseline in the group that received 0.5 mg. The
most common adverse events overall were anemia and headache.
CONCLUSIONS: In this preliminary trial, ozanimod at a daily dose of 1 mg
resulted in a slightly higher rate of clinical remission of ulcerative colitis
than placebo. The trial was not large enough or of sufficiently long duration to
establish clinical efficacy or assess safety. (Funded by Receptos; TOUCHSTONE
ClinicalTrials.gov number, NCT01647516.). The sphingosine-1-phosphate 1 receptor (S1P1R ) is expressed by lymphocytes,
dendritic cells, and vascular endothelial cells and plays a role in the
regulation of chronic inflammation and lymphocyte egress from peripheral
lymphoid organs. Ozanimod is an oral selective modulator of S1P1R and S1P5R
receptors in clinical development for the treatment of chronic immune-mediated,
inflammatory diseases. This first-in-human study characterized the safety,
pharmacokinetics (PK), and pharmacodynamics (PD) of ozanimod in 88 healthy
volunteers using a range of single and multiple doses (7 and 28 days) and a
dose-escalation regimen. Ozanimod was generally well tolerated up to a maximum
single dose of 3 mg and multiple doses of 2 mg/d, with no severe adverse events
(AEs) and no dose-limiting toxicities. The most common ozanimod-related AEs
included headache, somnolence, dizziness, nausea, and fatigue. Ozanimod
exhibited linear PK, high steady-state volume of distribution (73-101 L/kg),
moderate oral clearance (204-227 L/h), and an elimination half-life of
approximately 17 to 21 hours. Ozanimod produced a robust dose-dependent
reduction in total peripheral lymphocytes, with a median decrease of 65% to 68%
observed after 28 days of dosing at 1 and 1.5 mg/d, respectively. Ozanimod
selectivity affected lymphocyte subtypes, causing marked decreases in cells
expressing CCR7 and variable decreases in subsets lacking CCR7. A dose-dependent
negative chronotropic effect was observed following the first dose, with the
dose-escalation regimen attenuating the first-dose negative chronotropic effect.
Ozanimod safety, PK, and PD properties support the once-daily regimens under
clinical investigation. The past three decades have witnessed remarkable advances in our ability to
target specific elements of the immune and inflammatory response, fuelled by
advances in both biotechnology and disease knowledge. As well as providing
superior treatments for immune-mediated inflammatory diseases (IMIDs), such
therapies also offer unrivalled opportunities to study the underlying
immunopathological basis of these conditions.In this review, we explore recent
approaches to the treatment of IMIDs and the insights to pathobiology that they
provide. We review novel biologic agents targeting the T-helper 17 axis,
including therapies directed towards interleukin (IL)-17 (secukinumab,
ixekizumab, bimekizumab), IL-17R (brodalumab), IL-12/23p40 (ustekinumab,
briakinumab) and IL-23p19 (guselkumab, tildrakizumab, brazikumab, risankizumab,
mirikizumab). We also present an overview of biologics active against type I and
II interferons, including sifalumumab, rontalizumab, anifrolumab and
fontolizumab. Emerging strategies to interfere with cellular adhesion processes
involved in lymphocyte recruitment are discussed, including both integrin
blockade (natalizumab, vedolizumab, etrolizumab) and sphingosine-1-phosphate
receptor inhibition (fingolimod, ozanimod). We summarise the development and
recent application of Janus kinase (JAK) inhibitors in the treatment of IMIDs,
including first-generation pan-JAK inhibitors (tofacitinib, baricitinib,
ruxolitinib, peficitinib) and second-generation selective JAK inhibitors
(decernotinib, filgotinib, upadacitinib). New biologics targeting B-cells
(including ocrelizumab, veltuzumab, tabalumab and atacicept) and the development
of novel strategies for regulatory T-cell modulation (including low-dose IL-2
therapy and Tregitopes) are also discussed. Finally, we explore recent
biotechnological advances such as the development of bispecific antibodies
(ABT-122, COVA322), and their application to the treatment of IMIDs. Ozanimod is a novel, selective, oral sphingosine-1-phosphate (1 and 5) receptor
modulator in development for multiple sclerosis and inflammatory bowel disease.
This randomized, double-blind, placebo-controlled, positive-controlled,
parallel-group thorough QT study characterized the effects of ozanimod on
cardiac repolarization in healthy subjects. Eligible subjects were randomized to
1 of 2 groups: ozanimod (escalated from 0.25 to 2 mg over 14 days) or placebo
(for 14 days). A single dose of moxifloxacin 400 mg or placebo was administered
on days 2 and 17. The primary end point was the time-matched, placebo-corrected,
baseline-adjusted mean QTcF (ΔΔQTcF). A total of 113/124 (91.1%) subjects
completed the study. The upper limits of the 2-sided 90% confidence intervals
for ΔΔQTcF for both ozanimod 1 and 2 mg were below the 10-millisecond regulatory
threshold. No QTcF >480 milliseconds or postdose change in QTcF of
>60 milliseconds was observed. There was no evidence of a positive relationship
between concentrations of ozanimod and its active metabolites and ΔΔQTcF.
Although ozanimod blunted the observed diurnal increase in heart rate,
excursions below predose heart rates were no greater than with placebo. Results
demonstrate that ozanimod does not prolong the QTc interval or cause clinically
significant bradycardia, supporting ozanimod's evolving favorable cardiac safety
profile. Ozanimod (RPC1063) is an oral selective modulator of the sphingosine-1-phosphate
1 and 5 receptors under development for the treatment of relapsing multiple
sclerosis and inflammatory bowel disease. The effects of high-fat and low-fat
meals on the pharmacokinetics (PK) of a single oral dose of ozanimod were
evaluated in 24 healthy volunteers in a randomized, open-label crossover trial.
Each subject received a 1-mg dose of ozanimod hydrochloride under 3 meal
conditions (fasted, high-fat, and low-fat), each separated by 7 days. Mean
plasma concentration-time profiles for ozanimod and its active metabolites
(RP101988 [major], RP101075 [minor]) were similar under all 3 conditions.
Moreover, all PK parameters for ozanimod, RP101988, and RP101075 were similar
under the 3 meal conditions. The 90% confidence intervals (CIs) for the ratios
of geometric least-squares mean (fed/fasted) were within the equivalence limits
of 0.80 to 1.25 for area under the concentration-time curve from time 0 to
infinity (AUC0-∞ ) and maximum plasma concentration (Cmax ) for ozanimod,
RP101988, and RP101075, except for the high-fat effect on RP101075 Cmax (90%CI,
0.76-0.88). Given this lack of a food effect on the exposure of ozanimod and its
active metabolites, ozanimod can be taken without regard to meals. |
Describe swirl sign in intracerebral hemorrhage. | Swirl sign is described as areas of low attenuation, radiolucency or irregular density. It was previously described in epidural hematomas. In intracerebral hemorrhage swirl sign is associated with greater hematoma volume, unfavorable outcomes and greater mortality risk. | BACKGROUND: Swirl sign has previously been described in epidural hematomas as
areas of low attenuation, radiolucency or irregular density. The aims of this
study were to describe swirl sign in ICH, study its prevalence, study the
reliability of the subjective evaluation on computed tomography (CT), and to
explore its prognostic value.
METHODS: CTs of 203 patients with ICH were retrospectively evaluated for the
presence of swirl sign. Association between swirl sign and different clinical
and radiological variables was studied.
RESULTS: Inter- and intraobserver agreement with regard to the occurrence of
swirl sign was substantial (К 0.80) and almost perfect (К 0.87), respectively.
Swirl sign was found in 30% of the study population. 61% of patients with swirl
sign were dead at one month compared with 21% of those with no swirl sign (p <
0.001). Only 19% of patients with swirl sign exhibited favorable outcome at
three months compared with 53% of those with no swirl sign (p < 0.001). Patients
with swirl sign exhibited larger ICHs with average ICH-volume 52 ± 50 ml (median
42 ml) compared with 15 ± 25 ml (median 6) in patients whose CT did not show
swirl sign (p < 0.001). Swirl sign was independent predictor of death at one
month (p = 0.03; adjusted odds ratio 2.6, 95% CI 1.1 - 6), and functional
outcome at three months (p = 0.045; adjusted odds ratio 2.6, 95% CI 1.02 - 6.5).
CONCLUSIONS: As swirl sign showed to be an ominous sign, we recommend
identification of this sign in cases of ICHs. The aim of the study was to identify the predictors of brain death (BD) upon
admission to the intensive care unit (ICU) of comatose patients with spontaneous
intracerebral hemorrhage (ICH). Patients admitted in our ICU from 2002 to 2010
for spontaneous ICH and placed under mechanical ventilation were retrospectively
analyzed. Of the 72 patients, 49% evolved to BD, 39% died after withdrawal of
life support, and 12% were discharged alive. The most discriminating
characteristics to predict BD were included in two models; Model 1 contained ≥3
abolished brainstem responses [adjusted odds ratios (OR) = 8.4 (2.4, 29.1)] and
the swirl sign on the baseline CT-scan [adjusted OR = 5.0 (1.6, 15.9)] and Model
2 addressed the abolition of corneal reflexes [unilateral/bilateral: adjusted
OR = 4.2 (0.9, 20.1)/8.8 (2.4, 32.3)] and the swirl sign on the baseline CT-scan
[adjusted OR = 6.2 (1.9, 20.0)]. Two scores predicting BD were created
(sensitivity: 0.89 and 0.88, specificity: 0.68 and 0.65). Risk of evolution
toward BD was classified as low (corneal reflexes present and no swirl sign),
high (≥1 corneal reflexes abolished and swirl sign), and intermediate. Simple
signs at ICU admission can predict BD in comatose patients with ICH and could
increase the potential for organ donation. The swirl sign is identified as a small area of low attenuation within an
intracranial hyperattenuating clot on non-enhanced computed tomography (CT)
scans of the brain, which represents active bleeding. The purpose of this study
was to evaluate the incidence of the swirl sign among patients with acute
epidural hematoma (AEDH) and to identify its prognostic value and impact on
surgical treatment. A retrospective review was performed of patients with a
diagnosis of traumatic EDH by CT scan who were surgically treated at the
Department of Neurosurgery of the First People's Hospital of Jingmen between
January 2010 and January 2014. Patients with combined or open craniocerebral
injuries and those who did not undergo surgical treatment were excluded. Of the
147 patients evaluated, 21 (14%) exhibited the swirl sign on non-enhanced CT
scans of the brain. Univariate analysis revealed a significant correlation
between the occurrence of the swirl sign and preoperative Glasgow coma scale
scores, preoperative mydriasis, time from injury to CT scan, and intraoperative
hematoma volume. Compared with patients without this sign, those exhibiting the
swirl sign had a higher mortality rate (24 vs. 6%, respectively; P = 0.028) and
a worse outcome (Glasgow Outcome Scale score ≤ 3: 38 vs. 15%, respectively;
P = 0.027) at 3 months. An adjusted analysis showed that the occurrence of the
swirl sign was an independent predictor of poor outcome (death: odds ratio
(OR) = 4.61; 95% confidence interval (CI): 1.34-15.82; P < 0.05; 3-month Glasgow
Outcome Scale score ≤ 3: OR = 3.47; 95% CI: 1.27-9.49; P < 0.05). In conclusion,
the occurrence of the swirl sign on the head CT scan of patients with AEDH was
found to be significantly associated with poor outcome. Therefore, early
identification of this sign and aggressive management with early surgical
evacuation is crucial for improving patient outcome. |
What is Morgellons disease? | It is a skin condition in which individuals have skin lesions that contain some kind of fibers. Patients often complain of bugs crawling under their skin. The disease is of unknown origin and may be psychosomatic, however recent evidence indicates it could be transmitted by a tick. | Morgellons disease is a controversial and poorly defined symptom cluster of skin
lesions and somatic symptoms, most notably 'fibers' in the skin. Because of
widespread coverage in the media and on the Internet, there are an increasing
number of patients presenting to dermatologists. We present three patients who
believed that they had fibers in their skin. We offer a discussion of delusions
of parasitosis to demonstrate similarities between these conditions. It has been
suggested by a limited number of healthcare providers that an unknown infectious
agent underlies this symptom complex yet no available evidence supports this
assertion. Laboratory values that would be reflective of an infectious process
(e.g. elevated white blood cells, sedimentation rate, C reactive protein) are
routinely normal and biopsies often reflect only nonspecific findings such as
acute and chronic inflammation with erosion or ulceration. Patients with
Morgellons disease generally lack insight into their disease and reject the need
for psychiatric help. The goal is to build trust and refrain from minimizing
what the patient experiences. Attentive examination of the patient's skin and
fragments they present is necessary to rule out a true underlying pathologic
process and to establish a trusting relationship. A supportive,
non-confrontational approach is ideal. The patient is best treated by a team of
practitioners of several specialties, including dermatologists, psychiatrists,
and counselors. Morgellons Disease is a condition involving painful skin lesions, fibrous
growths protruding from the skin, and subcutaneous stinging and burning
sensations, along with symptoms of anxiety, depression, fatigue, and memory and
attention deficits. The etiological and physiological bases of these symptoms
are unclear, making the diagnosis controversial and challenging to treat. There
are currently no established treatments for Morgellons Disease. The following
case example depicts treatment of a woman with Morgellons Disease using
hypnotherapy. Data from this case example suggest that hypnotherapy is a
promising intervention for the physical and psychological symptoms associated
with Morgellons Disease. BACKGROUND: Morgellons disease is a controversial illness in which patients
complain of stinging, burning, and biting sensations under the skin. Unusual
subcutaneous fibers are the unique objective finding. The etiology of Morgellons
disease is unknown, and diagnostic criteria have yet to be established. Our goal
was to identify prevalent symptoms in patients with clinically confirmed
subcutaneous fibers in order to develop a case definition for Morgellons
disease.
METHODS: Patients with subcutaneous fibers observed on physical examination
(designated as the fiber group) were evaluated using a data extraction tool that
measured clinical and demographic characteristics. The prevalence of symptoms
common to the fiber group was then compared with the prevalence of these
symptoms in patients with Lyme disease and no complaints of skin fibers.
RESULTS: The fiber group consisted of 122 patients. Significant findings in this
group were an association with tick-borne diseases and hypothyroidism, high
numbers from two states (Texas and California), high prevalence in middle-aged
Caucasian women, and an increased prevalence of smoking and substance abuse.
Although depression was noted in 29% of the fiber patients, pre-existing
delusional disease was not reported. After adjusting for nonspecific symptoms,
the most common symptoms reported in the fiber group were: crawling sensations
under the skin; spontaneously appearing, slow-healing lesions; hyperpigmented
scars when lesions heal; intense pruritus; seed-like objects, black specks, or
"fuzz balls" in lesions or on intact skin; fine, thread-like fibers of varying
colors in lesions and intact skin; lesions containing thick, tough, translucent
fibers that are highly resistant to extraction; and a sensation of something
trying to penetrate the skin from the inside out.
CONCLUSIONS: This study of the largest clinical cohort reported to date provides
the basis for an accurate and clinically useful case definition for Morgellons
disease. Morgellons is a medically contested diagnosis with foremost dermatological
symptoms. Patients experience fibers emerging from the skin, together with a
range of other somatic, psychiatric, and neurological complaints. Within the
medical community, it is generally held to be a variation of delusional
parasitosis/delusional infestation, which is usually treated with
antipsychotics. Little attention has been paid in the literature to the ethical
aspects of treating patients with Morgellons disease. The communicative
strategies suggested in the literature display significant ethical issues,
primarily the use of therapeutic privilege, i.e. withholding information from
the patient. Since this limits patient autonomy, that approach is ethically
problematic. Instead, the physician has an ethical obligation to respect the
patient's autonomy, provide full information, and seek consent before initiating
a psychiatric referral. Morgellons disease is characterized by complaints of uncomfortable skin
sensations and fibers emanating from nonhealing skin lesions. Morgellons disease
is well-known in the dermatology and psychiatry literature, where it is
typically considered a subtype of delusional parasitosis, but it has not yet
been described in the ophthalmology literature. A patient with self-reported
Morgellons disease is presented, who was referred for evaluation of left lower
eyelid ectropion. She reported that her skin was infested with fibers that were
"trying to get down into the eyelid." On examination, she had ectropion of the
left lower eyelid, broken cilia, and an ulcerated left upper eyelid lesion
concerning for carcinoma. Biopsy of the lesion was consistent with excoriation.
Treatment of her ectropion was deferred out of concern for wound dehiscence,
given the patient's aggressive excoriation behavior. This case is presented to
make the ophthalmologist aware of this disorder and to highlight the appropriate
clinical management. BACKGROUND: Morgellons disease (MD) is a complex skin disorder characterized by
ulcerating lesions that have protruding or embedded filaments. Many clinicians
refer to this condition as delusional parasitosis or delusional infestation and
consider the filaments to be introduced textile fibers. In contrast, recent
studies indicate that MD is a true somatic illness associated with tickborne
infection, that the filaments are keratin and collagen in composition and that
they result from proliferation and activation of keratinocytes and fibroblasts
in the skin. Previously, spirochetes have been detected in the dermatological
specimens from four MD patients, thus providing evidence of an infectious
process.
METHODS & RESULTS: Based on culture, histology, immunohistochemistry, electron
microscopy and molecular testing, we present corroborating evidence of
spirochetal infection in a larger group of 25 MD patients. Irrespective of Lyme
serological reactivity, all patients in our study group demonstrated
histological evidence of epithelial spirochetal infection. Strength of evidence
based on other testing varied among patients. Spirochetes identified as Borrelia
strains by polymerase chain reaction (PCR) and/or in-situ DNA hybridization were
detected in 24/25 of our study patients. Skin cultures containing Borrelia
spirochetes were obtained from four patients, thus demonstrating that the
organisms present in dermatological specimens were viable. Spirochetes
identified by PCR as Borrelia burgdorferi were cultured from blood in seven
patients and from vaginal secretions in three patients, demonstrating systemic
infection. Based on these observations, a clinical classification system for MD
is proposed.
CONCLUSIONS: Our study using multiple detection methods confirms that MD is a
true somatic illness associated with Borrelia spirochetes that cause Lyme
disease. Further studies are needed to determine the optimal treatment for this
spirochete-associated dermopathy. Morgellons disease is an infrequent syndromic condition, that typically affects
middle-aged white women, characterized by crawling sensations on and under the
skin, associated with itchy rashes, stinging sores, fiber-like filaments
emerging from the sores, severe fatigue, concentrating difficulty, and memory
loss. The scientific community is prone to believe that Morgellons is the
manifestation of various psychiatric syndromes (Munchausen, Munchausen by proxy,
Ekbom, Wittmaack-Ekbom). Up until now, no investigative science-based evidence
about its psychogenesis has ever been provided. In order to close this gap, we
have analyzed the filaments extracted from the skin lesions of a 49-year-old
Caucasian female patient, by using a Field Emission Gun-Environmental Electron
Scanning Microscope equipped with an X-ray microprobe, for the chemico-elemental
characterization of the filaments, comparing them with those collected during a
detailed indoor investigation, with careful air monitoring, in her apartment.
Our results prove the self-introduction under the epidermis of environmental
filaments. For the first time in the literature, we have scientifically
demonstrated the self-induced nature of Morgellons disease, thereby wiping out
fanciful theories about its etiopathogenesis. Morgellons disease (MD) is a dermopathy characterized by multicolored filaments
that lie under, are embedded in, or project from skin. Although MD was initially
considered to be a delusional disorder, recent studies have demonstrated that
the dermopathy is associated with tickborne infection, that the filaments are
composed of keratin and collagen, and that they result from proliferation of
keratinocytes and fibroblasts in epithelial tissue. Culture, histopathological
and molecular evidence of spirochetal infection associated with MD has been
presented in several published studies using a variety of techniques.
Spirochetes genetically identified as Borrelia burgdorferi sensu stricto
predominate as the infective agent in most of the Morgellons skin specimens
studied so far. Other species of Borrelia including Borrelia garinii, Borrelia
miyamotoi, and Borrelia hermsii have also been detected in skin specimens taken
from MD patients. The optimal treatment for MD remains to be determined. Morgellons disease is a rare disease with unknown etiology. Herein, we report
the first case of Morgellons disease in Korea. A 30-year-old woman presented
with a 2-month history of pruritic erythematous patches and erosions on the
arms, hands, and chin. She insisted that she had fiber-like materials under her
skin, which she had observed through a magnifying device. We performed skin
biopsy, and observed a fiber extruding from the dermal side of the specimen.
Histopathological examination showed only mild lymphocytic infiltration, and
failed to reveal evidence of any microorganism. The polymerase chain reaction
for Borrelia burgdorferi was negative in her serum. BACKGROUND: In recent years, there has been a reported increase in affliction of
the skin with small fibres or other particles. The condition has been referred
to as Morgellons disease. Patients present with stinging, burning or crawling
sensations of the skin, with perceived extrusion of iimate material alongside
fatigue and other systemic symptoms. Sufferers often experience significant
morbidity and reduction in quality of life.
OBJECTIVES: We aimed to explore the various clinical presentations, management
strategies and outcomes employed to treat this condition in our patients.
METHODS: We conducted a retrospective case notes review of 35 patients referred
to our multidisciplinary psycho-dermatology clinic at the Royal London Hospital
between January 2004 and January 2017.
RESULTS: The majority of patients were women (25) 71.4%, with a mean age of
54.6 years (26-80 years). Most (26) 74.2% were living alone. The average
duration of illness prior to presentation was 3.8 years (4 months-20 years).
Many patients had perceived precipitating factors (54.2%) and often
self-diagnosed (28.5%). Psychiatric co-morbidities included 42.8% with
depressive symptoms and 25.7% with anxiety. Substance misuse was elicited in
five patients (14%). Management of patients included both the treatment of skin
disease and psychosocial co-morbidities. Out of the 35 patients who attended
(14) 40% cleared or showed significant improvement. Sixteen (45.7%) patients
were stable and under review. One patient declined treatment and three did not
attend review. One patient died from disease unrelated to her skin condition.
CONCLUSIONS: Morgellons disease is a condition, which is widely discussed on the
internet and patients often self-diagnose. The course of the disease can be
chronic and debilitating. For a positive outcome, it is important that a strong
physican-patient relationship is cultivated. As demonstrated in this case
series, managing patients holistically in an integrated multidisciplinary
dermatology setting helps achieve positive outcomes. Approximately half of all patients presenting to dermatologists exhibit signs
and symptoms of psychiatric conditions that are either primary or secondary to
cutaneous disease. Because patients typically resist psychiatric consult,
dermatologists often are on the front line in evaluating and treating these
patients. Accordingly, distinguishing the specific underlying or resulting
psychiatric condition is essential for effective treatment. The etiology,
epidemiology, clinical presentation, diagnosis, and first-line treatment of
specific primary psychiatric causes of dermatologic conditions, including
delusional infestation, Morgellons syndrome, olfactory reference syndrome, body
dysmorphic disorder, excoriation disorder, trichotillomania, and dermatitis
artefacta are discussed here, followed by a discussion of the recommended
treatment approach with an overview of the different first-line therapies
discussed in this review, specifically cognitive behavioral therapy, atypical
antipsychotics, selective serotonin reuptake inhibitors, and tricyclic
antidepressants. Included is a guide for dermatologists to use while prescribing
these medications. |
What is Cellbase? | CellBase, a comprehensive collection of RESTful web services for retrieving relevant biological information from heterogeneous sources. | During the past years, the advances in high-throughput technologies have
produced an unprecedented growth in the number and size of repositories and
databases storing relevant biological data. Today, there is more biological
information than ever but, unfortunately, the current status of many of these
repositories is far from being optimal. Some of the most common problems are
that the information is spread out in many small databases; frequently there are
different standards among repositories and some databases are no longer
supported or they contain too specific and unconnected information. In addition,
data size is increasingly becoming an obstacle when accessing or storing
biological data. All these issues make very difficult to extract and integrate
information from different sources, to analyze experiments or to access and
query this information in a programmatic way. CellBase provides a solution to
the growing necessity of integration by easing the access to biological data.
CellBase implements a set of RESTful web services that query a centralized
database containing the most relevant biological data sources. The database is
hosted in our servers and is regularly updated. CellBase documentation can be
found at http://docs.bioinfo.cipf.es/projects/cellbase. |
What is the asosciation between the eustachian tube and the palatine muscle of the uvula? | Palatal musculature is known to be responsible for the active opening of the eustachian tube. | |
Are Conserved Nonexonic Elements (CNEEs) important in phylogenomics research? | Yes. Conserved Nonexonic Elements (CNEEs) appear to be promising as phylogenomic markers, yielding phylogenetic resolution as high as for UCEs and introns but with fewer gaps, less ambiguity in alignments and with patterns of nucleotide substitution more consistent with the assumptions of commonly used methods of phylogenetic analysis. | Noncoding markers have a particular appeal as tools for phylogenomic analysis
because, at least in vertebrates, they appear less subject to strong variation
in GC content among lineages. Thus far, ultraconserved elements (UCEs) and
introns have been the most widely used noncoding markers. Here we analyze and
study the evolutionary properties of a new type of noncoding marker, conserved
nonexonic elements (CNEEs), which consists of noncoding elements that are
estimated to evolve slower than the neutral rate across a set of species.
Although they often include UCEs, CNEEs are distinct from UCEs because they are
not ultraconserved, and, most importantly, the core region alone is analyzed,
rather than both the core and its flanking regions. Using a data set of 16 birds
plus an alligator outgroup, and ∼3600-∼3800 loci per marker type, we found that
although CNEEs were less variable than bioinformatically derived UCEs or introns
and in some cases exhibited a slower approach to branch resolution as determined
by phylogenomic subsampling, the quality of CNEE alignments was superior to
those of the other markers, with fewer gaps and missing species. Phylogenetic
resolution using coalescent approaches was comparable among the three marker
types, with most nodes being fully and congruently resolved. Comparison of
phylogenetic results across the three marker types indicated that one branch,
the sister group to the passerine + falcon clade, was resolved differently and
with moderate (>70%) bootstrap support between CNEEs and UCEs or introns.
Overall, CNEEs appear to be promising as phylogenomic markers, yielding
phylogenetic resolution as high as for UCEs and introns but with fewer gaps,
less ambiguity in alignments and with patterns of nucleotide substitution more
consistent with the assumptions of commonly used methods of phylogenetic
analysis. |
What is the definition of General Regulatory Factors (GRFs)? | GRFs, which bind to sites scattered throughout the genome within promoters, would not only play a key role in regulating gene expression but also partition the genome in functionally independent domains. | Insulators are sequences that uncouple adjacent chromosome domains. Here we have
shown that Saccharomyces cerevisiae Rap1p and Abf1p proteins are endowed with a
potent insulating capacity. Insulating domains in Rap1p coincide with previously
described transcription activation domains, whereas four adjacent subdomains
spanning the whole of the Abf1p C terminus (440-731) were found to display
autonomous insulating capacity. That both Rap1p and Abf1p silencing domains
either contain or largely overlap with an insulating domain suggests that
insulation conveys some undefined chromosome organization capacity that also
contributes a function in silencing. Together with Reb1p and Tbf1p, previously
involved in the activity of Saccharomyces cerevisiae subtelomeric insulators,
insulating potential emerges as a supplementary common property of General
Regulatory Factors (GRFs). Thus GRFs, which bind to sites scattered throughout
the genome within promoters, would not only play a key role in regulating gene
expression but also partition the genome in functionally independent domains. Gene-regulation functions (GRF) provide a unique characteristic of a
cis-regulatory module (CRM), relating the concentrations of transcription
factors (input) to the promoter activities (output). The challenge is to predict
GRFs from the sequence. Here we systematically consider the lysogeny-lysis CRMs
of different temperate bacteriophages such as the Lactobacillus casei phage A2,
Escherichia coli phages lambda, and 186 and Lactococcal phage TP901-1. This
study allowed explaining a recent experimental puzzle on the role of Cro protein
in the lambda switch. Several general conclusions have been drawn: 1),
long-range interactions, multilayer assembly and DNA looping may lead to complex
GRFs that cannot be described by linear functions of binding site occupancies;
2), in general, GRFs cannot be described by the Boolean logic, whereas a
three-state non-Boolean logic suffices for the studied examples; 3), studied
CRMs of the intact phages seemed to have a similar GRF topology (the number of
plateaus and peaks corresponding to different expression regimes); we
hypothesize that functionally equivalent CRMs might have topologically
equivalent GRFs for a larger class of genetic systems; and 4) within a given GRF
class, a set of mechanistic-to-mathematical transformations has been identified,
which allows shaping the GRF before carrying out a system-level analysis. The packaging of eukaryotic genomes into nuclesomes plays critical roles in
chromatin organization and gene regulation. Studies in Saccharomyces cerevisiae
indicate that nucleosome occupancy is partially encoded by intrinsic
antinucleosomal DNA sequences, such as poly(A) sequences, as well as by binding
sites for trans-acting factors that can evict nucleosomes, such as Reb1 and the
Rsc3/30 complex. Here, we use genome-wide nucleosome occupancy maps in 13
Ascomycota fungi to discover large-scale evolutionary reprogramming of both
intrinsic and trans determits of chromatin structure. We find that poly(G)s
act as intrinsic antinucleosomal sequences, comparable to the known function of
poly(A)s, but that the abundance of poly(G)s has diverged greatly between
species, obscuring their antinucleosomal effect in low-poly(G) species such as
S. cerevisiae. We also develop a computational method that uses nucleosome
occupancy maps for discovering trans-acting general regulatory factor (GRF)
binding sites. Our approach reveals that the specific sequences bound by GRFs
have diverged substantially across evolution, corresponding to a number of major
evolutionary transitions in the repertoire of GRFs. We experimentally validate a
proposed evolutionary transition from Cbf1 as a major GRF in pre-whole-genome
duplication (WGD) yeasts to Reb1 in post-WGD yeasts. We further show that the
mating type switch-activating protein Sap1 is a GRF in S. pombe, demonstrating
the general applicability of our approach. Our results reveal that the
underlying mechanisms that determine in vivo chromatin organization have
diverged and that comparative genomics can help discover new determits of
chromatin organization. The microRNA (miRNA) miR396 regulates GROWTH-REGULATING FACTORs (GRFs), a plant
specific family of transcription factors. Overexpression of miR396 causes a
decrease in the GRFs that has been shown to affect cell proliferation in the
meristem and developing leaves. To bring further insights into the function of
the miR396 regulatory network we performed a mutant enhancer screen of a stable
Arabidopsis transgenic line expressing 35S:miR396b, which has a reduction in
leaf size. From this screen we recovered several mutants enhancing this
phenotype and displaying organs with lotus- or needle-like shape. Analysis of
these plants revealed mutations in as2 and rdr6. While 35S:miR396b in an as2
context generated organs with lotus-like shape, the overexpression of the miRNA
in an rdr6 mutant background caused more important developmental defects,
including pin-like organs and lobed leaves. Combination of miR396
overexpressors, and rdr6 and as2 mutants show additional organ defects,
suggesting that the three pathways act in concert. Genetic interactions during
leaf development were observed in a similar way between miR396 overexpression
and mutants in RDR6, SGS3 or AGO7, which are known to participate in
trans-acting siRNA (ta-siRNA) biogenesis. Furthermore, we found that miR396 can
cause lotus- and pin-like organs per se, once a certain expression threshold is
overcome. In good agreement, mutants accumulating high levels of TCP4, which
induces miR396, interacted with the AS1/AS2 pathway to generate lotus-like
organs. The results indicate that the miR396 regulatory network and the ta-siRNA
biogenesis pathway synergistically interact during leaf development and
morphogenesis. Deciphering the molecular basis of how modern human phenotypes have evolved is
one of the most fascinating challenges in biology. Here, we will focus on the
roles of gene regulatory factors (GRFs), in particular transcription factors
(TFs) and long non-coding RNAs (lncRNAs) during human evolution. We will present
examples of TFs and lncRNAs that have changed or show signs of positive
selection in humans compared to chimpanzees, in modern humans compared to
archaic humans, or within modern human populations. On the basis of current
knowledge about the functions of these GRF genes, we speculate that they have
been involved in speciation as well as in shaping phenotypes such as brain
functions, skeletal morphology, and metabolic processes. RNA polymerase II (Pol II) transcription termination by the Nrd1p-Nab3p-Sen1p
(NNS) pathway is critical for the production of stable noncoding RNAs and the
control of pervasive transcription in Saccharomyces cerevisiae To uncover
determits of NNS termination, we mapped the 3'-ends of NNS-terminated
transcripts genome-wide. We found that nucleosomes and specific DNA-binding
proteins, including the general regulatory factors (GRFs) Reb1p, Rap1p, and
Abf1p, and Pol III transcription factors enhance the efficiency of NNS
termination by physically blocking Pol II progression. The same DNA-bound
factors that promote NNS termination were shown previously to define the 3'-ends
of Okazaki fragments synthesized by Pol δ during DNA replication. Reduced
binding of these factors results in defective NNS termination and Pol II
readthrough. Furthermore, inactivating NNS enables Pol II elongation through
these roadblocks, demonstrating that effective Pol II termination depends on a
synergy between the NNS machinery and obstacles in chromatin. Consistent with
this finding, loci exhibiting Pol II readthrough at GRF binding sites are
depleted for upstream NNS signals. Overall, these results underscore how RNA
termination signals influence the behavior of Pol II at chromatin obstacles, and
establish that common genomic elements define boundaries for both DNA and RNA
synthesis machineries. Eukaryotic promoters generally contain nucleosome depleted regions near their
transcription start sites. In the model organism Saccharomyces cerevisiae, these
regions are adjacent to binding sites for general regulatory transcription
factors, and the Reb1 protein is commonly bound to promoter DNA near such
regions. The yeast TFC6 promoter is a unique RNA polymerase II promoter in that
it is autoregulated by its own gene product Tfc6p, which is part of the RNA
polymerase III transcription factor complex TFIIIC. We previously demonstrated
that mutation of a potential Reb1 binding site adjacent to the TFIIIC binding
site in the TFC6 promoter modestly reduces transcript levels, but leads to a
severe decrease in Tfc6 protein levels due to an upstream shift in the TFC6
transcription start site. Here we confirm that Reb1p indeed binds to the TFC6
promoter, and is important for proper transcription start site selection and
protein expression. Interestingly, loss of Reb1p association at this site has a
similar effect on the adjacent divergently transcribed ESC2 promoter, resulting
in a significant increase of 5'-extended ESC2 transcripts and reduction of Esc2
protein levels. This altered divergent transcription may be the result of
changes in nucleosome positioning at this locus in the absence of Reb1p binding.
We speculate that an important function of general regulatory factors such as
Reb1p is to establish and maintain proper transcription start sites at
promoters, and that when binding of such factors is compromised, resulting
effects on mRNA translation may be an underappreciated aspect of gene regulation
studies. In Saccharomyces cerevisiae, a group of more than 200 co-regulated genes (Ribi
genes) is involved in ribosome biogenesis. This regulon has recently been shown
to rely on a small set of transcriptional regulators (mainly Abf1, but also
Reb1, Tbf1 and Rap1) previously referred to as general regulatory factors (GRFs)
because of their widespread binding and action at many promoters and other
specialized genomic regions. Intriguingly, Abf1 binding to Ribi genes is
differentially modulated in response to distinct nutrition signaling pathways.
Such a dynamic promoter association has the potential to orchestrate both
activation and repression of Ribi genes in synergy with neighboring regulatory
sites and through the functional interplay of histone acetyltransferases and
deacetylases. |
Which virus can be diagnosed with the monospot test? | Epstein-Barr virus (EBV) can be detected with the monospot test. EBV is a highly prevalent virus, transmitted via saliva, which often causes asymptomatic infection in children but frequently results in infectious mononucleosis in adolescents. | Infectious Mononucleosis (IM), a benign lymphoproliferative disease, is the best
known clinical syndrome caused by Epstein-Barr Virus (EBV). It usually resolves
over a period of weeks or months without sequelae but may occasionally be
complicated by a wide variety of neurologic, hematologic, hepatic, respiratory,
and psychological complications. In this report we describe a patient with acute
hepatitis following EBV-IM in a previously healthy woman. A 26-year-old woman
who presented with fever, generalized weakness, nausea, sore throat, yellowing
of skin, and a generalized skin rash was admitted to our clinic. Tonsillar
enlargement, pharyngeal erythema, palatal petechiae, lymphadenopathy, and
jaundice were noted. Significant atypical lymphocytes ( > 10%) were seen on the
peripheral blood smear. Liver function tests such as ALT: 303 U/L, AST: 172 U/L,
ALP: 193 U/L and total bilirubin: 7.3 mg/dl were elevated. Serological tests for
EBV infection were consistent with acute infection (EBV virus capsid antigen was
reactive with IgM and IgG antibodies). The Monospot test was also positive. On
the seventh day, liver function tests and bilirubin had risen to peak level and
platelets were decreased. The patient was managed supportively and her critical
condition improved and was finally stabilized. Although the prognosis for IM is
very favorable, a variety of acute complications may occur. BACKGROUND: Infection with Epstein-Barr virus (EBV) is almost ubiquitous in
humans and generally occurs at two ages: infantile, which is usually
asymptomatic and associated with poorer socioeconomic conditions, and
adolescent, which causes infectious mononucleosis (IM) in ~25% cases. The
determits of whether the infection causes IM remain uncertain. We aimed to
evaluate seasonality and temporal trends in IM.
METHODS: Data from all Monospot tests, used as a marker for IM, were collected
from the Grampian population over 16 years.
RESULTS: Positive Monospot test results peaked at 17 years in females and 19 in
males. Females had 16% more diagnoses, although 55% more tests. IM was ~38% more
common in winter than summer. The annual rate of positive tests decreased
progressively over the study period, from 174/100 000 (95% CI 171-178) in 1997
to 67/100 000 (95% CI 65-69) in 2012.
CONCLUSIONS: IM appears to be decreasing in incidence, which may be caused by
changing environmental influences on immune systems. One such factor may be
exposure to sunlight.Words 168.
FUNDING: The Medical Research Council and NHS Grampian-MS endowments. A 29-year-old man presented with sudden left-sided pleuritic chest pain on a
background of sore throat during the preceding week. On examination he had
tender cervical lymphadenopathy, he was tachycardic and had a 24 mm Hg blood
pressure difference between the left and right arms. Bloods revealed deranged
liver function tests and a lymphocytosis. His D-dimer was raised, hence he was
treated for presumed pulmonary embolism before imaging was available. Monospot
test was positive. He subsequently had both a CT pulmonary angiogram and a CT
angiogram of the aorta to exclude pulmonary embolism and aortic dissection. The
CT revealed splenomegaly with a large subdiaphragmatic haematoma secondary to
splenic rupture. This had likely caused referred pain through diaphragmatic
irritation. He was taken to theatre for urgent splenectomy. The unifying
diagnosis was infectious mononucleosis complicated by spontaneous splenic
rupture secondary to Epstein-Barr virus infection. A 75-year-old woman presented with altered mental status, septic picture, and
influenza-like symptoms. Initial investigations revealed atypical lymphocytosis,
thrombocytopenia, elevated liver enzymes, and a positive monospot test result.
Further investigation showed the Epstein-Barr virus viral capsid antibody
IgM/IgG and Epstein-Barr virus DNA by polymerase chain reaction to be negative;
however, interestingly her cytomegalovirus (CMV) IgM and IgG were positive,
suggesting that her mononucleosis-like syndrome was due to acute CMV infection.
Herein, we report the first case of a heterophile-positive mononucleosis
syndrome caused by acute CMV infection in an elderly immunocompetent woman. This
case conveys that monospot test can yield false-positive result in the setting
of acute CMV infection. Epstein-Barr virus (EBV) is a highly prevalent virus, transmitted via saliva,
which often causes asymptomatic infection in children but frequently results in
infectious mononucleosis in adolescents. Heterophile antibody tests, including
the Monospot test, are red cell or latex agglutination assays, which detect
antired cell antibodies produced as part of a polyclonal antibody response
occurring during EBV infection. Heterophile antibody tests are rapid, cheap and
specific tests that can be performed from the onset of symptoms of infectious
mononucleosis. In adolescents, heterophile antibody tests have high specificity
and sensitivity in the diagnosis of primary acute EBV infection. However, the
tests have low sensitivity and low negative predictive value in young children
and are not useful under the age of 4. Heterophile tests may be positive in
other viral infections, autoimmune disease and haematological maligcies, but
do not appear to be positive in primary bacterial infection. Virus-specific
serology is required in children under the age of 4 or if an older child is
heterophile negative. Virus-specific serology allows diagnosis and the pattern
of positivity and negativity enables the clinician to stage the EBV infection.
Virus-specific serology appears to have better sensitivity in young children,
but there is cross-reaction with other herpesvirus infections, a longer
turnaround time and it is more expensive to perform. Further research is needed
to establish which children benefit from and hence require testing for
heterophile antibodies, the cost-effectiveness of EBV investigations and whether
heterophile titres have predictive value for the severity of infection and the
likelihood of complications. Henoch-Schönlein purpura (HSP) is the most common form of childhood vasculitis.
Various viral and bacterial infections, drugs, vaccines, food allergy and even
insect bites have been considered as triggering factors in pathogenesis of HSP.
Epstein-Barr virus (EBV) infection, which is associated with HSP, have been
rarely reported. Herein we present HSP patient possibly caused by EBV infection.
A 8-year old boy was admitted to our department with fever, rashes on legs and
arms and intermittent mild abdominal pain. Multiple purpuric rashes were on his
extremities, abdomen and buttock. Laboratory investigations revealed that
monospot test was positive, EBV serology tests; Anti-EA-D Ig G: 3+, Anti-VCA
gp125 Ig G: 3+, Anti-VCA p19 Ig M: 2+, Anti EBNA-1 Ig M: negative, Anti EBNA-1
Ig M: negative, Anti EBNA-1 Ig G: negative. The patient was interpreted as the
primary active acute EBV infection. A skin biopsy showed leucocytoclastic
vasculitis. The other viral and bacterial investigations were negative. The
patient was diagnosed as HSP vasculitis according to EULAR criteria and treated
with intravenous hydration and ibuprofen. He was discharged after 15 days with
normal laboratory findings and physical examination. We think that EBV infection
may be stimulant factor for autoimmune reactions and may cause HSP vasculitis.
Hence, it may be useful to investigate the EBV infection in etiology of HSP
cases. |
Why is the Fyn kinase considered a promising therapeutic target for Alzheimer's Disease? | Fyn is an attractive target for AD therapeutics, not only based on its activation by Aβ via cellular prion protein but also due to its known interaction with tau, uniquely linking the two key pathologies in AD. | The past decade has brought tremendous progress in unraveling the
pathophysiology of Alzheimer's disease (AD). While increasingly sophisticated
immunotherapy targeting soluble and aggregated brain amyloid-beta (Aβ) continues
to dominate clinical research in AD, a deeper understanding of Aβ physiology has
led to the recognition of distinct neuronal signaling pathways linking Aβ to
synaptotoxicity and neurodegeneration and to new targets for therapeutic
intervention. Identifying specific signaling pathways involving Aβ has allowed
for the development of more precise therapeutic interventions targeting the most
relevant molecular mechanisms leading to AD. In this review, I highlight the
discovery of cellular prion protein as a high-affinity receptor for Aβ
oligomers, and the downstream signaling pathway elucidated to date, converging
on nonreceptor tyrosine kinase Fyn. I discuss preclinical studies targeting Fyn
as a therapeutic intervention in AD and our recent experience with the safety,
tolerability, and cerebrospinal fluid penetration of the Src family kinase
inhibitor saracatinib in patients with AD. Fyn is an attractive target for AD
therapeutics, not only based on its activation by Aβ via cellular prion protein
but also due to its known interaction with tau, uniquely linking the two key
pathologies in AD. Fyn is also a challenging target, with broad expression
throughout the body and significant homology with other members of the Src
family kinases, which may lead to unintended off-target effects. A phase 2a
proof-of-concept clinical trial in patients with AD is currently under way,
providing critical first data on the potential effectiveness of targeting Fyn in
AD. |
What is the link between TADs and GRBs? | Topologically associating domains (TADs) are ancient features that coincide with Metazoan clusters of extreme noncoding conservation (aka GRBs). | Developmental genes in metazoan genomes are surrounded by dense clusters of
conserved noncoding elements (CNEs). CNEs exhibit unexplained extreme levels of
sequence conservation, with many acting as developmental long-range enhancers.
Clusters of CNEs define the span of regulatory inputs for many important
developmental regulators and have been described previously as genomic
regulatory blocks (GRBs). Their function and distribution around important
regulatory genes raises the question of how they relate to 3D conformation of
these loci. Here, we show that clusters of CNEs strongly coincide with
topological organisation, predicting the boundaries of hundreds of topologically
associating domains (TADs) in human and Drosophila. The set of TADs that are
associated with high levels of noncoding conservation exhibit distinct
properties compared to TADs devoid of extreme noncoding conservation. The close
correspondence between extreme noncoding conservation and TADs suggests that
these TADs are ancient, revealing a regulatory architecture conserved over
hundreds of millions of years.Metazoan genomes contain many clusters of
conserved noncoding elements. Here, the authors provide evidence that these
clusters coincide with distinct topologically associating domains in humans and
Drosophila, revealing a conserved regulatory genomic architecture. |
What is Target Explorer? | Target Explorer is an automated tool for the identification of new target genes for a specified set of transcription factors. It was specifically designed for the well-annotated Drosophila melanogaster genome, but most options can be used for sequences from other genomes as well. Target Explorer is available at http://trantor.bioc.columbia.edu/Target_Explorer/ | |
Can the yeast protein Abf1 act as insulator? | Saccharomyces cerevisiae Rap1p and Abf1p proteins are endowed with a potent insulating capacity | Insulators are sequences that uncouple adjacent chromosome domains. Here we have
shown that Saccharomyces cerevisiae Rap1p and Abf1p proteins are endowed with a
potent insulating capacity. Insulating domains in Rap1p coincide with previously
described transcription activation domains, whereas four adjacent subdomains
spanning the whole of the Abf1p C terminus (440-731) were found to display
autonomous insulating capacity. That both Rap1p and Abf1p silencing domains
either contain or largely overlap with an insulating domain suggests that
insulation conveys some undefined chromosome organization capacity that also
contributes a function in silencing. Together with Reb1p and Tbf1p, previously
involved in the activity of Saccharomyces cerevisiae subtelomeric insulators,
insulating potential emerges as a supplementary common property of General
Regulatory Factors (GRFs). Thus GRFs, which bind to sites scattered throughout
the genome within promoters, would not only play a key role in regulating gene
expression but also partition the genome in functionally independent domains. |
What does the human IVIG treatment for Alzheimer's disease contain? | Human intravenous immunoglobulin (IVIG) is a mixture of polyclonal IgG antibodies isolated and pooled from thousands of healthy human donors. | Alzheimer's disease (AD) is a chronic neurodegenerative disease associated with
intracerebral accumulation of aggregated amyloid-beta (Aβ) and tau proteins, as
well as neuroinflammation. Human intravenous immunoglobulin (IVIG) is a mixture
of polyclonal IgG antibodies isolated and pooled from thousands of healthy human
donors. The scientific rationale for testing IVIG as a potential AD treatment
include its natural anti-Aβ antibody activity, its favorable safety profile and
inherent anti-inflammatory/immunomodulatory properties. Over the past decade,
several clinical and pre-clinical experimental findings, advanced our knowledge
about biological and therapeutic properties of IVIG that are relevant to AD
therapy. Anti-amyloid antibodies in IVIG show significantly higher binding
avidity for amyloid oligomers and fibrils than for Aβ monomers. In a double
transgenic murine model of AD, intracerebral injection of IVIG causes
suppression of Aβ fibril pathology whereas long term peripheral IVIG treatments
causes elevation of total brain Aβ levels with no measurable impact on Aβ
deposits or tendency for inducing cerebral microhemmorhage. Furthermore, chronic
IVIG treatment suppressed neuroinflammation and fostered adult hippocampal
neurogenesis. In clinical studies with AD patients, IVIG showed an acceptable
safety profile and has not been reported to increase the incidence of amyloid
related imaging abnormalities. Preliminary studies on small number of patients
reported clinical benefits in mild to moderate stage AD patients. However,
double blind, placebo controlled studies later did not replicate those initial
findings. Interestingly though, in APOE4 carriers and in moderate disease stage
subgroups, positive cognitive signals were reported. Nevertheless, both clinical
and experimental (mouse) studies show that antibodies in IVIG can accumulate in
CNS and its biological activities include neutralization of Aβ oligomers,
suppression of neuroinflammation and immunomodulation. Identifying mediators of
IVIG's effects at the cellular and molecular level is warranted. In light of its
favourable safety profile and aforementioned biological properties, IVIG is
still an enigmatic experimental candidate with enormous potential for being an
AD therapeutic. |
Is there any linear-time and linear-space algorithm for the computation of avoided words in biological sequences? | Yes. There is a linear-time and linear-space algorithm for the computation of avoided words of length k in a given sequence x. | |
What is the mechanism of the auxin-inducible degron system? | Fusion of inducible degradation signals, so-called degrons, to cellular proteins is an elegant method of controlling protein levels in vivo. The auxin-inducible degron (AID) system allows the rapid and reversible proteolysis of proteins of interest, and enables the generation of conditional mutants of budding yeast. Strategies that use ubiquitin-mediated protein degradation to eliminate the product of a gene of interest, such as heat-inducible degron (td) and auxin-inducible degron (AID), are powerful methods for constructing conditional mutants. Auxin-inducible degron (AID) technology allows rapid depletion of proteins in animal cells and fungi, but its application to human cells has been limited by the difficulties of tagging endogenous proteins. The auxin-inducible degron harbors great potential for dynamic protein depletion in yeast. Here, we thoroughly and quantitatively characterize the auxin-inducible degron in single yeast cells. | BACKGROUND: Inducible inactivation of a protein is a powerful approach for
analysis of its function within cells. Fission yeast is a useful model for
studying the fundamental mechanisms such as chromosome maintece and cell
cycle. However, previously published strategies for protein-depletion are
successful only for some proteins in some specific conditions and still do not
achieve efficient depletion to cause acute phenotypes such as immediate cell
cycle arrest. The aim of this work was to construct a useful and powerful
protein-depletion system in Shizosaccaromyces pombe.
RESULTS: We constructed an auxin-inducible degron (AID) system, which utilizes
auxin-dependent poly-ubiquitination of Aux/IAA proteins by SCFTIR1 in plants, in
fission yeast. Although expression of a plant F-box protein, TIR1, decreased
Mcm4-aid, a component of the MCM complex essential for DNA replication tagged
with Aux/IAA peptide, depletion did not result in an evident growth defect. We
successfully improved degradation efficiency of Mcm4-aid by fusion of TIR1 with
fission yeast Skp1, a conserved F-box-interacting component of SCF (improved-AID
system; i-AID), and the cells showed severe defect in growth. The i-AID system
induced degradation of Mcm4-aid in the chromatin-bound MCM complex as well as
those in soluble fractions. The i-AID system in conjunction with transcription
repression (off-AID system), we achieved more efficient depletion of other
proteins including Pol1 and Cdc45, causing early S phase arrest.
CONCLUSION: Improvement of the AID system allowed us to construct conditional
null mutants of S. pombe. We propose that the off-AID system is the powerful
method for in vivo protein-depletion in fission yeast. The auxin-inducible degron (AID) system allows the rapid and reversible
proteolysis of proteins of interest, and enables the generation of conditional
mutants of budding yeast. The construction of budding yeast AID mutants is
simple, and the effect of depletion of essential proteins on proliferation can
be confirmed by analyzing their phenotype. In this protocol, we describe a
procedure to generate AID mutants of budding yeast via a simple transformation
using PCR-amplified DNA. We also describe methods to confirm the depletion of
proteins of interest that are required for proliferation by serial-dilution and
liquid-culture assays. The analysis of consequences resulting after experimental elimination of gene
function has been and will continue to be an extremely successful strategy in
biological research. Mutational elimination of gene function has been widely
used in the fly Drosophila melanogaster. RNA interference is used extensively as
well. In the fly, exceptionally precise temporal and spatial control over
elimination of gene function can be achieved in combination with sophisticated
transgenic approaches and clonal analyses. However, the methods that act at the
gene and transcript level cannot eliminate protein products which are already
present at the time when mutant cells are generated or RNA interference is
started. Targeted inducible protein degradation is therefore of considerable
interest for controlled rapid elimination of gene function. To this end, a
degradation system was developed in yeast exploiting TIR1, a plant F box
protein, which can recruit proteins with an auxin-inducible degron to an E3
ubiquitin ligase complex, but only in the presence of the phytohormone auxin.
Here we demonstrate that the auxin-inducible degradation system functions
efficiently also in Drosophila melanogaster. Neither auxin nor TIR1 expression
have obvious toxic effects in this organism, and in combination they result in
rapid degradation of a target protein fused to the auxin-inducible degron. Author information:
(1)Center of Frontier Research, National Institute of Genetics, Research
Organization of Information and Systems, Yata 1111, Mishima, Shizuoka 411-8540,
Japan.
(2)Division of Biological Science, Graduate School of Science, Nagoya
University, Chikusa-ku, Nagoya 464-8602, Japan; PRESTO, Japan Science and
Technology Agency, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012, Japan.
(3)Division of Mammalian Development, Genetic Strains Research Center, National
Institute of Genetics, Yata 1111, Mishima, Shizuoka 411-8540, Japan; Department
of Genetics, SOKENDAI, Yata 1111, Mishima, Shizuoka 411-8540, Japan; Department
of Biological Sciences, Graduate School of Science, The University of Tokyo,
7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
(4)Center of Frontier Research, National Institute of Genetics, Research
Organization of Information and Systems, Yata 1111, Mishima, Shizuoka 411-8540,
Japan; PRESTO, Japan Science and Technology Agency, 4-1-8 Honcho, Kawaguchi,
Saitama 332-0012, Japan; Department of Genetics, SOKENDAI, Yata 1111, Mishima,
Shizuoka 411-8540, Japan. Electronic address: [email protected]. Generation of cells with a loss-of-function mutation in a gene (knockout cells)
is a valuable technique for studying the function of a given gene product.
However, if the product of the target gene is essential for cell viability,
conditional knockout cell lines must be generated. Recently, as gene editing
technology using CRISPR/Cas9 has developed, it has become possible to produce
conditional knockout cell lines using this technique. However, to obtain final
conditional knockout cell lines, it is necessary to perform several experiments
with multiple complicated steps. In this paper, we introduce an easy and
efficient method to generate conditional knockout cell lines based on combining
auxin-inducible degron (AID) technology with CRISPR/Cas9 gene editing. Our
method only requires performing a single transfection and is therefore an easy
and rapid method to obtain a conditional knockout cell line. |
Which bacteria causes erythrasma? | Corynebacterium minutissimum is the bacteria that leads to cutaneous eruptions of erythrasma and is the most common cause of interdigital foot infections. | Infective embolic retinopathy as a sequela of bacterial endocarditis is
described in a 31-year-old woman with mitral valve prolapse. The infective
organism, Corynebacterium minutissimum, has not been previously found to cause
ocular or multisystem diseases. It is a common mucocutaneous inhabitant which
causes erythrasma. In our case report both ocular involvement and septicaemia
were present. The infection was confirmed by positive serial blood cultures.
Mitral valve prolapse was confirmed by echocardiography. On clinical examination
the retinopathy consisted of white intraretinal lesions which resolved with
antibiotic therapy. By fluorescein angiography focal areas of hypofluorescence
corresponding to the white fundus lesions were present. Optic disc oedema was
also seen. Bacterial skin infections are important to recognize because we have the means
to eradicate almost all of them. Primary skin infections are mainly caused by
staphylococci or streptococci. Staphylococci infections present as furuncles and
carbuncles, superficial folliculitis, impetigo or rarely the Scalded Skin
Syndrome. Streptococcal infections present as impetigo, ecthyma, erysipelas or
cellulitis. Corynebacteria causes erythrasma, trichomycosis or pitted
keratolysis. Gram-negative primary skin infections, although uncommon, may
occur; bacterial cultures are generally necessary for diagnosis. Secondary
bacterial infections of pre-existing wounds, burns, dermatitic skin, or
retention cysts are common events. BACKGROUND: Erythrasma is an uncommon vulvar infection, best diagnosed by its
fluorescence under the Wood lamp. This report shows that despite a negative Wood
lamp examination, the diagnosis can be made histologically.
CASE: A 42-year-old woman was referred to our clinic with a persistent candidal
infection. Evaluation included a Wood lamp examination, wet mount, and potassium
hydroxide test of the affected skin, all of which were negative. A biopsy of the
area demonstrated rods and filamentous organisms in the keratotic layer
consistent with a Corynebacterium minutissimum infection. The patient was
diagnosed as having erythrasma, and she responded to oral erythromycin.
CONCLUSION: Persistent vulvar diseases may be caused by erythrasma despite a
negative Wood lamp examination. The diagnosis can be made by biopsy of the
lesion. Skin infections are common and may be caused by bacteria, fungi or viruses.
Breaks in the skin integrity, particularly those that inoculate pathogens into
the dermis, frequently cause or exacerbate skin infections. Bacterial skin
infections caused by corynebacteria include erythrasma, trichomycosis axillaris
and pitted keratolysis. Staphylococci may cause impetigo, ecthyma and
folliculitis. Streptococcal skin infections include impetigo and erysipelas.
Human papillomavirus skin infections present as several different types of
warts, depending on the surface infected and its relative moisture, and the
patterns of pressure. The many dermatomycoses (skin infections caused by fungi
or yeasts) include tinea capitis, tinea barbae, tinea cruris, tinea manus, tinea
pedis and tinea unguium (onychomycosis). Candidal infections occur in moist
areas, such as the vulva, mouth, penis, skinfolds and diaper area. Wounds caused
by wood splinters or thorns may result in sporotrichosis. Animal bites may
result in complex, serious infections, requiring tetanus and, possibly, rabies
prophylaxis in addition to appropriate antibiotic therapy. Corynebacterium minutissimum is the bacteria that leads to cutaneous eruptions
of erythrasma and is the most common cause of interdigital foot infections. It
is found mostly in occluded intertriginous areas such as the axillae,
inframammary areas, interspaces of the toes, intergluteal and crural folds, and
is more common in individuals with diabetes mellitus than other clinical
patients. This organism can be isolated from a cutaneous site along with a
concurrent dermatophyte or Candida albicans infection. The differential
diagnosis of erythrasma includes psoriasis, dermatophytosis, candidiasis and
intertrigo, and methods for differentiating include Wood's light examination and
bacterial and mycological cultures. Erythromycin 250mg four times daily for 14
days is the treatment of choice and other antibacterials include tetracycline
and chloramphenicol; however, the use of chloramphenicol is limited by bone
marrow suppression potentially leading to neutropenia, agranulocytosis and
aplastic anaemia. Further studies are needed but clarithromycin may be an
additional drug for use in the future. Where there is therapeutic failure or
intertriginous involvement, topical solutions such as clindamycin, Whitfield's
ointment, sodium fusidate ointment and antibacterial soaps may be required for
both treatment and prophylaxis. Limited studies on the efficacy of these
medications exist, however, systemic erythromycin demonstrates cure rates as
high as 100%. Compared with tetracyclines, systemic erythromycin has greater
efficacy in patients with involvement of the axillae and groin, and similar
efficacy for interdigital infections. Whitfield's ointment has equal efficacy to
systemic erythromycin in the axillae and groin, but shows greater efficacy in
the interdigital areas and is comparable with 2% sodium fusidate ointment for
treatment of all areas. Adverse drug effects and potential drug interactions
need to be considered. No cost-effectiveness data are available but there are
limited data on cost-related treatment issues. A guideline is proposed for the
detection, evaluation, treatment and prophylaxis of this cutaneous eruption. BACKGROUND: Erythrasma is a superficial cutaneous infection caused by
Corynebacterium minutissimum and is characterized by fluorescence under Wood's
light (UV) because of the presence of porphyrins. These molecules are
photosensitizing and we propose to assess efficacy of red light that activates
porphyrins (photodynamic reaction) in treatment of this pathology.
OBJECTIVES: Assessment of effects of photodynamic action of red light for
treatment of erythrasma without exogenous photosensitizing molecules.
METHODS: Thirteen patients with erythrasma were treated by one illumination (80
J/cm2) by red light (broad band, peak at 635 nm) without exogenous
photosensitizing molecules. Disappearance or reduction of extent of lesions were
observed 2 weeks later. If lesions were still present, a second irradiation was
conducted with the same method.
RESULTS: Preliminary results are presented. As a result of red light
irradiation, we noticed a complete recovery for three patients and, in most
other cases, reduction of extent of lesions (mean: -29% after one session). The
treatment was well tolerated.
CONCLUSION: We report first cases of photodynamic treatment of erythrasma. There
are other reports of clinical applications of antimicrobial action of
photodynamic therapy in dermatology (acne vulgaris, leishmaniasis, warts, etc.).
But there are few applications without addition of exogenous photosensitizing
agent. The originality and interest of our study is to use spontaneous presence
of porphyrins in the lesions. This technique seems to be an interesting
alternative, inexpensive and easy, for the treatment of this localized
infection. But an optimal method is still to be determined to improve efficacy. BACKGROUND: Erythrasma is a superficial infection caused by Corynebacterium
minutissimum and affects the major skin folds and the interdigital regions of
the feet. It is characterized by erythematous, brown, scaly patches and
maceration, and exhibits coral-red fluorescence under Wood light.
OBJECTIVE: The aim of this study was to determine the frequency of erythrasma in
patients with interdigital lesions.
METHODS: An open, prospective, longitudinal, observational study was performed
in a hospital in Mexico City between March and December, 2006. All patients with
interdigital lesions were examined with a Wood lamp and direct examination was
performed with 20 % potassium hydroxide. Cultures were done in Sabouraud
dextrose agar and brain heart infusion agar, and smears were analyzed. General
characteristics and concomitant diseases were recorded.
RESULTS: We examined 73 patients, of whom 24 (32.8 %) were diagnosed with
erythrasma based on coral-red fluorescence under Wood light and identification
of corynebacteria by Gram staining. The disease was more common in women (83.33
%) and the mean age of the patients was 43.5 years. The main clinical findings
were scaling and maceration, and the fourth interdigital web was the most
commonly affected. Corynebacterium could not be isolated in any of the cases.
Mycology was positive in 15 cases (62.5 %) and the following microorganisms were
isolated: Candida (16.6 %), dermatophytes (12.5 %), and Trichosporon (4.1 %).
CONCLUSIONS: Interdigital erythrasma is a common condition and can be easily
confused with interdigital tinea. It persists if not treated appropriately.
Rapid diagnosis is easily obtained by examination with a Wood lamp, while
culture is difficult and unnecessary for diagnosis. The coexistence of
erythrasma with dermatophytes and Candida should be considered when the
interdigital webs are affected. BACKGROUND: Corynebacterium spp. are diphtheroid bacteria responsible for pitted
keratolysis, a common plantar infection confined to the thick stratum corneum.
AIM: To study a series of demographic features of patients suffering from pitted
keratolysis, and to present a review of the Corynebacterium-associated
infections, including pitted keratolysis, erythrasma, and trichobacteriosis.
MATERIALS AND METHODS: A 2-year, two-center, prospective survey assessed the
demographics of pitted keratolysis, including age, gender, site of infection,
symptoms, patients' complaints, the use of protective and/or occlusive shoes,
seasonality of diagnosis, drug intake, associated skin signs (including
dyshidrosis, erythrasma, and trichobacteriosis), recurrences, and previous
diagnoses and treatments.
RESULTS: The mean age of the 53 patients with pitted keratolysis was 24.9 years
(range, 10-57 years). The male to female ratio was 7.8:1. The soles of both feet
were commonly involved (92.4%). Pressure-bearing areas were the usual sites of
infection, ranging from restricted involvement of the toes (12/53, 22.6%) to
spreading to the entire plantar surface (15/53, 28.3%). A total of 36 (68%) of
the 53 patients complained of hyperhidrosis. An unpleasant smell and pain were
noted by 35 (66%) and 25 (47%) of the 53 patients, respectively. Occlusive and
protective shoes were worn in 51 (96.2%) and 31 (58.4%) of the 53 cases,
respectively.
CONCLUSION: Pitted keratolysis commonly affects young male patients wearing
protective shoes for professional reasons, inducing a moist and warm
environment. Hyperhidrosis, an unpleasant smell, and pain are the main clinical
complaints. Erythrasma is a superficial skin disease caused by Gram-positive Corynebacterium
species. Coral-red fluorescence under Wood's light, strongly suggestive of
erythrasma, can be attributed to the presence of porphyrins. Fractionated
porphyrin analysis in erythrasma lesions is yet to be reported. We attempted to
investigate erythrasma lesions by isolating the responsible bacteria and
determining their exogenous porphyrin production by HPLC analysis. We observed a
78-year-old woman with erythrasma who had a well-demarcated slightly scaling
patch on her left foot, between the fourth and fifth toes. Two kinds of colonies
on 5 % sheep blood agar were obtained from this lesion. Analysis of the 16S rRNA
sequence revealed the colonies to be Corynebacterium aurimucosum and
Microbacterium oxydans. HPLC analysis demonstrated that coproporphyrin III
(Copro III) levels were clearly elevated, although the amounts of protoporphyrin
were diminished. These results indicate that the fluorescent substance was Copro
III. This study supports the view that excess Copro III synthesis by C.
aurimucosum and M. oxydans leads to accumulation of porphyrin in cutaneous
tissue, which emits a coral-red fluorescence when exposed to Wood's light. BACKGROUND: Erythrasma is a skin infection which is caused by Corynebacterium
minutissimum. Interdigital erythrasma is the most common form.
OBJECTIVE: The aim of this study was to detect the frequency and risk factors of
interdigital erythrasma in patients with clinically suspected tinea pedis.
METHODS: This study was conducted between June and December 2010 and included
122 patients who had interdigital foot lesions. All patients were examined using
a Wood's lamp. The smears were stained using Gram's method. Direct examination
was performed using 20% potassium hydroxide. Sabouraud dextrose agar and brain
heart infusion agar were used for cultures. Moreover, the demographical
characteristics of patients, concomitant diseases and clinical findings were
also recorded. Cases that were found to be positive on Wood's lamp examination
and/or Gram staining/culture were considered as erythrasma.
RESULTS: The rate of erythrasma was found to be 46.7%. The mean age was 43.6
years, and the disease was more prevalent in men. The most common clinical
finding was desquamation. Mycological examination was found as positive in
40.35% of the patients. No growth was observed in bacteriological cultures. It
was found that using only Wood's lamp examination or Gram staining resulted in
11 (9%) and 19 positive patients (15.6%), respectively, whereas using both
Wood's lamp examination and Gram staining concurrently resulted in 27 positive
patients (22.1%).
CONCLUSION: Interdigital erythrasma is a commonly seen condition and can
clinically mimic tinea pedis. A Wood's lamp is a good diagnostic tool, but Gram
staining, particularly in those with a negative Wood's lamp result, may be a
useful method. Interdigital foot infections are mostly caused initially by dermatophytes,
yeasts and less frequently by bacteria. Erythrasma caused by Corynebacterium
minutissimum can be confused with superficial mycoses. The aim of the study was
to determine the prevalence of the etiologic agents of superficial mycoses and
the frequency of Corynebacterium minutissimum in interdigital foot infections.
All the samples obtained from the 121 patients with interdigital foot infections
were examined directly with the use of 20% potassium hydroxide mounts and Gram
stain under the microscope and cultured on Sabouraud's dextrose agar plates. In
identification of superficial mycoses, the rate was found to be 14% with the
cultural method and 14% with direct microscopic examination. Using a combination
of direct microscopic examination and culture, a 33.8% ratio was achieved. In
the culture of these samples, the most isolated factor was Trichophyton rubrum
(33.7%). In 24 of the patients (19.8%) Corynebacterium minutissimum was detected
by Gram staining, in 6 of these patients Trichophyton rubrum was found,
Trichophyton mentagrophytes was found in 2 and Trichosporon spp. was found in 1.
The examination of interdigital foot lesions in the laboratory, the coexistence
of erythrasma with dermatophytes and yeast should be considered. |
Mutations in which gene cause Schimke immune-osseous dysplasia? | SMARCAL1, also known as HARP, is an ATP-dependent annealing helicase that stabilizes replication forks during DNA damage. Mutations in this gene are the cause of Schimke immune-osseous dysplasia, an autosomal recessive disorder characterized by T-cell immunodeficiency and growth dysfunctions. | Schimke immuno-osseous dysplasia (SIOD, MIM 242900) is an autosomal-recessive
pleiotropic disorder with the diagnostic features of spondyloepiphyseal
dysplasia, renal dysfunction and T-cell immunodeficiency. Using genome-wide
linkage mapping and a positional candidate approach, we determined that
mutations in SMARCAL1 (SWI/SNF2-related, matrix-associated, actin-dependent
regulator of chromatin, subfamily a-like 1), are responsible for SIOD. Through
analysis of data from persons with SIOD in 26 unrelated families, we observed
that affected individuals from 13 of 23 families with severe disease had two
alleles with nonsense, frameshift or splicing mutations, whereas affected
individuals from 3 of 3 families with milder disease had a missense mutation on
each allele. These observations indicate that some missense mutations allow
retention of partial SMARCAL1 function and thus cause milder disease. Schimke-immuno-osseous dysplasia is a rare autosomal-recessive multisystem
disorder with the main clinical features of disproportionate growth deficiency,
defective cellular immunity, and progressive renal disease. It is caused by
mutations of SMARCAL1, a gene encoding a putative chromatin remodeling protein
of unknown function. Because a detailed description of the clinical features is
an essential first step in elucidating the function of SMARCAL1, we present the
first detailed anthropometric data for Schimke-immuno-osseous dysplasia
patients. By comprehensive anthropometric examination (28 parameters) of 8
patients (3 females) with the typical findings of Schimke-immuno-osseous
dysplasia (mean age: 14.8 years; range: 4.9-30.5 years) and 304 patients (117
females) with congenital and hereditary chronic kidney disease (mean age: 10.7
+/- 4.8 years; range: 3-21.8 years), we show that Schimke-immuno-osseous
dysplasia patients differ significantly from those with other forms of chronic
kidney disease. z scores were calculated with reference limits derived from 5155
healthy children (2591 females) aged 3 to 18 years. The key finding was that, in
the latter group, median leg length was significantly more reduced than sitting
height, whereas in Schimke-immuno-osseous dysplasia patients, the reduction of
sitting height was significantly more pronounced than for leg length. Therefore,
the ratio of sitting height/leg length might be a simple tool for the clinician
to distinguish Schimke-immuno-osseous dysplasia from other chronic kidney
disease patients. Schimke-immuno-osseous dysplasia is very likely if this ratio
is < 0.83. However, other forms of chronic kidney disease have to be discussed
in case of a ratio > 1.01. Schimke immuno-osseous dysplasia (OMIM 242900) is an uncommon
autosomal-recessive multisystem disease caused by mutations in SMARCAL1
(swi/snf-related, matrix-associated, actin-dependent regulator of chromatin,
subfamily a-like 1), a gene encoding a putative chromatin remodeling protein.
Neurologic manifestations identified to date relate to enhanced atherosclerosis
and cerebrovascular disease. Based on a clinical survey, we determined that half
of Schimke immuno-osseous dysplasia patients have a small head circumference,
and 15% have social, language, motor, or cognitive abnormalities. Postmortem
examination of 2 Schimke immuno-osseous dysplasia patients showed low brain
weights and subtle brain histologic abnormalities suggestive of perturbed
neuron-glial migration such as heterotopia, irregular cortical thickness,
incomplete gyral formation, and poor definition of cortical layers. We found
that SMARCAL1 is highly expressed in the developing and adult mouse and human
brain, including neural precursors and neuronal lineage cells. These
observations suggest that SMARCAL1 deficiency may influence brain development
and function in addition to its previously recognized effect on cerebral
circulation. Schimke immuno-osseous dysplasia is a rare autosomal recessive multisystem
disorder characterized by steroid-resistant nephrotic syndrome,
immunodeficiency, and spondyloepiphyseal dysplasia. Mutations in SWI/SNF2
related, matrix associated, actin dependent regulator of chromatin, subfamily
a-like 1 (SMARCAL1) gene are responsible for the disease. The present report
describes, for the first time, a Schimke immuno-osseous dysplasia child with
SMARCAL1 missense mutation (R561H) and manifestations of intussusception
secondary to Epstein-Barr virus-negative non-Hodgkin lymphoma, who expired due
to septicemia following chemotherapy. The report emphasizes the necessity of
more limited immunosuppressive protocols in Schimke immuno-osseous dysplasia
patients with lymphoproliferative disorders. BACKGROUND: Schimke immuno-osseous dysplasia (SIOD, OMIM #242900) is an
autosomal-recessive pleiotropic disorder characterized by spondyloepiphyseal
dysplasia, renal dysfunction and T-cell immunodeficiency. SIOD is caused by
mutations in the gene SMARCAL1.
CASE PRESENTATION: We report the clinical and genetic diagnosis of a 5-years old
girl with SIOD, referred to our Center because of nephrotic-range proteinuria
occasionally detected during the follow-up for congenital hypothyroidism.
Mutational analysis of SMARCAL1 gene was performed by polymerase chain reaction
(PCR) and bidirectional sequencing. Sequence analysis revealed that patient was
compound heterozygous for two SMARCAL1 mutations: a novel missense change
(p.Arg247Pro) and a well-known nonsense mutation (p.Glu848*).
CONCLUSION: This report provided the clinical and genetic description of a mild
phenotype of Schimke immuno-osseous dysplasia associated with nephrotic
proteinuria, decreasing after combined therapy with ACE inhibitors and sartans.
Our experience highlighted the importance of detailed clinical evaluation,
appropriate genetic counseling and molecular testing, to provide timely
treatment and more accurate prognosis. OBJECTIVE: Schimke immuno-osseous dysplasia (SIOD), is an autosomal recessive
inherited disease caused by SMARCAL1 (MIM:20606622) mutations, while in about
half of the patients no any mutation in SMARCAL1 could be found. This disease
involves multiple systems and is characterized by short and dissymmetric stature
with spondyloepiphyseal dysplasia, progressive renal failure, lymphopenia with
recurrent infections, and hyperpigmented macules. This study aimed to analyze
SMARCAL1 gene of 2 unrelated suspected SIOD children, to make definite
diagnosis, and find more SMARCAL1 mutation types of Chinese SIOD.
METHOD: Two suspected Chinese Han male SIOD children who visited our hospital
from 2008 to 2014, aged 3 y 6 m and 7 y 8 m, both were short and had
spondyloepiphyseal dysplasia, progressive renal failure, lymphopenia with
recurrent infections. After informed consent, they and their parents's DNA were
extracted from blood. PCRs for all 16 exons of SMARCAL1 were performed and PCR
products were purified by 2% gel electrophoresis and sequenced directly.
Pathogenicity of missense variations was confirmed by SIFT and sequencing
SMARCAL1 of fifty normal controls.
RESULT: (1) Four gene variations were found in the two children: Two reported
missense mutations c.1129G>C, p.Glu377Gln and c.1933C>T, p. Arg645Cys. Two
splicing mutations c.1334+1G>A and c.2142-1 G>A were detected. (2) c.1129G>C,
p.Glu377Gln were reported as a disease-causing mutations before, but it was an
single nucleotide polymorphism (SNP) which was found in 15 of 50 normal
controls. (3) Two novel splicing mutations were found in this study: c.1334+1G>A
and c.2142-1 G>A.
CONCLUSION: (1) We detected 3 disease-causing mutations in 2 SIOD children by
SMARCAL1 gene analysis, while 2 splicing mutations were novel mutations. (2)
c.1129G>C, p.Glu377Gln was a SNP but not a disease-causing mutation at least in
Chinese population. Schimke immuno-osseous dysplasia is an autosomal recessive multisystem disorder
caused by defects in SWI/SNF-related, matrix-associated, actin-dependent
regulator of chromatin, subfamily a-like 1 gene (SMARCAL1). SMARCAL1 product is
a helicase that has role in selective cellular proliferation. The disorder is
characterized by spondyloepiphyseal dysplasia with short stature, nephropathy, T
cell deficiency, neurologic and cutaneous signs. Patients may have
hyperpigmented skin lesions similar to café au lait spots. Symptoms and disease
severity in Schimke immuno-osseous dysplasia varies from patient to patient.
Genetic, epigenetic and environmental factors play role on the severity of the
disease. Here we report on a patient with short stature, steroid resistant
nephrotic syndrome and recurrent infections. Cutaneous findings and
developmental delay helped us to reach the diagnosis of Schimke immuno-osseous
dysplasia. A homozygous missense mutation in SMARCAL1 gene confirmed the
clinical diagnosis. Mutations in SMARCAL1, which encodes a DNA annealing helicase with roles in DNA
replication fork restart, DNA repair, and gene expression modulation, cause
Schimke immuno-osseous dysplasia (SIOD), an autosomal recessive disease
characterized by skeletal dysplasia, renal disease, T-cell immunodeficiency, and
arteriosclerosis. The clinical features of SIOD arise from pathological changes
in gene expression; however, the underlying mechanism for these gene expression
alterations remains unclear. We hypothesized that changes of the epigenome alter
gene expression in SIOD. To test this, we performed a genetic screen for
interaction between Marcal1, the Drosophila melanogaster ortholog of SMARCAL1,
and the genes of the trithorax group (trxG) and Polycomb group (PcG), which
encode epigenetic regulators. SMARCAL1 and Marcal1 genetically interacted with
trxG and PcG members. A homozygous null mutation of Marcal1 suppressed the
wing-to-haltere transformation, ectopic Ultrabithorax (Ubx) expression, and
ectopic Ubx minigene expression caused by PcG deficiency. The suppression of
ectopic Ubx expression correlated with reduced chromatin accessibility of the
Ubx promoter. To our knowledge, this is the first in vivo evidence for
deficiency of a SMARCAL1 ortholog altering the chromatin structure of a gene. SMARCAL1 (SWI/SNF Related, Matrix Associated, Actin Dependent Regulator Of
Chromatin, Subfamily A-Like 1), also known as HARP, is an ATP-dependent
annealing helicase that stabilizes replication forks during DNA damage.
Mutations in this gene are the cause of Schimke immune-osseous dysplasia (SIOD),
an autosomal recessive disorder characterized by T-cell immunodeficiency and
growth dysfunctions. In this review, we summarize the main roles of SMARCAL1 in
DNA repair, telomere maintece and replication fork stability in response to
DNA replication stress. |
What is the most common histological diagnosis of "butterfly glioma"? | Butterfly glioma is glioblastoma multiforme invading corpus callosum . | A 45 year-old male with a butterfly glioma received stereotactic biopsy for
histologic confirmation of the clinical diagnosis. Microscopically, the results
were controversial since some biopsy specimens showed distinct inflammatory
changes, while others displayed typical features of a maligt glioma. The
patient died four days after the stereotactic approach due to therapy-resistant
intracranial pressure rise. In addition to a large butterfly glioblastoma
originating from the frontal part of the corpus callosum, neuropathologic
examination revealed a mycotic encephalitis with formation of numerous
fungi-containing inflammatory foci in all parts of the brain and in the glioma.
General autopsy disclosed pulmonary aspergillosis as the source of the
inflammatory spread. A previous steroid medication over several weeks for
treatment of increased intracranial pressure may be considered as an important
factor in the origin of the pulmonary aspergillosis complicating the butterfly
glioma. The authors report a case of neuronal ceroid lipofuscinosis (Kufs' disease)
confirmed by stereotactically obtained brain biopsy findings and initially
diagnosed as a butterfly glioma. The presenting symptoms in the 64-year-old
patient were mental alterations with progressive dementia, followed by muscular
atrophy and myoclonia with distal preponderance. The mild initial disturbances
of coordination increased, and the patient developed a markedly ataxic gait.
Computerized tomography (CT) scanning and magnetic resoce imaging revealed
generalized cerebral atrophy and a bifrontal space-occupying lesion involving
the callosum. The original "clearcut" diagnosis of glioblastoma multiforme,
based on CT scans, was unexpectedly disproved by examination of stereotactically
obtained brain biopsy specimens, which revealed a neuronal ceroid lipofuscinosis
(Kufs' disease). To the authors' knowledge, this is the first report of a case
presenting with both diffuse brain atrophy and localized accumulation of
neuronal lipofuscin, mimicking a mass lesion on radiological studies. Glioblastoma multiforme (GBM), the most common maligt brain tumor of adults,
is relatively rare in children. In a GBM affecting a 16-year-old boy, the tumor
spread across the corpus callosum (butterfly glioma). This type of bilateral
hemispheric growth has previously been thought to result from spread along the
white matter tracts. Two samples obtained from opposite sides of the same tumor
were analyzed comprehensively for loss of heterozygosity (LOH) and
microsatellite instability (MSI). Amplification of EGFR and MDM2 was studied by
means of multiplex polymerase chain reaction. Exons 5, 6, 7, and 8 of TP53 were
screened for mutations by sequencing. In neither specimen were molecular
alterations found in the EGFR, MDM2, or TP53 genes. The specimen obtained from
the right hemisphere exhibited a high level of MSI and LOH in chromosome arms
5q, 9p, and 13q. The specimen from the left hemisphere exhibited LOH in
chromosome arms 3p, 5q, 9p, 9q, 10p, 10q, and 13q. Here we propose four
plausible hypothetical scenarios underlying the tumorigenesis of this GBM. In humans, high-grade gliomas may infiltrate across the corpus callosum
resulting in bihemispheric lesions that may have symmetrical, winged-like
appearances. This particular tumor manifestation has been coined a "butterfly"
glioma (BG). While canine and human gliomas share many neuroradiological and
pathological features, the BG morphology has not been previously reported in
dogs. Here, we describe the magnetic resoce imaging (MRI) characteristics of
BG in three dogs and review the potential differential diagnoses based on
neuroimaging findings. All dogs presented for generalized seizures and
interictal neurological deficits referable to multifocal or diffuse forebrain
disease. MRI examinations revealed asymmetrical (2/3) or symmetrical (1/3),
bihemispheric intra-axial mass lesions that predomitly affected the
frontoparietal lobes that were associated with extensive perilesional edema, and
involvement of the corpus callosum. The masses displayed heterogeneous T1, T2,
and fluid-attenuated inversion recovery signal intensities, variable contrast
enhancement (2/3), and mass effect. All tumors demonstrated classical
histopathological features of glioblastoma multiforme (GBM), including glial
cell pseudopalisading, serpentine necrosis, microvascular proliferation as well
as invasion of the corpus callosum by neoplastic astrocytes. Although rare, GBM
should be considered a differential diagnosis in dogs with an MRI evidence of
asymmetric or symmetric bilateral, intra-axial cerebral mass lesions with signal
characteristics compatible with glioma. A 54-year-old man presented with change in behaviour, nocturnal enuresis,
abnormal limb movement and headache of one week's duration. The diagnosis of
butterfly glioma (glioblastoma multiforme) was made based on imaging
characteristics and was further confirmed by biopsy findings. As the corpus
callosum is usually resistant to infiltration by tumours, a mass that involves
and crosses the corpus callosum is suggestive of an aggressive neoplasm. Other
neoplastic and non-neoplastic conditions that may involve the corpus callosum
and mimic a butterfly glioma, as well as associated imaging features, are
discussed. |
Which chromosomes are implicated in the Emanuel syndrome? | Emanuel syndrome is associated with supernumerary chromosome t(11;22)(q23;q11), which consists of the extra genetic material from chromosomes 11 and 22. | Emanuel syndrome results from +der(22)t(11q23;22q11). Cleft palate, ear
anomalies, heart defects, genital anomalies, hypotonia, and mental retardation
are the main features of the syndrome. We report a nine-year-old boy with the
t(11;22)(q23;q11) chromosome, transmitted in an unbalanced fashion from his
mother, and originated in the maternal grandmother's meiosis. In addition to
mental retardation, hypotonia, craniofacial anomalies, and cryptorchidism, he
has novel findings such as, joint hyperextensibility, left liver lobe agenesis,
left sided malposition of the gallbladder and pancreas hypoplasia. This is the
first report associating these features with Emanuel syndrome. Emanuel syndrome is characterized by multiple congenital anomalies and
developmental disability. It is caused by the presence of a supernumerary
derivative chromosome that contains material from chromosomes 11 and 22. The
origin of this imbalance is 3:1 malsegregation of a parental balanced
translocation between chromosomes 11 and 22, which is the most common recurrent
reciprocal translocation in humans. Little has been published on the clinical
features of this syndrome since the 1980s and information on natural history is
limited. We designed a questionnaire to collect information from families
recruited through an international online support group, Chromosome 22 Central.
Data gathered include information on congenital anomalies, medical and surgical
history, developmental and behavioral issues, and current abilities. We received
information on 63 individuals with Emanuel syndrome, ranging in age from newborn
to adulthood. As previously recognized, congenital anomalies were common, the
most frequent being ear pits (76%), micrognathia (60%), heart malformations
(57%), and cleft palate (54%). Our data suggest that vision and hearing
impairment, seizures, failure to thrive and recurrent infections, particularly
otitis media, are common in this syndrome. Psychomotor development is uniformly
delayed, however the majority of individuals (over 70%) eventually learn to walk
with support. Language development and ability for self-care are also very
impaired. This study provides new information on the clinical spectrum and
natural history of Emanuel syndrome for families and physicians caring for these
individuals. Many human diseases share a developmental origin that manifests during childhood
or maturity. Aneuploid syndromes are caused by supernumerary or reduced number
of chromosomes and represent an extreme example of developmental disease, as
they have devastating consequences before and after birth. Investigating how
alterations in gene dosage drive these conditions is relevant because it might
help treat some clinical aspects. It may also provide explanations as to how
quantitative differences in gene expression determine phenotypic diversity and
disease susceptibility among natural populations. Here, we aimed to produce
induced pluripotent stem cell (iPSC) lines that can be used to improve our
understanding of aneuploid syndromes. We have generated iPSCs from monosomy X
[Turner syndrome (TS)], trisomy 8 (Warkany syndrome 2), trisomy 13 (Patau
syndrome) and partial trisomy 11;22 (Emanuel syndrome), using either skin
fibroblasts from affected individuals or amniocytes from antenatal diagnostic
tests. These cell lines stably maintain the karyotype of the donors and behave
like embryonic stem cells in all tested assays. TS iPSCs were used for further
studies including global gene expression analysis and tissue-specific directed
differentiation. Multiple clones displayed lower levels of the pseudoautosomal
genes ASMTL and PPP2R3B than the controls. Moreover, they could be transformed
into neural-like, hepatocyte-like and heart-like cells, but displayed
insufficient up-regulation of the pseudoautosomal placental gene CSF2RA during
embryoid body formation. These data support that abnormal organogenesis and
early lethality in TS are not caused by a tissue-specific differentiation
blockade, but rather involves other abnormalities including impaired
placentation. PURPOSE: Emanuel syndrome is a rare chromosomal disorder characterized by severe
mental retardation and multiple anomalies. The syndrome is caused by chromosomal
imbalance due to a supernumerary derivative chromosome 22. Little is known
regarding the characteristics of prenatal biochemical screening, or
ultrasonographic markers in this syndrome. We aimed to identify a prenatal
screening pattern characteristic of Emanuel Syndrome.
METHODS: We report the prenatal characteristics of five fetuses with Emanuel
syndrome, four of which were diagnosed prenatally.
RESULTS: We found no consistent pattern of prenatal biochemical markers or other
prenatal characteristics. Nevertheless, increased NT, low PAPP-A and ultrasound
features such as intra uterine growth restriction, posterior fossa, cardiac and
bowel abnormalities may be helpful in raising the suspicion for this rare
genetic syndrome.
CONCLUSION: Review of the biochemical screening results, ultrasound findings,
and demographic characteristics of this Emanuel syndrome case series, as well as
of the relevant literature fail to suggest a characteristic prenatal pattern. Emanuel syndrome is an inherited chromosomal abnormality resulting from 3:1
meiotic segregation from parental balanced translocation carrier
t(11;22)(q23;q11), mostly of maternal origin. It is characterized by mental
retardation, microcephaly, preauricular tag or sinus, ear anomalies, cleft or
high arched palate, micrognathia, congenital heart diseases, kidney
abnormalities, structural brain anomalies and genital anomalies in male. Here
in, we describe a female patient with supernumerary der(22) syndrome (Emanuel
syndrome) due to balanced translocation carrier father t(11;22) (q23;q11). She
was mentally and physically disabled and had most of the craniofacial
dysmorphism of this syndrome. Our patient had cleft palate, maldeveloped corpus
callosum and hind brain with normal internal organs. Additionally,
arachnodactyly, hyperextensibility of hand joints, abnormal deep palmar and
finger creases, extra finger creases and bilateral talipus were evident and not
previously described with this syndrome. Cytogenetic analysis and FISH
documented that the patient had both translocation chromosomes plus an
additional copy of der(22) with karyotyping: 47,XX,t(11;
22)(q23;q11),+der(22)t(11;22)(q23;q11). We postulated that this rare chromosomal
complement can arise from; 2:2 segregation in the first meiotic division of the
balanced translocation father followed by non-disjunction at meiosis II in the
balanced spermatocyte. We observed a t(11;22)(q23-24;q11.2-12) and monosomy 3 in renal tumor cells from
a 72-year-old man. The hypothesis of a primitive peripheral neuroectodermal
tumor (PPNET) located in the kidney was promptly excluded: Histologically, the
tumor was a clear cell renal cell carcinoma (RCC) and we did not observe an
EWSR1 gene rearrangement. The constitutional origin of this alteration was
established. We report on the second case of RCC in a patient with a
constitutional t(11;22). The t(11;22)(q23;q11.2) is the main recurrent germline
translocation in humans. Unbalanced translocation can be transmitted to the
progeny and can cause Emanuel syndrome. Our observation alerts cancer
cytogeneticists to the fortuitous discovery of the constitutional t(11;22) in
tumor cells. This translocation appears grossly similar to the t(11;22)(q24;q12)
of PPNET and should be evoked if present in all cells of a tumor other than
PPNET. This is important when providing appropriate genetic counseling.
Moreover, the potential oncogenic role of the t(11;22) and its predisposing risk
of cancer are under debate. The family history of the patient revealed a
disabled brother who died at an early age from colon cancer and a sister with
breast cancer. This observation reopens the issue of a link between the
constitutional t(11;22) and cancer, and the utility of cancer prevention workups
for t(11;22) carriers. BACKGROUND: Complex small supernumerary marker chromosomes (sSMC) constitute one
of the smallest subgroups of sSMC in general. Complex sSMC consist of
chromosomal material derived from more than one chromosome; the best known
representative of this group is the derivative chromosome 22 {der(22)t(11;22)}
or Emanuel syndrome. In 2008 we speculated that complex sSMC could be part of an
underestimated entity.
RESULTS: Here, the overall yet reported 412 complex sSMC are summarized. They
constitute 8.4% of all yet in detail characterized sSMC cases. The majority of
the complex sSMC is contributed by patients suffering from Emanuel syndrome
(82%). Besides there are a der(22)t(8;22)(q24.1;q11.1) and a
der(13)t(13;18)(q11;p11.21) or der(21)t(18;21)(p11.21;q11.1) = der(13 or 21)t(13
or 21;18) syndrome. The latter two represent another 2.6% and 2.2% of the
complex sSMC-cases, respectively. The large majority of complex sSMC has a
centric minute shape and derives from an acrocentric chromosome. Nonetheless,
complex sSMC can involve material from each chromosomal origin. Most complex
sSMC are inherited form a balanced translocation in one parent and are
non-mosaic. Interestingly, there are hot spots for the chromosomal breakpoints
involved.
CONCLUSIONS: Complex sSMC need to be considered in diagnostics, especially in
non-mosaic, centric minute shaped sSMC. As yet three complex-sSMC-associated
syndromes are identified. As recurrent breakpoints in the complex sSMC were
characterized, it is to be expected that more syndromes are identified in this
subgroup of sSMC. Overall, complex sSMC emphasize once more the importance of
detailed cytogenetic analyses, especially in patients with idiopathic mental
retardation. Constitutional t(11;22)(q23;q11) is the most frequent recurrent non-Robertsonian
translocation in humans. Balanced carriers of t(11;22) usually manifest no
clinical symptoms, and are often identified after the birth of offspring with an
unbalanced form of this translocation, known as Emanuel syndrome. To determine
the prevalence of the disorder, we sent surveillance questionnaires to 735 core
hospitals in Japan. The observed number of Emanuel syndrome cases was 36 and
that of t(11;22) balanced translocation carriers, 40. On the basis of the de
novo t(11;22) translocation frequency in sperm from healthy men, we calculated
the frequency of the translocations in the general population. Accordingly, the
prevalence of Emanuel syndrome was estimated at 1 in 110,000. Based on this
calculation, the estimated number of Emanuel syndrome cases in Japan is 1063 and
of t(11;22) balanced translocation carriers, 16,604, which are much higher than
the numbers calculated from the questionnaire responses. It is possible that
this discordance is partly attributable to a lack of disease identification.
Further efforts should be made to increase the awareness of Emanuel syndrome to
ensure a better quality of life for affected patients and their families. Emanuel syndrome is associated with supernumerary chromosome, which consists of
the extra genetic material from chromosome 11 and 22. The frequency of this
syndrome has been reported as 1 in 110,000. It is a rare anomaly associated with
multiple systemic malformations such as micrognathia and congenital heart
disease. In addition, patients with Emanuel syndrome may have seizure disorders.
We experienced anesthetic management of a patient with Emanuel syndrome who
underwent palatoplasty. This patient had received tracheotomy due to
micrognathia. In addition, he had atrial septal defect, mild pulmonary artery
stenosis, and cleft palate. Palatoplasty was performed without any complication
during anesthesia. Close attention was directed to cardiac function, seizure,
and airway management. Emanuel syndrome (ES) is the most frequent type of recurrent non‑Robertsonian
translocation that is characterized by numerous anomalies. Over 100 patients
with ES have been described in the literature. The phenotype of this syndrome
varies but often consists of facial dysmorphism, microcephaly, severe
intellectual disability, developmental retardation, congenital heart disease and
genital anomalies. The present study describes a 2‑year‑old boy with multiple
malformations, including facial dysmorphism, severe intellectual disability,
growth retardation, congenital heart disease, cleft lip and palate, genital
malformation (micropenis), amblyopia, thymic dysplasia and hearing impairment.
The karyotype of the patient was 47,XY,+del(22)(q13), and the maternal karyotype
was 46,XX,t(11;22)(q25;q13),9qh‑,15p+. Single‑nucleotide polymorphism‑array
analysis of the proband indicated a partial duplication of chromosomes 22 and 11
at 22q11.1‑q11.21 and 11q23.3‑q25, respectively, which confirmed the diagnosis
of ES. To date, no cases of ES have been reported in mainland China. The present
case further emphasizes the necessity and importance of high‑resolution
techniques for genetic diagnosis and for subsequent genetic counseling. The
present study contributed to the phenotypic delineation of ES and confirmed the
first ES patient in mainland China. OBJECTIVE: To carry out genetic analysis for a fetus with Dandy-Walker
malformation and provide prenatal diagnosis for its parents during the
subsequent pregcy.
METHODS: Routine G-banding was carried out to analyze the karyotype of the fetus
and its parents, and next-generation sequencing (NGS) and fluorescence in situ
hybridization (FISH) were used to verify the result.
RESULTS: The father showed a normal karyotype, while the mother was found to
carry a balanced t(11; 22) (q23; q11) translocation. NGS and FISH analysis
verified that the supernumerary marker chromosome carried by the fetus was
der(22) t(11; 22) (q23;q11). The fetus was diagnosed with Emanuel syndrome.
During the next pregcy, the fetus was found to carry the same balanced
translocation as its mother. After genetic counseling, the couple decided to
continue with the pregcy, and eventually delivered a healthy baby.
CONCLUSION: A fetal case of Emanuel syndrome has been identified. The derivative
der(22) t(11; 22)(q23; q11) chromosome probably underlies the Dandy-Walker
malformation in the fetus. Combined cytogenetic and molecular analyses can
attain a more precise diagnosis for fetal abnormalities detected by
ultrasonography. |
Is a CpG island methylator phenotype involved in ependymomas? | Although devoid of recurrent single nucleotide variants and focal copy number aberrations, poor-prognosis hindbrain ependymomas exhibit a CpG island methylator phenotype | Epigenetic alterations, including methylation, have been shown to be an
important mechanism of gene silencing in cancer. Ependymoma has been well
characterized at the DNA copy number and mRNA expression levels. However little
is known about DNA methylation changes. To gain a more global view of the
methylation profile of ependymoma we conducted an array-based analysis. Our data
demonstrated tumors to segregate according to their location in the CNS, which
was associated with a difference in the global level of methylation.
Supratentorial and spinal tumors displayed significantly more hypermethylated
genes than posterior fossa tumors, similar to the 'CpG island methylator
phenotype' (CIMP) identified in glioma and colon carcinoma. This hypermethylated
profile was associated with an increase in expression of genes encoding for
proteins involved in methylating DNA, suggesting an underlying mechanism. An
integrated analysis of methylation and mRNA expression array data allowed us to
identify methylation-induced expression changes. Most notably genes involved in
the control of cell growth and death and the immune system were identified,
including members of the JNK pathway and PPARG. In conclusion, we have generated
a global view of the methylation profile of ependymoma. The data suggests
epigenetic silencing of tumor suppressor genes is an important mechanism in the
pathogenesis of supratentorial and spinal, but not posterior fossa ependymomas.
Hypermethylation correlated with a decrease in expression of a number of tumor
suppressor genes and pathways that could be playing an important role in tumor
pathogenesis. Author information:
(1)1] Developmental & Stem Cell Biology Program, Arthur and Sonia Labatt Brain
Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario M5G
1L7, Canada [2] Laboratory Medicine and Pathobiology, University of Toronto,
Toronto, Ontario M5S 1A8, Canada [3] Division of Neurosurgery, University of
Toronto, Toronto, Ontario M5S 1A8, Canada [4].
(2)1] Division of Pediatric Neurooncology, German Cancer Research Center (DKFZ),
69120 Heidelberg, Germany [2] Department of Pediatric Oncology, Hematology and
Immunology, University of Heidelberg, Heidelberg 69120, Germany [3] German
Cancer Consortium (DKTK), Heidelberg 69120, Germany [4].
(3)1] German Cancer Consortium (DKTK), Heidelberg 69120, Germany [2] Division of
Molecular Genetics, German Cancer Research Center (DKFZ), Heidelberg 69120,
Germany.
(4)1] German Cancer Consortium (DKTK), Heidelberg 69120, Germany [2] Division of
Theoretical Bioinformatics, German Cancer Research Center (DKFZ), Heidelberg
69120, Germany.
(5)Department of Molecular Genetics, Banting and Best Department of Medical
Research, The Donnelly Centre, University of Toronto, Toronto, Ontario M4N 1X8,
Canada.
(6)1] German Cancer Consortium (DKTK), Heidelberg 69120, Germany [2] Genome
Biology, European Molecular Biology, Laboratory Meyerhofstr. 1, Heidelberg
69117, Germany.
(7)1] Developmental & Stem Cell Biology Program, Arthur and Sonia Labatt Brain
Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario M5G
1L7, Canada [2] Laboratory Medicine and Pathobiology, University of Toronto,
Toronto, Ontario M5S 1A8, Canada.
(8)Developmental & Stem Cell Biology Program, Arthur and Sonia Labatt Brain
Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario M5G
1L7, Canada.
(9)Department of Genetics, Norris Cotton Cancer Center, Dartmouth Medical
School, Lebanon, New Hampshire 03756, USA.
(10)1] Division of Pediatric Neurooncology, German Cancer Research Center
(DKFZ), 69120 Heidelberg, Germany [2] German Cancer Consortium (DKTK),
Heidelberg 69120, Germany.
(11)1] German Cancer Consortium (DKTK), Heidelberg 69120, Germany [2] Division
of Bioinformatics, German Cancer Research Center (DKFZ), Heidelberg 69120,
Germany.
(12)Department of Neurology, Harvard Medical School, Children's Hospital Boston,
MIT, Boston, Massachusetts 02115, USA.
(13)Department of Pathology, The University of Texas MD Anderson Cancer Center,
Houston, Texas 77030, USA.
(14)1] Ontario Cancer Institute, Princess Margaret Cancer Centre-University
Health Network, Toronto, Ontario M5G 1L7, Canada [2] Ontario Institute for
Cancer Research, Toronto, Ontario M5G 1L7, Canada.
(15)Cancer Epigenetics Discovery Performance Unit, GlaxoSmithKline
Pharmaceuticals, Collegeville, Pennsylvania 19426, USA.
(16)Department of Oncogenomics, Academic Medical Center, Amsterdam 1105, The
Netherlands.
(17)1] Department of Pediatric Oncology, Hematology and Immunology, University
of Heidelberg, Heidelberg 69120, Germany [2] German Cancer Consortium (DKTK),
Heidelberg 69120, Germany [3] CCU Pediatric Oncology, German Cancer Research
Center (DKFZ), Heidelberg 69120, Germany.
(18)1] Centre for High-Throughput Biology, Department of Microbiology &
Immunology, University of British Columbia, Vancouver, V6T 1Z4 British Columbia,
Canada [2] Canada's Michael Smith Genome Sciences Centre, BC Cancer Agency,
Vancouver, British Columbia V5Z 1L3, Canada.
(19)1] Canada's Michael Smith Genome Sciences Centre, BC Cancer Agency,
Vancouver, British Columbia V5Z 1L3, Canada [2] Department of Medical Genetics,
University of British Columbia, Vancouver, British Columbia V6H 3N1, Canada.
(20)Department of Pediatrics and National Capital Consortium, Uniformed Services
University, Bethesda, Maryland 20814, USA.
(21)Department of Neurosurgery, University of Utah School of Medicine, Salt Lake
City, Utah 84132, USA.
(22)Pediatric Neurosurgery, Catholic University Medical School, Gemelli
Hospital, Rome 00168, Italy.
(23)Department of Neurology and Neurological Sciences, Stanford University
School of Medicine, Stanford, California 94305, USA.
(24)Department of Pediatrics, Virginia Commonwealth, Richmond, Virginia
23298-0646, USA.
(25)Department of Pathology, University of Warsaw, Children's Memorial Health
Institute University of Warsaw, Warsaw 04-730, Poland.
(26)Division of Anatomical Pathology, Department of Pathology and Molecular
Medicine, McMaster University, Hamilton General Hospital, Hamilton, Ontario L8S
4K1, Canada.
(27)1] Department of Pediatric Oncology, Hematology and Immunology, University
of Heidelberg, Heidelberg 69120, Germany [2] German Cancer Consortium (DKTK),
Heidelberg 69120, Germany.
(28)1] German Cancer Consortium (DKTK), Heidelberg 69120, Germany [2] Department
of Neuropathology Ruprecht-Karls-University Heidelberg, Institute of Pathology,
Heidelberg 69120, Germany.
(29)1] University of Michigan Cell and Developmental Biology, Ann Arbor,
Michigan 48109-2200, USA [2] Department of Neurosurgery, University of Michigan
Medical School, Ann Arbor, Michigan 48109, USA.
(30)Department of Neurosurgery, University of Michigan Medical School, Ann
Arbor, Michigan 48109, USA.
(31)Department of Neurosurgery, University of California San Francisco, San
Francisco, California 94143-0112, USA.
(32)Departments of Neurology, Pediatrics, and Neurosurgery, University of
California, San Francisco, The Helen Diller Family Cancer Research Building, San
Francisco, California 94158, USA.
(33)1] Developmental & Stem Cell Biology Program, Arthur and Sonia Labatt Brain
Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario M5G
1L7, Canada [2] Department of Neuro-oncology, The Hospital for Sick Children,
Toronto, Ontario M5G 1X8, Canada.
(34)Department of Haematology and Oncology, The Hospital for Sick Children,
Toronto, Ontario M5G 1X8, Canada.
(35)1] Developmental & Stem Cell Biology Program, Arthur and Sonia Labatt Brain
Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario M5G
1L7, Canada [2] Laboratory Medicine and Pathobiology, University of Toronto,
Toronto, Ontario M5S 1A8, Canada [3] Division of Neurosurgery, University of
Toronto, Toronto, Ontario M5S 1A8, Canada.
(36)Departments of Pediatrics and Human Genetics, McGill University and the
McGill University Health Center Research Institute, Montreal, Quebec H3Z 2Z3,
Canada.
(37)Department of Neuro-oncology, The Hospital for Sick Children, Toronto,
Ontario M5G 1X8, Canada.
(38)Genome Biology, European Molecular Biology, Laboratory Meyerhofstr. 1,
Heidelberg 69117, Germany.
(39)1] Ontario Cancer Institute, Princess Margaret Cancer Centre-University
Health Network, Toronto, Ontario M5G 1L7, Canada [2] Ontario Institute for
Cancer Research, Toronto, Ontario M5G 1L7, Canada [3] Department of Medical
Biophysics, University of Toronto, Toronto, Ontario M5G 1X8, Canada.
(40)1] Developmental & Stem Cell Biology Program, Arthur and Sonia Labatt Brain
Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario M5G
1L7, Canada [2] Laboratory Medicine and Pathobiology, University of Toronto,
Toronto, Ontario M5S 1A8, Canada [3] Division of Neurosurgery, University of
Toronto, Toronto, Ontario M5S 1A8, Canada [4] Department of Molecular Genetics,
University of Toronto, Toronto, Ontario M5S 1A8, Canada.
(41)1] Division of Pediatric Neurooncology, German Cancer Research Center
(DKFZ), 69120 Heidelberg, Germany [2] Department of Pediatric Oncology,
Hematology and Immunology, University of Heidelberg, Heidelberg 69120, Germany
[3] German Cancer Consortium (DKTK), Heidelberg 69120, Germany.
(42)1] German Cancer Consortium (DKTK), Heidelberg 69120, Germany [2] University
of Michigan Cell and Developmental Biology, Ann Arbor, Michigan 48109-2200, USA
[3] CCU Neuropathology, German Cancer Research Center (DKFZ), Heidelberg 69120,
Germany. Here, we review the recent literature on molecular discoveries in ependymomas
and pediatric diffuse gliomas. Ependymomas can now be categorized into three
location-related subgroups according to their biological profile: posterior
fossa ependymomas, group A (PFA) and B (PFB), and supratentorial ependymomas.
Although no recurrently mutated genes were found throughout these groups of
ependymomas, PFA exhibited a CpG island methylator phenotype, PFB was associated
with extensive chromosomal aberrations, and the C11orf95-RELA fusion gene was
frequently observed in supratentorial ependymomas. Meanwhile, it has now become
apparent that pediatric diffuse gliomas have a distinct genetic status from
their adult counterparts, even though they share an indistinguishable histology.
In pediatric low-grade diffuse gliomas, an intragenic duplication of the portion
of FGFR1 encoding the tyrosine kinase domain (TKD) and rearrangements of
MYB/MYBL1 were found recurrently and mutually exclusively. As for non-brainstem
high-grade tumors, in addition to H3F3A, TP53, and ATRX mutations, which were
frequently observed in older children, recurrent fusions involving NTRK1, NTRK2,
and NTRK3 were reported in infants younger than 3 years of age. Moreover, in
diffuse intrinsic pontine gliomas (DIPG), recurrent somatic mutations of ACVR1
were found in association with HIST1H3B mutations. |
Are loop domains preserved upon cohesin loss? | No. Degradation of cohesin leads to elimination of loop domains. Neither compartment domains nor histone marks are affected. Loss of loop domains does not lead to widespread ectopic gene activation but does affect a significant minority of active genes. | Author information:
(1)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA; Department of Structural Biology, Stanford
University School of Medicine, Stanford, CA 94305, USA.
(2)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA.
(3)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA; Center for Theoretical Biological Physics,
Rice University, Houston, TX 77030, USA.
(4)Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA.
(5)Lymphocyte Nuclear Biology, NIAMS, NIH, Bethesda, MD 20892, USA.
(6)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Center for Theoretical Biological Physics, Rice University, Houston,
TX 77030, USA; Department of Computer Science, Stanford University, Stanford, CA
94305, USA.
(7)Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA; Department of
Pathology and Center for Cancer Research, Massachusetts General Hospital and
Harvard Medical School, Boston, MA 02114, USA.
(8)Department of Chemistry, New York University, New York, NY 10003, USA.
(9)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Departments of Computer Science and Computational and Applied
Mathematics, Rice University, Houston, TX 77030, USA.
(10)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA; Departments of Computer Science and
Computational and Applied Mathematics, Rice University, Houston, TX 77030, USA;
Medical Scientist Training Program, Baylor College of Medicine, Houston, TX
77030, USA.
(11)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Medicine, University of California, San Diego, La
Jolla, CA 92037, USA.
(12)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA;
Department of Medicine, University of California, San Diego, La Jolla, CA 92037,
USA.
(13)Department of Chemistry, New York University, New York, NY 10003, USA;
Courant Institute of Mathematical Sciences, New York University, New York, NY
10012, USA; NYU-ECNU Center for Computational Chemistry, NYU Shanghai, Shanghai
200062, China.
(14)Lymphocyte Nuclear Biology, NIAMS, NIH, Bethesda, MD 20892, USA; Center of
Cancer Research, NCI, NIH, Bethesda, MD 20892, USA.
(15)Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA; Department of
Biology, MIT, Cambridge, MA 02139, USA; Department of Systems Biology, Harvard
Medical School, Boston, MA 02115, USA.
(16)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA; Center for Theoretical Biological Physics,
Rice University, Houston, TX 77030, USA; Broad Institute of MIT and Harvard,
Cambridge, MA 02139, USA; Departments of Computer Science and Computational and
Applied Mathematics, Rice University, Houston, TX 77030, USA. Electronic
address: [email protected]. |
Which workflow in Bioconductor has been developed for accessing human RNA-seq samples? | The recount2 resource is composed of over 70,000 uniformly processed human RNA-seq samples spanning TCGA and SRA, including GTEx. | The recount2 resource is composed of over 70,000 uniformly processed human
RNA-seq samples spanning TCGA and SRA, including GTEx. The processed data can be
accessed via the recount2 website and the recount Bioconductor package. This
workflow explains in detail how to use the recount package and how to integrate
it with other Bioconductor packages for several analyses that can be carried out
with the recount2 resource. In particular, we describe how the coverage count
matrices were computed in recount2 as well as different ways of obtaining public
metadata, which can facilitate downstream analyses. Step-by-step directions show
how to do a gene-level differential expression analysis, visualize base-level
genome coverage data, and perform an analyses at multiple feature levels. This
workflow thus provides further information to understand the data in recount2
and a compendium of R code to use the data. |
What does VBP15 do to skeletal muscle? | VBP15 protects and promotes efficient repair of skeletal muscle cells. Potent inhibition of NF-κB is mediated through protein interactions of the glucocorticoid receptor, however VBP15 shows significantly reduced hormonal receptor transcriptional activity. In DMD model mice it improves muscle strength, live-imaging and pathology through both preventive and post-onset intervention regimens. | Absence of dystrophin makes skeletal muscle more susceptible to injury,
resulting in breaches of the plasma membrane and chronic inflammation in
Duchenne muscular dystrophy (DMD). Current management by glucocorticoids has
unclear molecular benefits and harsh side effects. It is uncertain whether
therapies that avoid hormonal stunting of growth and development, and/or
immunosuppression, would be more or less beneficial. Here, we discover an oral
drug with mechanisms that provide efficacy through anti-inflammatory signaling
and membrane-stabilizing pathways, independent of hormonal or immunosuppressive
effects. We find VBP15 protects and promotes efficient repair of skeletal muscle
cells upon laser injury, in opposition to prednisolone. Potent inhibition of
NF-κB is mediated through protein interactions of the glucocorticoid receptor,
however VBP15 shows significantly reduced hormonal receptor transcriptional
activity. The translation of these drug mechanisms into DMD model mice improves
muscle strength, live-imaging and pathology through both preventive and
post-onset intervention regimens. These data demonstrate successful improvement
of dystrophy independent of hormonal, growth, or immunosuppressive effects,
indicating VBP15 merits clinical investigation for DMD and would benefit other
chronic inflammatory diseases. |
What is filgotinib? | Filgotinib is an oral selective Janus kinase inhibitor. It has been tested in patients with rheumatoid arthritis and Chroni's disease, and has been shown to be safe and efficacious. | BACKGROUND: Filgotinib (GLPG0634, GS-6034) is a once-daily, orally administered,
Janus kinase 1 (JAK1)-selective inhibitor. The FITZROY study examined the
efficacy and safety of filgotinib for the treatment of moderate-to-severe
Crohn's disease.
METHODS: We did a randomised, double-blind, placebo-controlled phase 2 study,
which recruited patients from 52 centres in nine European countries. We enrolled
eligible patients aged 18-75 years with a documented history of ileal, colonic,
or ileocolonic Crohn's disease for 3 months or more before screening, as
assessed by colonoscopy and supported by histology, and a Crohn's Disease
Activity Index (CDAI) score during screening between 220 and 450 inclusive.
Patients were randomly assigned (3:1) to receive filgotinib 200 mg once a day or
placebo for 10 weeks. Patients were stratified according to previous anti-tumour
necrosis factor alpha exposure, C-reactive protein concentration at screening
(≤10 mg/L or >10 mg/L), and oral corticosteroid use at baseline, using an
interactive web-based response system. The primary endpoint was clinical
remission, defined as CDAI less than 150 at week 10. After week 10, patients
were assigned based on responder status to filgotinib 100 mg once a day,
filgotinib 200 mg once a day, or placebo for an observational period lasting a
further 10 weeks. The filgotinib and placebo treatment groups were compared
using ANCOVA models and logistic regression models containing baseline values
and randomisation stratification factors as fixed effects. Analyses were done on
the intention-to-treat non-responder imputation set. The trial was registered at
ClinicalTrials.gov, number NCT02048618.
FINDINGS: Between Feb 3, 2014, and July 10, 2015, we enrolled 174 patients with
active Crohn's disease confirmed by centrally read endoscopy (130 in the
filgotinib 200 mg group and 44 in the placebo group). In the intention-to-treat
population, 60 (47%) of 128 patients treated with filgotinib 200 mg achieved
clinical remission at week 10 versus ten (23%) of 44 patients treated with
placebo (difference 24 percentage points [95% CI 9-39], p=0·0077). In a pooled
analysis of all periods of filgotinib and placebo exposure over 20 weeks,
serious treatment-emergent adverse effects were reported in 14 (9%) of 152
patients treated with filgotinib and three (4%) of 67 patients treated with
placebo.
INTERPRETATION: Filgotinib induced clinical remission in significantly more
patients with active Crohn's disease compared with placebo, and had an
acceptable safety profile.
FUNDING: Galapagos. OBJECTIVES: To evaluate the efficacy and safety of different doses of
filgotinib, an oral Janus kinase 1 inhibitor, as monotherapy in patients with
active rheumatoid arthritis (RA) and previous inadequate response to
methotrexate (MTX).
METHODS: In this 24-week phase IIb study, patients with moderately to severely
active RA were randomised (1:1:1:1) to receive 50, 100 or 200 mg filgotinib once
daily, or placebo, after a ≥4-week washout from MTX. The primary end point was
the percentage of patients achieving an American College of Rheumatology (ACR)20
response at week 12.
RESULTS: Overall, 283 patients were randomised and treated. At week 12,
significantly more patients receiving filgotinib at any dose achieved ACR20
responses versus placebo (≥65% vs 29%, p<0.001). For other key end points at
week 12 (ACR50, ACR70, ACR-N, Disease Activity Score based on 28 joints and C
reactive protein, Clinical Disease Activity Index, Simplified Disease Activity
Index and Health Assessment Questionnaire-Disability Index) significant
differences from baseline in favour of filgotinib 100 and 200 mg versus placebo
were seen; responses were maintained or improved through week 24. Rapid onset of
action was observed for most efficacy end points. Dose-dependent increases in
haemoglobin were observed. The percentage of patients with treatment-emergent
adverse events (TEAE) was similar in the placebo and filgotinib groups (∼40%).
Eight patients on filgotinib and one on placebo had a serious TEAE, and four
patients, all of whom received filgotinib, experienced a serious infection. No
tuberculosis or opportunistic infections were reported.
CONCLUSIONS: Over 24 weeks, filgotinib as monotherapy was efficacious in
treating the signs and symptoms of active RA, with a rapid onset of action.
Filgotinib was generally well tolerated.
TRIAL REGISTRATION NUMBER: NCT01894516. |
What is Q-nexus? | Q-nexus is a comprehensive and efficient analysis pipeline designed for ChIP-nexus. | Author information:
(1)Institute for Medical and Human Genetics, Charité-Universitätsmedizin Berlin,
Augustenburger Platz 1, Berlin, 13353, Germany.
(2)Berlin Brandenburg Center for Regenerative Therapies (BCRT),
Charité-Universitätsmedizin Berlin, Augustenburger Platz 1, Berlin, 13353,
Germany.
(3)Centre for Genomic Regulation (CRG), The Barcelona Institute of Science and
Technology, Dr. Aiguader 88, Barcelona, 08003, Spain.
(4)Universitat Pompeu Fabra (UPF), Barcelona, Spain.
(5)Faculty of Biology, Johannes Gutenberg University Mainz, Ackermannweg 4,
Mainz, 55128, Germany.
(6)Institute of Molecular Biology, Ackermannweg 4, Mainz, 55128, Germany.
(7)Institute for Bioinformatics, Department of Mathematics and Computer Science,
Freie Universität Berlin, Arnimallee 14, Berlin, 14195, Germany.
(8)Labor für Pädiatrische Molekularbiologie, Charité-Universitätsmedizin Berlin,
Augustenburger Platz 1, Berlin, 13353, Germany.
(9)Institute for Medical and Human Genetics, Charité-Universitätsmedizin Berlin,
Augustenburger Platz 1, Berlin, 13353, Germany. [email protected].
(10)Berlin Brandenburg Center for Regenerative Therapies (BCRT),
Charité-Universitätsmedizin Berlin, Augustenburger Platz 1, Berlin, 13353,
Germany. [email protected].
(11)Institute for Bioinformatics, Department of Mathematics and Computer
Science, Freie Universität Berlin, Arnimallee 14, Berlin, 14195, Germany.
[email protected].
(12)Max Planck Institute for Molecular Genetics, Inhestr. 63-73, Berlin, 14195,
Germany. [email protected].
(13)Current address: The Jackson Laboratory for Genomic Medicine, 10 Discovery
Drive, Farmington, 06032, CT, USA. [email protected]. |
What is the mechanism of action of Tisagenlecleucel? | Tisagenlecleucel CD19-directed chimeric antigen receptor T cells (CART19) product that cause reprogramming a patient's own T cells with a transgene encoding a chimeric antigen receptor to identify and eliminate CD19-expressing cells. Its is is approved for children and young adults with relapsed or refractory B-cell acute lymphoblastic leukemia. | An expert panel recommended approval of Novartis's experimental chimeric antigen
T-cell therapy, tisagenlecleucel, for children and young adults with relapsed or
refractory B-cell acute lymphoblastic leukemia. The therapy would be the first
of its kind approved for cancer and has the potential to transform standard of
care for advanced blood cancers. The first chimeric antigen receptor T-cell therapy, tisagenlecleucel, received
FDA approval for the treatment of patients up to 25 years of age with B-cell
precursor acute lymphoblastic leukemia who haven't responded to standard therapy
or who have relapsed at least twice. PURPOSE OF REVIEW: In this review, we discuss the most recent developments in
gene-editing technology and discuss their application to adoptive T cell
immunotherapy.
RECENT FINDINGS: Engineered T cell therapies targeting cancer antigens have
demonstrated significant efficacy in specific patient populations. Most
impressively, CD19-directed chimeric antigen receptor T cells (CART19) have led
to impressive responses in patients with B-cell leukemia and lymphoma. CTL019,
or KYMRIAH™ (tisagenlecleucel), a CD19 CAR T cell product developed by Novartis
and the University of Pennsylvania, was recently approved for clinical use by
the Food and Drug Administration, representing a landmark in the application of
adoptive T cell therapies. As CART19 enters routine clinical use, improving the
efficacy of this exciting platform is the next step in broader application.
Novel gene-editing technologies like CRISPR-Cas9 allow facile editing of
specific genes within the genome, generating a powerful platform to further
optimize the activity of engineered T cells. Autologous, patient-specific chimeric antigen receptor T-cell (CART) therapy has
emerged as a powerful and potentially curative therapy for cancer, especially
for CD19-positive hematological maligcies. Indeed, on August 30, 2017, the
University of Pennsylvania-designed CD19-directed CART (CART-19) cell therapy
(CTL019, tisagenlecleucel-t, Kymriah - Novartis) became the first CART therapy
approved by the Food and Drug Administration (FDA) for acute lymphoblastic
leukemia. However, the development of CART technology and its wider application
is partly limited by the patient-specific nature of such a platform and by the
time required for manufacturing. The efficacious generation of universal
allogeneic CART cells would overcome these limitations and represent a major
advance in the field. However, several obstacles in the generation of universal
CART cells need to be overcome, namely the risk of CART rejection and the risk
of graft-versus-host disease mediated by the allogeneic CART. In this review, we
discuss the different strategies being employed to generate universal CART and
provide our perspective on the successful development of a truly off-the-shelf
CART product. |
What is LHON, also known as Lebers syndrome? | Leber's hereditary optic neuropathy (LHON) is a common inherited mitochondrial disorder that is characterized by the degeneration of the optic nerves, leading to vision loss. | Leber hereditary optic neuropathy (LHON) presents with sudden onset of visual
loss mainly in young adult males. LHON is not uncommon in Australia, accounting
for 2% of invalid blind pensions. We have identified 20 unrelated families
carrying mitochondrial DNA mutations associated with LHON and 135 of 291
individuals with documented LHON are currently alive in Australia. The mean age
of onset of visual loss for males was 26 years and for females 27 years, with a
range from six to 65 years. The mean risk of visual loss was 20% for males and
4% for females. There are over 1750 male and female carriers living in Australia
who have not yet lost vision; 600 carriers are under 24 years of age. The
expected number of new cases of blindness from LHON is three to four per year. Pre-excitation syndrome is common in families with Leber's hereditary optic
neuropathy (LHON). 24 Finnish families with LHON were screened for the 11778 and
the 3460 mitochondrial DNA mutations. 5 of 30 individuals with LHON and the
11778 mutation had the Wolff-Parkinson-White pre-excitation syndrome. None of 10
with the 3460 mutation or of 11 with "other" mutations had this syndrome.
Overall, 5 of 51 LHON patients and 9 of 112 symptom-free maternal relatives had
Wolff-Parkinson-White syndrome (9%). In paternal relatives, the frequency was
1.6%. Mitochondrial DNA causal for LHON may contribute to pre-excitation
syndrome. We review the main features of human mitochondrial function and structure, and
in particular mitochondrial transcription, translation, and replication cycles.
Furthermore, some pecularities such as mitochondria's high polymorphism, the
existence of mitochondrial pseudogenes, and the various considerations to take
into account when studying mitochondrial diseases will also be mentioned.
Mitochondrial syndromes mostly affecting the nervous system have, during the
past few years, been associated with mitochondrial DNA (mt DNA) alterations such
as deletions, duplications, mutations and depletions. We suggest a possible
classification of mitochondrial diseases according to the kind of mt DNA
mutations: structural mitochondrial gene mutation as in LHON (Leber's Hereditary
Optic Neuropathy) and NARP (Neurogenic muscle weakness, Ataxia and Retinitis
Pigmentosa) as well as some cases of Leigh's syndrome; transfer RNA and
ribosomal RNA mitochondrial gene mutation as in MELAS (Mitochondrial
Encephalomyopathy, Lactic Acidosis and Strokelike Episodes) or MERRF (Myoclonic
Epilepsy with Ragged Red Fibers) or deafness with aminoglycoside; structural
with transfer RNA mitochondrial gene mutations as observed in large-scale
deletions or duplications in Kearns-Sayre syndrome, Pearson's syndrome, diabetes
mellitus with deafness, and CPEO (Chronic Progressive External Ophtalmoplegia).
Depletions of the mt DNA may also be classified in this category. Even though
mutations are generally maternally inherited, most of the deletions are
sporadic. However, multiple deletions or depletions may be transmitted in a
mendelan trait which suggests that nuclear gene products play a primary role in
these processes. The relationship between a mutation and a particular phenotype
is far from being fully understood. Gene dosage and energic threshold, which are
tissue-specific, appear to be the best indicators. However, the recessive or
domit behavior of both the wild type or the mutated genome appears to play a
significant role, which can be verified with in vitro studies. BACKGROUND: Mitochondrial encephalomyopathies are heterogeneous diseases with
common clinical features of muscle and/or the central nervous system. Although
molecular and histological diagnoses have been established, imaging modalities
for the functioning evaluation of these patients are still obscure. In this
study, we tried to use 99mTc-HMPAO brain SPECT images to analyze various
mitochondrial encephalomyopathies.
METHODS: We examined 99mTc-HMPAO Brain SPECT studies of 15 patients with various
types of mitochondrial encephalomyopathy (3 Leber's hereditary optic neuropathy
(LHON), 4 Kearns-Sayre syndrome (KSS), 4 mitochondrial myopathy, encephalopathy,
lactic acidosis, and stroke-like episodes (MELAS), 3 myoclonic epilepsy and
ragged-red fiber disease (MERRF), and 1 Leigh syndrome), diagnosed by molecular
studies.
RESULTS: The results of our studies show obviously decreased radiotracer
accumulation in the parieto-temporal regions after stroke-like episodes in
patients with MELAS and MERRF: Relatively diminished brain perfusion in patients
with KSS is noted, probably secondary to severe encephalomyopathy or subdural
effusion. However, there seems to be no significant correlation between the
clinical manifestations and the imaging findings in LHON and Leigh.
CONCLUSIONS: 99mTc-HMPAO Brain SPECT is useful in the diagnosing and assessment
of the progress of MERRF, MELAS and KSS. However, its role in LHON and Leigh
syndrome seems to be debatable. DNA sequence analysis of the gene encoding subunit 6 of the
NADH-ubiquinone-oxidoreductase complex (ND6) in human mitochondria was performed
in 25 independent patients who suffer from Lebers hereditary optic neuropathy
(LHON). In 10 cases the well-known LHON mutation at nucleotide position (np)
14484 was detected. Furthermore, silent substitutions at np14167 and np14527 and
missense mutations at np14498, np14564, np14568, and np14582 were found in
individual patients. The np14498 and np14568 mutations were found in patients
who present a typical clinical picture and course of LHON but lack any of the
canonical mtDNA mutations. The np14568 mutation, which replaces a moderately
conserved glycine by a serine residue, was observed in a single male patient and
subsequently excluded in 175 independent controls. The mutation at np14498,
which replaces an evolutionarily highly conserved tyrosine with a cysteine, was
found in a multigeneration family with four affected members, the eldest
carrying a heteroplasmic mixture of mutated and wildtype mtDNA molecules. None
of 170 analyzed control subjects carried this mutation. These findings provide
evidence that several allelic ND6 gene mutations may be involved in Lebers
hereditary optic neuropathy. Wolfram or DIDMOAD (Diabetes Insipidus, Diabetes Mellitus, Optic Atrophy and
Deafness) syndrome, which has long been known as an autosomal-recessive
disorder, has recently been proposed to be a mitochondrial-mediated disease with
either a nuclear or a mitochondrial genetic background. The phenotypic
characteristics of the syndrome resemble those found in other mitochondrial
(mt)DNA mediated disorders such as Leber's hereditary optic neuropathy (LHON) or
maternally inherited diabetes and deafness (MIDD). Therefore, we looked for
respective mtDNA alterations in blood samples from 7 patients with DIDMOAD
syndrome using SSCP-analysis of PCR-amplified fragments, encompassing all
mitochondrial ND and tRNA genes, followed by direct sequencing. Subsequently, we
compared mtDNA variants identified in this disease group with those detected in
a group of LHON patients (n = 17) and in a group of 69 healthy controls. We
found that 4/7 (57%) DIDMOAD patients harbored a specific set of point mutations
in tRNA and ND genes including the so-called class II or secondary LHON
mutations at nucleotide positions (nps) 4216 and 4917 (haplogroup B). In
contrast, LHON-patients were frequently (10/17, 59%) found in association with
another cluster of mtDNA variants including the secondary LHON mutations at nps
4216 and 13708 and further mtDNA polymorphisms in ND genes (haplogroup A),
overlapping with haplogroup B only by variants at nps 4216 and 11251. The
frequencies of both haplogroups were significantly lower in the control group
versus the respective disease groups. We propose that haplogroup B represents a
susceptibility factor for DIDMOAD which, by interaction with further exogeneous
or genetic factors, might increase the risk for disease. The authors present a case report of 26 years old man with bilateral optic nerve
neuropathy. Detection of heteroplasmic mutation of mitochondrial DNA at G3460A
site confirmed the suspicion on Lebers hereditary optic nerve neuropathy (LHON).
Genetic and environmental factors of the disease and various accompanying
neurologic and other symptoms, which can together with the optic nerve defect
participate in the development of of the LOHN clinical pattern are discussed.
(Ref. 12.) Myocardial thickening and isolated left ventricular abnormal trabeculation
(ILVAT) have not been described in patients with Leber's hereditary optic
neuropathy (LHON) before. Wolff-Parkinson-White syndrome, myocardial thickening
and ILVAT were found by electrocardiogram, echocardiography and cardiac magnetic
resoce imaging in a 48-year-old man with bilateral, severely reduced visual
acuity since age 24 years, palpitations since age 43 years and lower limb muscle
cramps since age 47 years. Because ILVAT is frequently associated with
respiratory chain disorders, neurological investigations were initiated,
revealing the primary LHON mutation G3460A in lymphocytic mitochondrial DNA. On
the basis of the clinical and genetic data, LHON was diagnosed in the index
patient, but also in the patient's brother who showed ILVAT as well.
Wolff-Parkinson-White syndrome, myocardial thickening and ILVAT may be rare
manifestations of LHON. BACKGROUND: Leber's hereditary optic neuropathy (LHON) is a maternally inherited
optic neuropathy caused by mutations in mitochondrial DNA (mtDNA). It is also
believed that several epigenetic factors have an influence on the development of
LHON.
METHODS: A case series was observed.
RESULTS: Three patients who developed bilateral optic neuropathy are presented.
All patients had a primary LHON mutation in their mtDNA, but also a subnormal
vitamin B12 serum level at the time of presentation.
CONCLUSIONS: The clinical picture of optic neuropathy associated with vitamin
B12 deficiency shows similarity to that of LHON. Both involve the nerve fibres
of the papillomacular bundle. The present case reports suggest that optic
neuropathy in patients carrying a primary LHON mtDNA mutation may be
precipitated by vitamin B12 deficiency. Therefore, known carriers should take
care to have an adequate dietary intake of vitamin B12 and malabsorption
syndromes like those occurring in familial pernicious anaemia or after gastric
surgery should be excluded. Leber hereditary optic neuropathy (LHON) is a mitochondrial disorder
characterized by bilateral painless optic atrophy and blindness. It usually
occurs in young men in association with three major mutations in the
mitochondrial genome (mtDNA). We report a patient with a history of alcohol
abuse who developed at age 63 years visual impairment, sensorineural hearing
loss, and memory dysfunction, suggestive of Susac's syndrome. The patient
carried the heteroplasmic mt. 11778G>A mutation on the T2e mtDNA haplogroup. It
remains unclear if chronic alcohol abuse combined with the mitochondrial genetic
background prompted an aged-related neurodegeneration or deferred the onset of
the LHON disease. BACKGROUND: The aim of this paper is to describe the clinical features and
molecular findings of a unique case of Leber's hereditary optic neuropathy
(LHON)/mitochondrial encephalopathy, lactic acidosis and stroke-like episodes
(MELAS) overlap syndrome presenting as nonischemic central retinal vein
occlusion (CRVO).
METHODS: An 11-year-old Chinese girl presented with sudden onset of bilateral
blurred vision. The clinical history, imaging studies, and molecular analysis
results were reviewed. The PubMed and OVID databases were used for literature
review.
RESULTS: Nonischemic CRVO in the subject's right eye and tortuosity of small and
medium-sized retinal arterioles in the left eye were found at initial
presentation. Bilateral optic disc pallor was then noted with recovery of CRVO.
Severe headache and several stroke-like episodes occurred subsequently, with
elevated lactate levels in serum and cerebrospinal fluid. LHON/MELAS overlap
syndrome was diagnosed, and mitochondrial DNA sequencing revealed G13513A
heteroplasmic mutation. Vision was 20/30 in the right eye and 20/800 in the left
eye at the last visit.
CONCLUSIONS: Mitochondrial DNA G13513A mutation can cause LHON/MELAS overlap
syndrome. Nonischemic CRVO is a rare manifestation of LHON/MELAS. Atypical
findings in cases of LHON should raise the suspicion of overlap syndrome or
other mitochondrial diseases. Leber's hereditary optic neuropathy (LHON) is a maternally transmitted disease.
Clinically, no efficient assay protocols have been available. In this study, we
aimed to develop an oligonucleotide biochip specialized for detection of known
base substitution mutations in mitochondrial DNA causing LHON and to investigate
frequencies of LHON relevant variants in Anhui region of China. Thirty-two pairs
of oligonucleotide probes matched with the mutations potentially linked to LHON
were covalently immobilized. Cy5-lablled targets were amplified from blood DNA
samples by a multiplex PCR method. Two kinds of primary mutations 11778 G > A
and 14484 T > C from six confirmed LHON patients were interrogated to validate
this biochip format. Further, fourteen Chinese LHON pedigrees and twenty-five
unrelated healthy individuals were investigated by the LHON biochip, direct
sequencing and pyrosequencing, respectively. The biochip was found to be able
efficiently to discriminate homoplasmic and heteroplasmic mtDNA mutations in
LHON. Biochip analysis revealed that twelve of eighteen LHON symptomatic cases
from the 14 Chinese pedigree harbored the mutations either 11778G > A, 14484T >
C or 3460G <formula>></formula> A, respectively, accounting for 66.7%. The
mutation 11778G > A in these patients was homoplasmic and prevalent (55.5%, 10
of 18 cases). The mutations 3460G > A and 3394T > C were found to co-exist in
one LHON case. The mutation 13708G > A appeared in one LHON pedigree. Smaller
amount of sampling and reaction volume, easier target preparation, fast and
high-throughput were the main advantages of the biochip over direct DNA
sequencing and pyrosequencing. Our findings suggested that primary mutations of
11778G > A, 14484T > C or 3460G > A are main variants of mtDNA gene leading to
LHON in China. The biochip would easily be implemented in clinical diagnosis. BACKGROUND: Leber hereditary optic neuropathy (LHON) is an inherited form of
bilateral optic atrophy leading to the loss of central vision. The primary cause
of vision loss is mutation in the mitochondrial DNA (mtDNA), however, unknown
secondary genetic and/or epigenetic risk factors are suggested to influence its
neuropathology. In this study folate gene polymorphisms were examined as a
possible LHON secondary genetic risk factor in Iranian patients.
METHODS: Common polymorphisms in the MTHFR (C677T and A1298C) and MTRR (A66G)
genes were tested in 21 LHON patients and 150 normal controls.
RESULTS: Strong associations were observed between the LHON syndrome and C677T
(P= 0.00) and A66G (P= 0.00) polymorphisms. However, no significant association
was found between A1298C (P =0.69) and the LHON syndrome.
CONCLUSION: This is the first study that shows MTHFR C677T and MTRR A66G
polymorphisms play a role in the etiology of the LHON syndrome. This finding may
help in the better understanding of mechanisms involved in neural degeneration
and vision loss by LHON and hence the better treatment of patients. Lebers hereditary optic neuropathy (LHON) is a maternally inherited disease
characterized by subacute severe visual loss in both eyes, which usually
manifests in young adulthood. The disease has maternal inheritance due to
mitochondrial DNA mutation. The final diagnosis is genetic. There is still no
proven treatment, but there is significant progress in developments on the
genetics of the disease to reach gene therapy. In this article we review the
latest literature relevant to this disease. Author information:
(1)Institute of Genetics, School of Medicine, Zhejiang University, Hangzhou,
Zhejiang, China 2Collaborative Innovation Center for Diagnosis and Treatment of
Infectious Diseases, Zhejiang University, Hangzhou, China.
(2)Institute of Genetics, School of Medicine, Zhejiang University, Hangzhou,
Zhejiang, China 3Attardi Institute of Mitochondrial Biomedicine, Wenzhou Medical
University, Wenzhou, Zhejiang, China.
(3)Institute of Genetics, School of Medicine, Zhejiang University, Hangzhou,
Zhejiang, China 4School of Ophthalmology and Optometry, Wenzhou Medical
University, Wenzhou, Zhejiang, China.
(4)Attardi Institute of Mitochondrial Biomedicine, Wenzhou Medical University,
Wenzhou, Zhejiang, China.
(5)Attardi Institute of Mitochondrial Biomedicine, Wenzhou Medical University,
Wenzhou, Zhejiang, China 4School of Ophthalmology and Optometry, Wenzhou Medical
University, Wenzhou, Zhejiang, China.
(6)Institute of Genetics, School of Medicine, Zhejiang University, Hangzhou,
Zhejiang, China.
(7)Department of Ophthalmology, Xingtai Eye Hospital, Xingtai, Hebei, China.
(8)Department of Ophthalmology, The Third Affiliated Hospital, Xinxiang Medical
College, Xinxiang, He, China.
(9)School of Ophthalmology and Optometry, Wenzhou Medical University, Wenzhou,
Zhejiang, China.
(10)Department of Ophthalmology, Dongfang Hospital, Beijing University of
Chinese Medicine and Pharmacology, Beijing, China.
(11)Division of Human Genetics, Cincinnati Children's Hospital Medical Center,
Cincinnati, Ohio, United States.
(12)Institute of Genetics, School of Medicine, Zhejiang University, Hangzhou,
Zhejiang, China 2Collaborative Innovation Center for Diagnosis and Treatment of
Infectious Diseases, Zhejiang University, Hangzhou, China 8Division of Human
Genetics, Cincinnati Chi. BACKGROUND: Leber hereditary optic neuropathy (LHON) and mitochondrial
encephalopathy, myopathy, lactic acidosis and stroke-like episodes (MELAS)
syndromes are mitochondrially inherited disorders characterized by acute visual
failure and variable multiorgan system presentation, respectively.
MATERIALS AND METHODS: A 12-year-old girl with otherwise unremarkable medical
history presented with abrupt, painless loss of vision. Over the next few
months, she developed moderate sensorineural hearing loss, vertigo, migraines,
anhedonia and thyroiditis. Ocular examination confirmed bilateral optic nerve
atrophy. Metabolic workup documented elevated cerebrospinal fluid lactate.
Initial genetic analyses excluded the three most common LHON mutations.
Subsequently, Sanger sequencing of the entire mitochondrial DNA (mtDNA) genome
was performed.
RESULTS: Whole mtDNA sequencing revealed a pathogenic heteroplasmic mutation
m.13046T>C in MTND5 encoding the ND5 subunit of complex I. This particular
variant has previously been described in a single case report of MELAS/Leigh
syndrome (subacute necrotizing encephalopathy). Based on the constellation of
clinical symptoms in our patient, we diagnose the condition as LHON/MELAS
overlap syndrome.
CONCLUSIONS: We describe a unique presentation of LHON/MELAS overlap syndrome
resulting from a m.13046T>C mutation in a 12-year-old girl. In patients with
sudden vision loss in which three of the most prevalent LHON mitochondrial
mutations have been ruled out, molecular genetic examination should be extended
to other mtDNA-encoded subunits of MTND5 complex I. Furthermore, atypical
clinical presentations must be considered, even in well-described phenotypes. Author information:
(1)Departament de Patología Mitocondrial i Neuromuscular, Hospital Universitari
Vall d'Hebron Institut de Recerca (VHIR), Universitat Autònoma de Barcelona,
Barcelona, Spain; Centro de Investigación Biomédica en Red de Enfermedades Raras
(CIBERER), ISCIII, Barcelona, Spain.
(2)Departament de neurología, Hospital Universitari Vall d'Hebron Institut de
Recerca (VHIR), Universitat Autònoma de Barcelona, Barcelona, Spain.
(3)Translational Bioinformatics, Hospital Universitari Vall d'Hebron Institut de
Recerca (VHIR), Universitat Autònoma de Barcelona, Barcelona, Spain.
(4)Departament de neurofisiología, Hospital Universitari Vall d'Hebron Institut
de Recerca (VHIR), Universitat Autònoma de Barcelona, Barcelona, Spain.
(5)Departament d'oftalmología, Hospital Universitari Vall d'Hebron Institut de
Recerca (VHIR), Universitat Autònoma de Barcelona, Barcelona, Spain.
(6)Translational Bioinformatics, Hospital Universitari Vall d'Hebron Institut de
Recerca (VHIR), Universitat Autònoma de Barcelona, Barcelona, Spain; Institució
Catalana de Recerca i Estudis Avançats, Spain.
(7)Departament de Patología Mitocondrial i Neuromuscular, Hospital Universitari
Vall d'Hebron Institut de Recerca (VHIR), Universitat Autònoma de Barcelona,
Barcelona, Spain; Centro de Investigación Biomédica en Red de Enfermedades Raras
(CIBERER), ISCIII, Barcelona, Spain; Àrea de Genètica Clínica i Molecular,
Hospital Universitari Vall d'Hebron, Barcelona, Spain. Electronic address:
[email protected]. Leber's hereditary optic neuropathy (LHON) is a maternally inherited
mitochondrial disorder with bilateral loss of central vision primarily due to
mitochondrial DNA (mtDNA) mutations in subunits of complex I in the respiratory
chain (primary LHON mutations), while other mtDNA mutations can also be
causative. Since the first description, it is known that LHON is not restricted
to the eyes but is a multisystem disorder additionally involving the central
nervous system, ears, endocrinological organs, heart, bone marrow, arteries,
kidneys, or the peripheral nervous system. Multisystem involvement may start
before or after the onset of visual impairment. Involvement of organs other than
the eyes may be subclinical depending on age, ethnicity, and possibly the
heteroplasmy rate of the responsible primary LHON mutation. Primary LHON
mutations may rarely manifest without ocular compromise but with arterial
hypertension, various neurodegenerative diseases, or Leigh syndrome. Patients
with LHON need to be closely followed up to detect at which point organs other
than the eyes become affected. Multiorgan disease in LHON often responds more
favorably to symptomatic treatment than the ocular compromise. Author information:
(1)IRCCS Institute of Neurological Sciences of Bologna (VC, MC, CLM), Bellaria
Hospital, Bologna, Italy; Unit of Neurology (VC, CLM), Department of Biomedical
and Neuromotor Sciences (DIBINEM), University of Bologna, Bologna, Italy;
Department of Neurology (IFdC), Erasmus Medical Center, Rotterdam, the
Netherlands; Neuro-Ophthalmology Unit (AK), University of Lausanne, Jules Gonin
Eye Hospital, Lausanne, Switzerland; Department of Neurology (TK),
Friedreich-Baur-Institute, Ludwing-Maximilians-University, Munich, Germany;
Munich Cluster for Systems Neurology (SyNergy) (TK), Munich, Germany; German
Center for Neurodegenerative Diseases (DZNE) (TK), Munich, Germany; Eye Center
(WAL), Medical Center, Faculty of Medicine, University of Freiburg, Breisgau,
Germany; Departments of Ophthalmology, Neurology and Neurological Surgery (NJN),
Emory University School of Medicine, Atlanta, Georgia; Department of
Ophthalmology (CO); Referral Center for Rare Diseases OPHTARA, Hôpital Européen
Georges Pompidou, Assistance Publique-Hôpitaux de Paris, Paris, France;
Department of Ophthalmology (JWRP), University Medical Center Groningen,
University of Groningen, Groningen, the Netherlands; Doheny Eye Institute (AAS),
Los Angeles, California; Department of Ophthalmology (AAS), David Geffen School
of Medicine at UCLA, Los Angeles, California; Department of Neuro-ophthalmology
(JvE), The Rotterdam Eye Hospital, Rotterdam, the Netherlands; Rotterdam
Ophthalmic Institute (ROI) (JvE), Rotterdam, the Netherlands; Fondation
Ophtalmologique Adolphe de Rothschild (CV-C), Paris, France; School of Optometry
and Vision Sciences (MV), Cardiff University, and Cardiff Eye Clinic, University
Hospital of Wales, Cardiff, United Kingdom; Wellcome Trust Center for
Mitochondrial Research (PY-W-M), Institute of Genetic Medicine, Newcastle
University, Newcastle Upon Tyne, United Kingdom; Newcastle Eye Center (PY-W-M),
Royal Victoria Infirmary, Newcastle Upon Tyne, United Kingdom; NIHR Biomedical
Research Center at Moorfields Eye Hospital and UCL Institute of Ophthalmology
(PY-W-M), London, United Kingdom; Department of Clinical Neurosciences (PY-W-M),
School of Clinical Medicine, University of Cambridge, Cambridge, United Kingdom;
Department of Ophthalmology (PB), San Raffaele Scientific Institute, Milan,
Italy; and Studio Oculistico d'Azeglio (PB), Bologna, Italy. |
What is the ReliableGenome? | The increasing adoption of clinical whole-genome resequencing (WGS) demands for highly accurate and reproducible variant calling (VC) methods. The observed discordance between state-of-the-art VC pipelines, however, indicates that the current practice still suffers from non-negligible numbers of false positive and negative SNV and INDEL calls that were shown to be enriched among discordant calls but also in genomic regions with low sequence complexity. ReliableGenome is a method for partitioning genomes into high and low concordance regions with respect to a set of surveyed VC pipelines. It combines call sets derived by multiple pipelines from arbitrary numbers of datasets and interpolates expected concordance for genomic regions without data. | MOTIVATION: The increasing adoption of clinical whole-genome resequencing (WGS)
demands for highly accurate and reproducible variant calling (VC) methods. The
observed discordance between state-of-the-art VC pipelines, however, indicates
that the current practice still suffers from non-negligible numbers of false
positive and negative SNV and INDEL calls that were shown to be enriched among
discordant calls but also in genomic regions with low sequence complexity.
RESULTS: Here, we describe our method ReliableGenome (RG) for partitioning
genomes into high and low concordance regions with respect to a set of surveyed
VC pipelines. Our method combines call sets derived by multiple pipelines from
arbitrary numbers of datasets and interpolates expected concordance for genomic
regions without data. By applying RG to 219 deep human WGS datasets, we
demonstrate that VC concordance depends predomitly on genomic context rather
than the actual sequencing data which manifests in high recurrence of regions
that can/cannot be reliably genotyped by a single method. This enables the
application of pre-computed regions to other data created with comparable
sequencing technology and software. RG outperforms comparable efforts in
predicting VC concordance and false positive calls in low-concordance regions
which underlines its usefulness for variant filtering, annotation and
prioritization. RG allows focusing resource-intensive algorithms (e.g. consensus
calling methods) on the smaller, discordant share of the genome (20-30%) which
might result in increased overall accuracy at reasonable costs. Our method and
analysis of discordant calls may further be useful for development, benchmarking
and optimization of VC algorithms and for the relative comparison of call sets
between different studies/pipelines.
AVAILABILITY AND IMPLEMENTATION: RG was implemented in Java, source code and
binaries are freely available for non-commercial use at
https://github.com/popitsch/wtchg-rg/ CONTACT: [email protected]
information: Supplementary data are available at Bioinformatics online. |
List polyubiquitin binding proteins involved in NF-kappaB signaling. | NEMO
A20
ABIN-1
ABIN-2
optineurin
p62 | Attachment of ubiquitin to proteins represents a central mechanism for the
regulation of protein metabolism and function. In the NF-kappaB pathway, binding
of NEMO to polyubiquitinated substrates initiates the pathway in response to
cellular stimuli. Other polyubiquitin binding proteins can antagonize this
pathway by competing with NEMO for polyubiquitin. We have used protein arrays to
identify polyubiquitin binding proteins that regulate NF-kappaB activity. Using
polyubiquitin as bait, protein arrays were screened and polyubiquitin binders
identified. Novel polyubiquitin binders AWP1, CALCOCO2, N4BP1, RIO3, TEX27,
TTC3, UBFD1 and ZNF313 were identified using this approach, while known
NF-kappaB regulators including NEMO, A20, ABIN-1, ABIN-2, optineurin and p62
were also identified. Overexpressed AWP1 and RIO3 repressed NF-kappaB activity
in a manner similar to optineurin, while siRNAs directed against AWP1 and RIO3
also reduced NF-kappaB activity. TNFalpha-dependent degradation of IkappaBalpha
was also suppressed by overexpression of AWP1 and RIO3, possibly due to the
polyubiquitin binding activity of these proteins. Protein array screening using
polyubiquitin enabled rapid identification of many known and novel polyubiquitin
binding proteins and the identification of novel NF-kappaB regulators. Polyubiquitination of proteins plays a critical role in the activation of immune
cells. K63-linked polyubiquitin-binding proteins TGF-β-activated kinase 1
(TAK1)-binding protein (TAB)2 and TAB3 are implicated in NF-κB signaling via
TAK1 activation. However, TAB2 alone is dispensable for NF-κB activation in
embryonic fibroblasts, and the functional roles of TAB2 and TAB3 in immune cells
has yet to be clarified. In this study, we demonstrate that TAB2 and TAB3 are
essential for B cell activation leading to Ag-specific Ab responses, as well as
B-1 and marginal zone B cell development. TAB2 and TAB3 are critical for the
activation of MAPKs, especially ERK, but not NF-κB, in response to TLR and CD40
stimulation in B cells. Surprisingly, TAB2 and TAB3 are dispensable for TAK1
activation in B cells, indicating that TAB2 and TAB3 activate MAPKs via a
pathway independent of TAK1. In contrast to B cells, macrophages lacking TAB2
and TAB3 did not show any defects in the cytokine production and the signaling
pathway in response to TLR stimulation. Furthermore, TAB2 and TAB3 were
dispensable for TNF-induced cytokine production in embryonic fibroblasts. Thus,
TAB2- and TAB3-mediated K63-linked polyubiquitin recognition controls B cell
activation via MAPKs, but not the TAK1/NF-κB axis. Research over the past decade has revealed how NF-κB essential modulator (NEMO;
also known as IKKγ) regulates the IKKα-IKKβ signalling axis in the innate immune
system. The discovery that NEMO is a polyubiquitin-binding protein and that the
IKK complex is modulated by other protein kinases that are themselves controlled
by polyubiquitin chains has provided a deeper molecular understanding of the
non-degradative roles of ubiquitylation. New mechanistic insights of NEMO and
related polyubiquitin-binding proteins have become a paradigm for how the
interplay between phosphorylation and ubiquitylation controls cell signalling
networks in health and disease. |
What are the prednisone side effects in DMD patients? | Side effects of prednisone in DMD patients include reduced growth rate and increase in body weight. | |
Which is the specificity of deubiquitinase USP25? | Ubiquitin Specific Protease 25 (USP25), a member of the deubiquitinase family, is involved in several disease-related signal pathways including myogenesis, immunity and protein degradation. It specially catalyzes the hydrolysis of the K48-linked and K63-linked polyubiquitin chains. | Ubiquitin Specific Protease 25 (USP25), a member of the deubiquitinase family,
is involved in several disease-related signal pathways including myogenesis,
immunity and protein degradation. It specially catalyzes the hydrolysis of the
K48-linked and K63-linked polyubiquitin chains. USP25 contains one
ubiquitin-associated domain and two ubiquitin-interacting motifs (UIMs) in its
N-terminal region, which interact with ubiquitin and play a role in substrate
recognition. Besides, it has been shown that the catalysis activity of USP25 is
either impaired by sumoylation or enhanced by ubiquitination within its UIM. To
elucidate the structural basis of the cross-regulation of USP25 function by
non-covalent binding and covalent modifications of ubiquitin and SUMO2/3, a
systematic structural biology study of USP25 is required. Here, we report the
(1)H, (13)C and (15)N backbone and side-chain resoce assignments of the
N-terminal ubiquitin binding domains (UBDs) of USP25 with BMRB accession number
of 19111, which is the first step of the systematic structural biology study of
the enzyme. Ubiquitination and deubiquitination have emerged as critical regulatory
processes in the virus-triggered type I interferon (IFN) induction pathway. In
this study, we carried out a targeted siRNA screen of 54 ubiquitin-specific
proteases (USPs) and identified USP25 as a negative regulator of the
virus-triggered type I IFN signaling pathway. Overexpression of USP25 inhibited
virus-induced activation of IFN-β, interferon regulation factor 3 (IRF3) and
nuclear factor-kappa B (NF-κB), as well as the phosphorylation of IRF3 and NF-κB
subunit p65. Furthermore, Knockdown of USP25 potentiated virus-induced induction
of the IFN-β. In addition, detailed analysis demonstrated that USP25 cleaved
lysine 48- and lysine 63-linked polyubiquitin chains in vitro and in vivo, and
its deubiquitinating enzyme (DUB) activity, were dependent on a cysteine residue
(Cys178) and a histidine residue (His607). USP25 mutants lacking DUB activity
lost the ability to block virus-induced type I IFN to some degree.
Mechanistically, USP25 deubiquitinated retinoic acid-inducible gene I (RIG-I),
tumornecrosis factor (TNF) receptor-associated factor 2 (TRAF2), and TRAF6 to
inhibit RIG-I-like receptor-mediated IFN signaling. Our findings suggest that
USP25 is a novel DUB negatively regulating virus-induced type I IFN production. Ubiquitin-specific protease (USP) 25, belonging to the USP family of
deubiquitinases, harbors two tandem ubiquitin-interacting motifs (UIMs), a
~20-amino-acid α-helical stretch that binds to ubiquitin. However, the role of
the UIMs in USP25 remains unclear. Here we show that the tandem UIM region binds
to Lys48-, but not Lys63-, linked ubiquitin chains, where the two UIMs played a
critical and cooperative role. Purified USP25 exhibited higher ubiquitin
isopeptidase activity to Lys48-, than to Lys63-, linked ubiquitin chains.
Mutations that disrupted the ubiquitin-binding ability of the tandem UIMs
resulted in a reduced ubiquitin isopeptidase activity of USP25, suggesting a
role for the UIMs in exerting the full catalytic activity of USP25. Moreover,
when mutations that convert the binding preference from Lys48- to Lys63-linked
ubiquitin chains were introduced into the tandem UIM region, the USP25 mutants
acquired elevated and reduced isopeptidase activity toward Lys63- and
Lys48-linked ubiquitin chains, respectively. These results suggested that the
binding preference of the tandem UIMs toward Lys48-linked ubiquitin chains
contributes not only to the full catalytic activity but also to the ubiquitin
chain substrate preference of USP25, possibly by selectively holding the
Lys48-linked ubiquitin chain substrates in the proximity of the catalytic core. |
Is Loss of function one of the cardinal signs of inflammation? | Functio Laesa also known as loss of function is considered to be the 5th cardinal sign of inflammation. | The concept of the four cardinal signs of acute inflammation comes from
antiquity as rubor et tumor cum calore et dolore, (redness and swelling with
heat and pain) extended later by functio laesa (loss of function). The
contemporary understanding of this process we owe to 19th-century milestone
discoveries by Rudolph Virchow, Julius Cohnheim and Elie Metchnikoff. In the
20th century, the development of potent technological tools allowed the rapid
expansion of knowledge of the cells and mediators of inflammatory processes, as
well as the molecular mechanisms of their interactions. It turned out that some
mediators of inflammation have both local and distant targets, among them the
liver (responding by the production of several acute phase reactants) and
neurohormonal centers. In the last decades it has become clear that the immune
system shares mediators and their receptors with the neurohormonal system of the
body; thus, they form a common homeostatic entity. Such an integrative view,
introduced by J. Edwin Blalock, when combined with Hans Selye's concept of
stress, led to the contemporary understanding of sickness behavior, defined by
Robert Dantzer as a highly organized strategy of the organism to fight
infections and to respond to other environmental stressors. According to semiotics, which may be defined as the doctrine of the essential
nature and fundamental varieties of signs, objects, and interpretants, pain is
considered to be a sign (significant) with very different meanings
(significance) either as a naturalistic symptom (of disease) or as a symbol used
in a metaphorical context. When following this methodological perspective it is
possible to interpret medical as well as poetic writings on equal terms. In
Graeco-Roman medical texts pain was mostly understood as a result and an
indicator of disease, but nonetheless as a symptom which seemed to be actively
produced by the affected body. Especially in the Corpus Hippocraticum dating
from the 5th and 4th century B. C. this materialistic and at the same time
psychosomatic attitude can be noticed. Aristotle (4th century B. C.), Celsus
(1st century A. D.), and the famous experimental physiologist Galen (2nd century
A. D.) agreed that pain was a sign of evil which should be fought without
exception. It was Galen who added the disturbance of function (functio laesa) as
the fifth cardinal sign of inflammation to the four well-known cardinal signs of
Celsus (rubor, calor, tumor, dolor). He also coined the term [see text] to
characterize an attack of migraine. In algotherapy, Galen used a complex
pharmacological system which was based upon the four cardinal qualities of
humoral pathology. On the other hand, pain was designed as a multi-dimensional
symbol by the famous Graeco-Roman epic poets. In Homer's Odyssey (8th century B.
C.), pain appears transformed into the shape of a scar which is visible and
palpable on the hero's leg like an identification tag, whereas in Virgil's
Aeneids (1st century B. C.) pain symbolizes weakness and defencelessness which
can only be alleviated by the goddess Venus. Inflammation is the body's response to injury or infection. As early as 2000
years ago, the Roman encyclopaedist Aulus Cornelius Celsus recognised four
cardinal signs of this response-redness, heat, swelling and pain; a fifth sign
is loss of function.[...]. |
Does the association of PARP1 and CTCF follow a circadian rhythm? | Synchronization of the circadian rhythm by serum shock induces oscillations in PARP1-CTCF interactions, which is accompanied by oscillating recruitment of circadian loci to the lamina, followed by the acquisition of repressive H3K9me2 marks and transcriptional attenuation. Furthermore, depletion of H3K9me2/3, inhibition of PARP activity by olaparib, or downregulation of PARP1 or CTCF expression counteracts both recruitment to the envelope and circadian transcription. PARP1- and CTCF-regulated contacts between circadian loci and the repressive chromatin environment at the lamina therefore mediate circadian transcriptional plasticity. | |
What induces Arabidopsis ROF1 expression? | The abundance of ROF1 increased several-fold under stress conditions such as wounding, heat stress or exposure to elevated NaCl levels. | We have isolated clones of an Arabidopsis gene (ROF1, for rotamase FKBP)
encoding a high molecular weight member of the FK506 binding protein (FKBP)
family. The deduced amino acid sequence of ROF1 predicts a 551-amino acid, 62
kDa polypeptide which is 44% identical to human FKBP59 - a 59 kDa FKBP which
binds to the 90 kDa heat shock protein and is associated with inactive steroid
hormone receptors. ROF1 contains three FKBP12-like domains in the N-terminal
portion of the protein (in contrast to two domains in mammalian FKBP59), an
internal repeat structure associated with protein-protein interactions
(tetratricopeptide repeats), and a putative calmodulin binding domain near the
C-terminal region of the protein. No sequences associated with protein
translocation out of the cytosol were found in ROF1. ROF1 mRNA was found at
equivalent low levels in light-grown roots, stems, and flowers and at slightly
higher levels in leaves. The abundance of ROF1 mRNA increased several-fold under
stress conditions such as wounding or exposure to elevated NaCl levels. The provece, half-life and biological activity of malondialdehyde (MDA) were
investigated in Arabidopsis thaliana. We provide genetic confirmation of the
hypothesis that MDA originates from fatty acids containing more than two
methylene-linked double bonds, showing that tri-unsaturated fatty acids are the
in vivo source of up to 75% of MDA. The abundance of the combined pool of free
and reversibly bound MDA did not change dramatically in stress, although a
significant increase in the free MDA pool under oxidative conditions was
observed. The half-life of infiltrated MDA indicated rapid metabolic
turnover/sequestration. Exposure of plants to low levels of MDA using a recently
developed protocol powerfully upregulated many genes on a cDNA microarray with a
bias towards those implicated in abiotic/environmental stress (e.g. ROF1 and
XERO2). Remarkably, and in contrast to the activities of other reactive
electrophile species (i.e. small vinyl ketones), none of the
pathogenesis-related (PR) genes tested responded to MDA. The use of structural
mimics of MDA isomers suggested that the propensity of the molecule to act as a
cross-linking/modifying reagent might contribute to the activation of gene
expression. Changes in the concentration/localisation of unbound MDA in vivo
could strongly affect stress-related transcription. The plant co-chaperones FK506-binding proteins (FKBPs) are peptidyl prolyl
cis-trans isomerases that function in protein folding, signal transduction and
chaperone activity. We report the characterization of the Arabidopsis large
FKBPs ROF1 (AtFKBP62) and ROF2 (AtFKBP65) expression and protein accumulation
patterns. Transgenic plants expressing ROF1 promoter fused to GUS reporter gene
reveal that ROF1 expression is organ specific. High expression was observed in
the vascular elements of roots, in hydathodes and trichomes of leaves and in
stigma, sepals, and anthers. The tissue specificity and temporal expression of
ROF1 and ROF2 show that they are developmentally regulated. Although ROF1 and
ROF2 share 85% identity, their expression in response to heat stress is
differentially regulated. Both genes are induced in plants exposed to 37 degrees
C, but only ROF2 is a bonafide heat-stress protein, undetected when plants are
grown at 22 degrees C. ROF1/ROF2 proteins accumulate at 37 degrees C, remain
stable for at least 4 h upon recovery at 22 degrees C, whereas, their mRNA level
is reduced after 1 h at 22 degrees C. By protein interaction assays, it was
demonstrated, that ROF1 is a novel partner of HSP90. The five amino acids
identified as essential for recognition and interaction between the mammalian
chaperones and HSP90 are conserved in the plant ROF1-HSP90. We suggest that
ROF/HSP90 complexes assemble in vivo. We propose that specific complexes
formation between an HSP90 and ROF isoforms depends on their spatial and
temporal expression. Such complexes might be regulated by environmental
conditions such as heat stress or internal cues such as different hormones. A direct interaction of the Arabidopsis thaliana immunophilin ROF1 with
phosphatidylinositol-3-phosphate and phosphatidylinositol-3,5-bisphosphate was
identified using a phosphatidylinositol-phosphate affinity chromatography of
cell suspension extracts, combined with a mass spectrometry (o LC ESI-MS/MS)
analysis. The first FK506 binding domain was shown sufficient to bind to both
phosphatidylinositol-phosphate stereoisomers. GFP-tagged ROF1 under the control
of a 35S promoter was localised in the cytoplasm and the cell periphery of
Nicotiana tabacum leaf explants. Immunofluorescence microscopy of Arabidopsis
thaliana root tips verified its cytoplasmic localization and membrane
association and showed ROF1 localization in the elongation zone which was
expanded to the meristematic zone in plants grown on high salt media. Endogenous
ROF1 was shown to accumulate in response to high salt treatment in Arabidopsis
thaliana young leaves as well as in seedlings germinated on high salt media
(0.15 and 0.2 M NaCl) at both an mRNA and protein level. Plants over-expressing
ROF1, (WSROF1OE), exhibited enhanced germination under salinity stress which was
significantly reduced in the rof1(-) knock out mutants and abolished in the
double mutants of ROF1 and of its interacting homologue ROF2 (WSrof1(-)/2(-)).
Our results show that ROF1 plays an important role in the osmotic/salt stress
responses of germinating Arabidopsis thaliana seedlings and suggest its
involvement in salinity stress responses through a
phosphatidylinositol-phosphate related protein quality control pathway. |
What is the preferred orientation of CTCF binding sites for chromatin looping? | chromatin loops preferentially form between CTCF binding sites oriented in a convergent manner. CTCF sites at loop anchors occur predominantly (>90%) in a convergent orientation, with the asymmetric motifs "facing" one another. | Author information:
(1)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA; Department of Computer Science, Department of
Computational and Applied Mathematics, Rice University, Houston, TX 77005, USA;
Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA.
(2)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA; Department of Computer Science, Department of
Computational and Applied Mathematics, Rice University, Houston, TX 77005, USA;
Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA; School of
Engineering and Applied Sciences, Harvard University, Cambridge, MA 02138, USA.
(3)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA; Department of Computer Science, Department of
Computational and Applied Mathematics, Rice University, Houston, TX 77005, USA.
(4)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA.
(5)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA; Department of Computer Science, Department of
Computational and Applied Mathematics, Rice University, Houston, TX 77005, USA;
Department of Computer Science, Stanford University, Stanford, CA 94305, USA.
(6)Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA; Department of
Biology, Massachusetts Institute of Technology (MIT), Cambridge, MA 02139, USA;
Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
Electronic address: [email protected].
(7)The Center for Genome Architecture, Baylor College of Medicine, Houston, TX
77030, USA; Department of Molecular and Human Genetics, Baylor College of
Medicine, Houston, TX 77030, USA; Department of Computer Science, Department of
Computational and Applied Mathematics, Rice University, Houston, TX 77005, USA;
Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA; Center for
Theoretical Biological Physics, Rice University, Houston, TX 77030, USA.
Electronic address: [email protected]. Author information:
(1)Center for Comparative Biomedicine, MOE Key Laboratory of Systems
Biomedicine, Institute of Systems Biomedicine, Collaborative Innovation Center
of Systems Biomedicine, Shanghai Jiao Tong University (SJTU), Shanghai 200240,
China; State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer
Institute, Renji Hospital, School of Medicine, SJTU, Shanghai 200240, China; Key
Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders
(MOE), Bio-X Center, School of Life Sciences and Biotechnology, SJTU, Shanghai
200240, China.
(2)Department of Biochemistry and Molecular Biophysics, Columbia University
Medical Center, 701 West 168(th) Street, New York, NY 10032, USA.
(3)Ludwig Institute for Cancer Research and Department of Cellular and Molecular
Medicine, University of California, San Diego School of Medicine, 9500 Gilman
Drive, La Jolla, CA 92093, USA.
(4)Department of Molecular and Cell Biology, Center for Systems Biology,
University of Texas at Dallas, Richardson, TX 75080, USA; MOE Key Laboratory of
Bioinformatics and Bioinformatics Division, Center for Synthetic and System
Biology, TNLIST/Department of Automation, Tsinghua University, Beijing 100084,
China.
(5)Cold Spring Harbor Laboratory, 1 Bungtown Rd, NY 11724, USA.
(6)Department of Biochemistry and Molecular Biophysics, Columbia University
Medical Center, 701 West 168(th) Street, New York, NY 10032, USA. Electronic
address: [email protected].
(7)Center for Comparative Biomedicine, MOE Key Laboratory of Systems
Biomedicine, Institute of Systems Biomedicine, Collaborative Innovation Center
of Systems Biomedicine, Shanghai Jiao Tong University (SJTU), Shanghai 200240,
China; State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer
Institute, Renji Hospital, School of Medicine, SJTU, Shanghai 200240, China; Key
Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders
(MOE), Bio-X Center, School of Life Sciences and Biotechnology, SJTU, Shanghai
200240, China. Electronic address: [email protected]. We recently used in situ Hi-C to create kilobase-resolution 3D maps of mammalian
genomes. Here, we combine these maps with new Hi-C, microscopy, and
genome-editing experiments to study the physical structure of chromatin fibers,
domains, and loops. We find that the observed contact domains are inconsistent
with the equilibrium state for an ordinary condensed polymer. Combining Hi-C
data and novel mathematical theorems, we show that contact domains are also not
consistent with a fractal globule. Instead, we use physical simulations to study
two models of genome folding. In one, intermonomer attraction during polymer
condensation leads to formation of an anisotropic "tension globule." In the
other, CCCTC-binding factor (CTCF) and cohesin act together to extrude unknotted
loops during interphase. Both models are consistent with the observed contact
domains and with the observation that contact domains tend to form inside loops.
However, the extrusion model explains a far wider array of observations, such as
why loops tend not to overlap and why the CTCF-binding motifs at pairs of loop
anchors lie in the convergent orientation. Finally, we perform 13 genome-editing
experiments examining the effect of altering CTCF-binding sites on chromatin
folding. The convergent rule correctly predicts the affected loops in every
case. Moreover, the extrusion model accurately predicts in silico the 3D maps
resulting from each experiment using only the location of CTCF-binding sites in
the WT. Thus, we show that it is possible to disrupt, restore, and move loops
and domains using targeted mutations as small as a single base pair. CCCTC-binding factor (CTCF) is an architectural protein involved in the
three-dimensional (3D) organization of chromatin. In this study, we assayed the
3D genomic contact profiles of a large number of CTCF binding sites with
high-resolution 4C-seq. As recently reported, our data also suggest that
chromatin loops preferentially form between CTCF binding sites oriented in a
convergent manner. To directly test this, we used CRISPR/Cas9 genome editing to
delete core CTCF binding sites in three loci, including the CTCF site in the
Sox2 super-enhancer. In all instances, CTCF and cohesin recruitment were lost,
and chromatin loops with distal, convergent CTCF sites were disrupted or
destabilized. Re-insertion of oppositely oriented CTCF recognition sequences
restored CTCF and cohesin recruitment, but did not re-establish chromatin loops.
We conclude that CTCF binding polarity plays a functional role in the formation
of higher-order chromatin structure. Mammalian interphase chromosomes fold into a multitude of loops to fit the
confines of cell nuclei, and looping is tightly linked to regulated function.
Chromosome conformation capture (3C) technology has significantly advanced our
understanding of this structure-to-function relationship. However, all 3C-based
methods rely on chemical cross-linking to stabilize spatial interactions. This
step remains a "black box" as regards the biases it may introduce, and some
discrepancies between microscopy and 3C studies have now been reported. To
address these concerns, we developed "i3C", a novel approach for capturing
spatial interactions without a need for cross-linking. We apply i3C to intact
nuclei of living cells and exploit native forces that stabilize chromatin
folding. Using different cell types and loci, computational modeling, and a
methylation-based orthogonal validation method, "TALE-iD", we show that native
interactions resemble cross-linked ones, but display improved signal-to-noise
ratios and are more focal on regulatory elements and CTCF sites, while strictly
abiding to topologically associating domain restrictions. |
Does erythromycin increase risk of hypertrophic pyloric stenosis? | Yes, post-natal erythromycin exposure is associated with increased risk of hypertrophic pyloric stenosis. The association is stronger when exposure occurs in the first 2 weeks of life. | OBJECTIVES: To evaluate the risk for infantile hypertrophic pyloric stenosis
(IHPS) among infants prescribed systemic erythromycin, infants prescribed a
course of erythromycin ophthalmic ointment, and infants whose mothers were
prescribed a macrolide antibiotic during pregcy.
STUDY DESIGN: Retrospective cohort study of infants born at an urban hospital
from June 1993 through December 1999.
RESULTS: Of 14,876 eligible infants, 43 (0.29%) developed IHPS. Infants
prescribed systemic erythromycin had increased risk of IHPS, with the highest
risk in the first 2 weeks of age (relative risk = 10.51 for erythromycin in
first 2 weeks, 95% CI 4.48, 24.66). Erythromycin ophthalmic ointment for
conjunctivitis was not associated with increased risk of IHPS. Maternal
macrolide antibiotics within 10 weeks of delivery may have been associated with
higher risk of IHPS but the data were not conclusive.
CONCLUSIONS: This study confirms an association between systemic erythromycin in
infants and subsequent IHPS, with the highest risk in the first 2 weeks of age.
No association was found with erythromycin ophthalmic ointment. A possible
association with maternal macrolide therapy in late pregcy requires further
study. Systemic erythromycin should be used with prudence in early infancy. OBJECTIVE: To assess the association between prenatal antibiotics, including
erythromycin, and infantile hypertrophic pyloric stenosis in a large cohort of
infants.
METHODS: This was a retrospective cohort study of births to women enrolled in
Tennessee Medicaid/TennCare, 1985-1997. Prescriptions for erythromycin,
nonerythromycin macrolides, and other antibiotics were identified from pharmacy
files linked with birth certificate files. The primary study outcome was
development of pyloric stenosis in the infant, identified from linked hospital
discharge diagnosis and surgical procedure codes.
RESULTS: The cohort included 260,799 mother/infant pairs. Among these women,
13,146 filled prescriptions for erythromycin (50.4 per 1000), and 621 filled
prescriptions for nonerythromycin macrolides (2.4 per 1000). There was no
association with prenatal erythromycin prescription and infantile hypertrophic
pyloric stenosis either after 32 weeks' gestation (adjusted odds ratio 1.17, 95%
confidence interval, 0.84, 1.64, P =.33) or at any time during pregcy
(adjusted odds ratio 1.15, 95% confidence interval 0.84, 1.56, P =.36). There
was an association between maternal prescriptions for nonerythromycin macrolides
and infantile hypertrophic pyloric stenosis (adjusted odds ratio 2.77, 95%
confidence interval 1.22, 6.30, P =.01).
CONCLUSION: The hypothesized association between erythromycin and infantile
pyloric stenosis was not seen. Causal inference from the association between
prenatal nonerythromycin macrolides and infantile hypertrophic pyloric stenosis
is limited by the small number of affected children and the evidence of other
differences between users of nonerythromycin macrolides and controls. |
How many amino acids does davunetide consist of? | Davunetide or NAP is an eight amino acid peptide. | Genome-wide association studies have identified strong associations between the
risk of developing Parkinson's disease (PD) and polymorphisms in the genes
encoding α-synuclein and the microtubule-associated protein tau. However, the
contribution of tau and its phosphorylated form (p-tau) to α-synuclein-induced
pathology and neuronal dysfunction remains controversial. We have assessed the
effects of NAP (davunetide), an eight-amino acid peptide that decreases tau
hyperphosphorylation, in mice overexpressing wild-type human α-synuclein
(Thy1-aSyn mice), a model that recapitulates aspects of PD. We found that the
p-tau/tau level increased in a subcortical tissue block that includes the
striatum and brain stem, and in the cerebellum of the Thy1-aSyn mice compared to
nontransgenic controls. Intermittent intranasal NAP administration at 2 μg/mouse
per day, 5 days a week, for 24 weeks, starting at 4 weeks of age, significantly
decreased the ratio of p-tau/tau levels in the subcortical region while a higher
dose of 15 μg/mouse per day induced a decrease in p-tau/tau levels in the
cerebellum. Both NAP doses reduced hyperactivity, improved habituation to a
novel environment, and reduced olfactory deficits in the Thy1-aSyn mice, but
neither dose improved the severe deficits of motor coordination observed on the
challenging beam and pole, contrasting with previous data obtained with
continuous daily administration of the drug. The data reveal novel effects of
NAP on brain p-tau/tau and behavioral outcomes in this model of synucleinopathy
and suggest that sustained exposure to NAP may be necessary for maximal
benefits. |
Can a circRNA be translated into protein? | Circ-ZNF609 is associated with heavy polysomes, and it is translated into a protein in a splicing-dependent and cap-independent manner, providing an example of a protein-coding circRNA in eukaryotes.
Circular RNAs (circRNAs) represent a large class of noncoding RNAs (ncRNAs) that have recently emerged as regulators of gene expression
However, whether circRNAs encode functional proteins remains elusive, although translation of several circRNAs was recently reported | Circular RNAs (circRNAs) are long, non-coding RNAs that result from the
non-canonical splicing of linear pre-mRNAs. However, the characteristics and the
critical role of circRNA in co-/post-transcriptional regulation were not well
recognized until the "microRNA sponge" function of circRNA is discovered. Recent
studies have mainly been devoted to the function of the circular RNA sponge for
miR-7 (ciRS-7) and sex-determining region Y (SRY) by targeting microRNA-7
(miR-7) and microRNA-138 (miR-138), respectively. In this review, we illustrate
the specific role of circRNAs in a wide variety of cancers and in regulating the
biological behavior of cancers via miR-7 or miR-138 regulation. Furthermore,
circRNA, together with its gene silencing ability, also shows its potential in
RNA interference (RNAi) therapy by binding to target RNAs, which provides a
novel perspective in cancer treatment. Thus, this review concerns the
biogenesis, biological function, oncogenesis, progression and possible therapies
for cancer involving circRNAs. Circular RNAs (circRNAs) are a large type of noncoding RNAs characterized by
their circular shape resulting from covalently closed continuous loops. They are
known to regulate gene expression in mammals. These tissue-specific transcripts
are largely generated from exonic or intronic sequences of their host genes.
Although several models of circRNA biogenesis have been proposed, the
understanding of their origin is far from complete. Unlike other noncoding RNAs,
circRNAs are widely expressed, highly conserved and stable in cytoplasm, which
confer special functionalities to them. They are known to serve as microRNA
(miRNA) sponges, regulators of alternative splicing, transcription factors and
encode for proteins. The expression of circRNAs is associated with several
pathological states and may potentially serve as novel diagnostic or predictive
biomarkers. CircRNAs are known to regulate the expression of numerous
cancer-related miRNAs. The circRNA-miRNA-mRNA axis is a known regulatory pattern
of several cancer-associated pathways, with both agonist and antagonist effects
on carcinogenesis. In consideration of their potential clinical relevance,
circRNAs are at the center of ongoing research initiatives on cancer prevention
and treatment. In this review, we discuss the current understanding of circRNAs
and the prospects for their potential clinical application in the management of
cancer patients. Circular RNAs (circRNAs) represent a large class of noncoding RNAs (ncRNAs) that
have recently emerged as regulators of gene expression. They have been shown to
suppress microRNAs, thereby increasing the translation and stability of the
targets of such microRNAs. In this review, we discuss the emerging functions of
circRNAs, including RNA transcription, splicing, turnover, and translation. We
also discuss other possible facets of circRNAs that can influence their function
depending on the cell context, such as circRNA abundance, subcellular
localization, interacting partners (RNA, DNA, and proteins), dynamic changes in
interactions following stimulation, and potential circRNA translation. The
ensuing changes in gene expression patterns elicited by circRNAs are proposed to
drive key cellular processes, such as cell proliferation, differentiation, and
survival, that govern health and disease. WIREs RNA 2017, 8:e1386. doi:
10.1002/wrna.1386 For further resources related to this article, please visit
the WIREs website. Circular RNAs (circRNAs) are abundant and evolutionarily conserved RNAs of
largely unknown function. Here, we show that a subset of circRNAs is translated
in vivo. By performing ribosome footprinting from fly heads, we demonstrate that
a group of circRNAs is associated with translating ribosomes. Many of these
ribo-circRNAs use the start codon of the hosting mRNA, are bound by
membrane-associated ribosomes, and have evolutionarily conserved termination
codons. In addition, we found that a circRNA generated from the muscleblind
locus encodes a protein, which we detected in fly head extracts by mass
spectrometry. Next, by performing in vivo and in vitro translation assays, we
show that UTRs of ribo-circRNAs (cUTRs) allow cap-independent translation.
Moreover, we found that starvation and FOXO likely regulate the translation of a
circMbl isoform. Altogether, our study provides strong evidence for translation
of circRNAs, revealing the existence of an unexplored layer of gene activity. Circular RNAs (circRNAs) constitute a family of transcripts with unique
structures and still largely unknown functions. Their biogenesis, which proceeds
via a back-splicing reaction, is fairly well characterized, whereas their role
in the modulation of physiologically relevant processes is still unclear. Here
we performed expression profiling of circRNAs during in vitro differentiation of
murine and human myoblasts, and we identified conserved species regulated in
myogenesis and altered in Duchenne muscular dystrophy. A high-content functional
genomic screen allowed the study of their functional role in muscle
differentiation. One of them, circ-ZNF609, resulted in specifically controlling
myoblast proliferation. Circ-ZNF609 contains an open reading frame spanning from
the start codon, in common with the linear transcript, and terminating at an
in-frame STOP codon, created upon circularization. Circ-ZNF609 is associated
with heavy polysomes, and it is translated into a protein in a
splicing-dependent and cap-independent manner, providing an example of a
protein-coding circRNA in eukaryotes. BACKGROUND: Circular RNAs (circRNAs) are RNA transcripts that are widespread in
the eukaryotic genome. Recent evidence indicates that circRNAs play important
roles in tissue development, gene regulation, and carcinogenesis. However,
whether circRNAs encode functional proteins remains elusive, although
translation of several circRNAs was recently reported.
METHODS: CircRNA deep sequencing was performed by using 10 pathologically
diagnosed glioblastoma samples and their paired adjacent normal brain tissues.
Northern blotting, Sanger sequencing, antibody, and liquid chromatograph Tandem
Mass Spectrometer were used to confirm the existence of circ-FBXW7 and its
encoded protein in in two cell lines. Lentivirus-transfected stable U251 and
U373 cells were used to assess the biological functions of the novel protein
invitro and invivo (five mice per group). Clinical implications of circ-FBXW7
were assessed in 38 pathologically diagnosed glioblastoma samples and their
paired periphery normal brain tissues by using quantitative polymerase chain
reaction (two-sided log-rank test).
RESULTS: Circ-FBXW7 is abundantly expressed in the normal human brain (reads per
kilobase per million mapped reads [RPKM] = 9.31). The spanning junction open
reading frame in circ-FBXW7 driven by internal ribosome entry site encodes a
novel 21-kDa protein, which we termed FBXW7-185aa. Upregulation of FBXW7-185aa
in cancer cells inhibited proliferation and cell cycle acceleration, while
knockdown of FBXW7-185aa promoted maligt phenotypes invitro and invivo.
FBXW7-185aa reduced the half-life of c-Myc by antagonizing USP28-induced c-Myc
stabilization. Moreover, circ-FBXW7 and FBXW7-185aa levels were reduced in
glioblastoma clinical samples compared with their paired tumor-adjacent tissues
(P < .001). Circ-FBXW7 expression positively associated with glioblastoma
patient overall survival (P = .03).
CONCLUSIONS: Endogenous circRNA encodes a functional protein in human cells, and
circ-FBXW7 and FBXW7-185aa have potential prognostic implications in brain
cancer. SUMMARY: Circular RNAs (circRNAs), a novel class of endogenous RNAs, are
widespread in eukaryotic cells. Emerging roles in diverse biological processes
suggest that circRNA is a promising key player in RNA world. Most circRNAs are
generated through back-splicing of pre-mRNAs, forming a covalently closed loop
structure with no 5' caps or 3' polyadenylated tails. In addition, most circRNAs
were not associated with translating ribosomes, therefore, circRNAs were deemed
to be noncoding. However, the latest research findings revealed that some
circRNAs could generate proteins in vivo, which expands the landscape of
transcriptome and proteome. To gain insights into the new area of circRNA
translation, we introduce an integrated tool capable of detecting circRNAs with
protein-coding potential from high-throughput sequencing data.
AVAILABILITY AND IMPLEMENTATION: CircPro is available at
http://bis.zju.edu.cn/CircPro.
CONTACT: [email protected].
SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics
online. |
What is the phenomenon of gene kissing? | Clustering of genes with similar expression patterns constitutes a phenomenon sometimes called "gene kissing." | At certain evolutionary junctures, two or more mutations participating in the
build-up of a new complex function may be required to become available
simultaneously in the same individuals. How could this happen in higher
organisms whose populations are small compared to those of microbes, and in
which chances of combined nearly simultaneous highly specific favorable
mutations are correspondingly low? The question can in principle be answered for
regulatory evolution, one of the basic processes of evolutionary change. A
combined resetting of transcription rates in several genes could occur in the
same individual. It is proposed that, in eukaryotes, changes in epigenetic
trends and epigenetically transforming encounters between alternative chromatin
structures could arise frequently enough so as to render probable particular
conjunctions of changed transcription rates. Such conjunctions could involve
mutational changes with low specificity requirements in gene-associated regions
of non-protein-coding sequences. The effects of such mutations, notably when
they determine the use of histone variants and covalent modifications of
histones, can be among those that migrate along chromatin. Changes in chromatin
structure are often cellularly inheritable over at least a limited number of
generations of cells, and of individuals when the germ line is involved. SINEs
and LINEs, which have been considered "junk DNA", are among the repeat sequences
that would appear liable to have teleregulatory effects on the function of a
nearby promoter, through changes in their numbers and distribution. There may
also be present preexisting unstably inheritable epigenetic trends leading to
cellular variegation, trends endemic in a cell population based on DNA sequences
previously established in the neighborhood. Either way, epigenetically
conditioned teleregulatory trends may display only limited penetrance. The
imposition at a distance of new chromatin structures with regulatory impact can
occur in cis as well as in trans, and is examined as intrachromosomally
spreading teleregulation and interchromosomal "gene kissing". The chances for
two or more particular epigenetically determined regulatory trends to occur
together in a cell are increased thanks to the proposed low specificity
requirements for most of the pertinent sequence changes in intergenic and
intronic DNA or in the distribution of middle repetitive sequences that have
teleregulatory impact. Inheritable epigenetic changes ("epimutations") with
effects at a distance would then perdure over the number of generations required
for "assimilation" of the several regulatory novelties through the occurrence
and selection, gene by gene, of specific classical mutations. These mutations
would have effects similar to the epigenetic effects, yet would provide
stability and penetrance. The described epigenetic/genetic partnership may well
at times have opened the way toward certain complex new functions. Thus, the
presence of "junk DNA", through co-determining the (higher or lower) order and
the variants of chromatin structure with regulatory effects at a distance, might
make an important contribution to the evolution of complex organisms. Expression of Hox genes located on different chromosomes is precisely regulated
and synchronized during development. In order to test the hypothesis that the
Hox loci might cluster in nuclear space in order to share regulatory components,
we performed 3D FISH on cryosections of developing mouse embryos and
differentiating embryoid bodies. We did not observe any instances of
co-localization of 4 different Hox alleles. Instances of 2 different alleles
touching each other were found in 20-47% of nuclei depending on the tissue. The
frequency of such "kissing" events was not significantly different in cells
expressing a high proportion of the Hox clusters when compared to cells
expressing none or only a few Hox genes. We found that the HoxB and HoxC
clusters, which are located in gene-rich regions, were involved more frequently
in gene kissing in embryonic nuclei. In the case of HoxB, this observation
correlated with the positioning of the corresponding chromosome towards the
interior of the nucleus. Our results indicate that co-regulation of the
different Hox clusters is not associated with co-localization of the loci at a
single regulatory compartment and that the chromosomal context may influence the
extent to which they contact each other in the nucleus. Genomes are spatially organized on many levels and the positioning of genes
within the nucleus contributes to their proper expression. This positioning can
also result in the clustering of genes with similar expression patterns, a
phenomenon sometimes called "gene kissing." We have found that yeast genes are
targeted to the nuclear periphery through interaction of the nuclear pore
complex with small, cis-acting "DNA zip codes" in their promoters. Our recent
study demonstrated that genes with the same zip codes cluster together at the
nuclear periphery. The zip codes were necessary and sufficient to induce
interchromosomal clustering. Finally, we identified a transcription factor
(Put3) that binds to the GRS I zip code. Put3 binds to GRS I and is required for
both GRS I-dependent positioning at the nuclear periphery and interchromosomal
clustering of GRS I-targeted genes. We speculate that our findings might provide
insight into other types of gene kissing, some of which also require cis-acting
DNA sequences and trans-acting proteins. |
What protein is the most common cause of hereditary renal amyloidosis? | The most common cause of hereditary renal amyloidosis is over expression of a mutant form of the Fibrinogen A Alpha protein | A French kindred with autosomal domit hereditary renal amyloidosis was found
to have a novel mutation in the fibrinogen Aalpha-chain gene. In this kindred,
renal disease appeared early in life and led to terminal renal failure at an
early age. Renal transplantation resulted in rapid destruction of the allograft
by amyloid deposition within 2 years. Amyloid fibril protein isolated from a
transplanted kidney was found to contain a novel, hybrid peptide of 49 residues
whose N-terminal 23 amino acids were identical to residues 499 to 521 of normal
fibrinogen Aalpha-chain. The remainder of the peptide (26 residues) represented
a completely new sequence for mammalian proteins. DNA sequencing documented that
the new sequence was the result of a single nucleotide deletion at position 4897
of the fibrinogen Aalpha-chain gene that gives a frame-shift at codon 522 and
premature termination at codon 548. The contributions toward fibrillogenesis of
the two portions of the amyloid fibril protein, ie, N-terminal fibrinogen
sequence and C-terminal novel sequence, are presently unknown. However, the
early onset and rapid reoccurrence of amyloid in renal transplants is unlike the
clinical course with other amyloid proteins having single amino acid
substitutions that give hereditary renal amyloidosis. Liver transplantation to
stop synthesis of this abnormal hepatic derived protein should be considered
early in the course of the disease. A middle age Portuguese woman was investigated for renal amyloidosis. She
presented with progressive renal failure, proteinuria, hypertension, and sensory
symptoms in the feet. Clinical and neurophysiological evaluation disclosed
sensory-autonomic neuropathy. Cardiovascular tests and 123-MIBG investigation
showed parasympathetic dysfunction and decrease of myocardial innervation, in
accordance with small fiber neuropathy, as usually observed in amyloidosis.
Immunohistochemical studies revealed AFib amyloidosis and genetic studies the
amino acid exchange Glu526Val of the fibrinogen Aalpha-chain mutation, which was
also present in one of her sons. The mutant gene in this patient was associated
with the same haplotype as all other reported cases of Glu526Val mutations. This
is the first reported AFibamyloidosis in Portugal, and the first case of AFib in
which sensory and autonomic nerve fiber dysfunction is described, indicating
that small nerve fiber lesion can occur in the fibrinogen Aalpha chain mutation.
This can be important for prognosis, in particular when liver transplantation is
considered for treatment. The predomit cause of hereditary renal amyloidosis is a mutation of the
fibrinogen Aalpha chain (AFib), the most common being the E526V mutation. The
evolution towards terminal renal insufficiency is constant and raises the
question of renal transplantation and the risk of recurrence. We describe the
case of a Portuguese woman with the E526V mutation without any renal or hepatic
history in her family which developed a nephrotic syndrome at the age of 35,
followed by stage 5 renal insufficiency. Because of the risk of recurrence of
amyloidosis on its transplant, we carried out a combined transplantation
liver-kidney despite the absence of clinical or biological hepatic
abnormalities. Four years later, the result is excellent and there is no sign of
the disease on the new organs. This successful experience as well as the five
other published cases of combined liver-kidney transplantation in Aalpha
fibrinogen chain amyloidosis, demonstrates the feasibility and efficacy of this
treatment in AFib amyloidosis. Mutations in the fibrinogen A alpha-chain gene are the most common cause of
hereditary renal amyloidosis in the United Kingdom. Previous reports of
fibrinogen A alpha-chain amyloidosis have been in isolated kindreds, usually in
the context of a novel amyloidogenic mutation. Here, we describe 71 patients
with fibrinogen amyloidosis, who were prospectively studied at the UK National
Amyloidosis Centre. Median age at presentation was 58 yr, and renal involvement
led to diagnosis in all cases. Even after a median follow-up of 4 yr, clinically
significant extra-renal disease was rare. Renal histology was characteristic:
striking glomerular enlargement with almost complete obliteration of the normal
architecture by amyloid deposition and little or no vascular or interstitial
amyloid. We discovered four amyloidogenic mutations in fibrinogen (P552H, E540V,
T538K, and T525fs). A family history of renal disease was frequently absent.
Median time from presentation to ESRD was 4.6 yr, and the estimated median
patient survival from presentation was 15.2 yr. Among 44 patients who reached
ESRD, median survival was 9.3 yr. Twelve renal transplants survived for a median
of 6.0 (0-12.2) yr. Seven grafts had failed after median follow up from
transplantation of 5.8 yr, including three from recurrent amyloid after 5.8,
6.0, and 7.4 yr; three grafts failed immediately for surgical reasons and one
failed from transplant glomerulopathy after 5.8 yr with no histological evidence
of amyloid. At censor, the longest surviving graft was 12.2 yr. In summary,
fibrinogen amyloidosis is predomitly a renal disease characterized by
variable penetrance, distinctive histological appearance, proteinuria, and
progressive renal impairment. Survival is markedly better than observed with
systemic AL amyloidosis, and outcomes with renal replacement therapy are
comparable to those for age-matched individuals with nondiabetic renal disease. BACKGROUND: Hereditary amyloidosis with predomit renal disease can be caused
by mutations in the gene encoding the fibrinogen Aα-chain (AFib). Here, we
describe the clinical course of AFib amyloidosis associated with the rare R554L
mutation, and the significance of extrarenal amyloid deposits and their possible
influence on cardiovascular morbidity.
METHODS: We report on 101 members of a family after having conducted patient
interviews, chart review, genetic testing, renal biopsies and assessment for
extrarenal amyloid deposition.
RESULTS: Ten family members had chronic kidney disease with late-onset gross
proteinuria and a variable course of declining renal function, starting in the
fourth decade of life. In two affected living members, we identified the AFib
R554L mutation. Renal biopsies from two affected members revealed almost
complete obliteration of the mesangial glomerular architecture, although kidney
function was only moderately impaired. There was neither evidence of extrarenal
amyloidosis nor accelerated atherosclerosis.
CONCLUSIONS: Renal amyloidosis associated with the R554L AFib variant dominated
the clinical picture in this family, which was similar to that associated with
the much more prevalent E526V mutation. Although it has been hypothesized that
vascular deposits of fibrinogen amyloid may be associated with accelerated
atherosclerosis, there was no suggestion of this in this particular kindred. Mutations in the fibrinogen Aα-chain genes are the most common cause of
hereditary renal amyloidosis. The renal histologic appearance in the patient is
characteristic and shows striking glomerular enlargement with almost complete
obliteration of the normal glomerular architecture by extensive amyloid
deposition. In contrast, the vessels and renal tubular interstitium of such
patient contains almost no amyloid at all. Here, we described a patient with
hereditary fibrinogen amyloidosis, who presented with proteinuria, hypertension
and renal failure. He was shown to be heterozygous for the relevant mutation
encoding the E526V fibrinogen variant. Hereditary renal amyloidosis is an autosomal domit condition with
considerable overlap with other amyloidosis types. Differential diagnosis is
complicated, but is relevant for prognosis and treatment. We describe a patient
with nephrotic syndrome and progressive renal failure, who had a mother with
renal amiloidosis. Renal biopsy revealed amyloid deposits in glomerular space,
with absence of light chains and protein AA. We suspected amyloidosis with
fibrinogen A alpha chain deposits, which is the most frequent cause of
hereditary amyloidosis in Europe, with a glomerular preferential affectation.
However, the genetic study showed a novel mutation in apolipoprotein AI. On
reviewing the biopsy of the patient's mother similar glomerular deposits were
found, but there were significant deposits in the renal medulla as well, which
is typical in APO AI amyloidosis. The diagnosis was confirmed by
immunohistochemistry. Apo AI amyloidosis is characterized by slowly progressive
renal disease and end-stage renal disease occurs aproximately 3 to 15 years from
initial diagnosis. Renal transplantation offers an acceptable graft survival and
in these patients with hepatorenal involvement simultaneous liver and kidney
transplantation could be considered. INTRODUCTION: Fibrinogen A alpha chain amyloidosis is an autosomal domit
disease associated with mutations in the fibrinogen A alpha chain (FGA) gene,
and it is the most common cause of hereditary renal amyloidosis in the UK.
Patients typically present with kidney impairment and progress to end-stage
renal disease over a median time of 4.6 years.
METHODS: Six patients presented with proteinuria, hypertension, and/or lower
limb edema and underwent detailed clinical and laboratory investigations.
RESULTS: A novel FGA gene mutation was identified in each case: 2 frameshift
mutations F521Sfs*27 and G519Efs*30 and 4 single base substitutions G555F,
E526K, E524K, R554H. In 5 subjects, extensive amyloid deposits were found solely
within the glomeruli, which stained specifically with antibodies to fibrinogen A
alpha chain, and in one of these cases, we found coexistent fibrinogen A alpha
chain amyloidosis and anti-glomerular basement membrane antibody disease. One
patient was diagnosed with light-chain amyloidosis after a bone marrow
examination revealed a small clonal plasma cell population, and laser
microdissection of the amyloid deposits followed by liquid chromatography and
tandem mass spectrometry identified kappa light chain as the fibril protein.
DISCUSSION: We report 6 novel mutations in the FGA gene: 5 were associated with
renal fibrinogen A alpha chain amyloidosis and 1 was found to be incidental to
light-chain amyloid deposits discovered in a patient with a plasma cell
dyscrasia. Clinical awareness and suspicion of hereditary amyloidosis
corroborated by genetic analysis and adequate typing using combined
immunohistochemistry and laser microdissection and mass spectrometry is valuable
to avoid misdiagnosis, especially when a family history of amyloidosis is
absent. |
What are the side effects of deflazacort in DMD patients? | The side effects observed in DMD patients following deflazacort treatment include growth failure and weight gain, facial fullness, blood pressure, bone health, cataracts, gastrointestinal symptoms, behavior, hypertrichosis, and need for medication interventions. |
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