Summary
The altered expression of micro‐RNA (miRNA) has been associated with Crohn's disease (CD) and ulcerative colitis (UC). The aim of this study was to establish specific miRNA expression ...patterns in the serum and mucosa of inflammatory bowel disease (IBD) patients (UC and CD with colonic involvement) at different stages of the disease. Serum and biopsies from nine active CD (aCD), nine inactive CD (iCD), nine active UC (aUC) and nine inactive UC (iUC) and serum from 33 healthy subjects were collected. Up to 700 miRNAs were evaluated by the TaqMan® human miRNA array. The ΔCt values were obtained using the mean expression values of all expressed miRNAs in a given sample as a normalization factor for miRNA real‐time quantitative polymerase chain reaction data. The levels of serum miRNAs in CD and UC patients were different to healthy subjects. Thirteen serum miRNAs were expressed commonly in CD and UC patients. Two miRNAs were higher and four miRNAs were lower in the serum of aCD than iCD. No serum miRNA was regulated exclusively in aUC compared with iUC patients. Four miRNAs were higher and three miRNAs were lower in the mucosa of aCD than iCD. Two miRNAs were higher and three miRNAs were lower in the mucosa of aUC than iUC. No serum miRNAs coincided with tissue miRNAs in aCD and aUC patients. Our results suggest the existence of specific miRNA expression patterns associated with IBD and their different stages and support the utility of miRNA as possible biomarkers. This pilot study needs to be validated in a large prospective cohort.
Summary
The diagnosis of primary sclerosing cholangitis (PSC) is difficult due to the lack of sensitive and specific biomarkers, as is the early diagnosis of cholangiocarcinoma (CC), a complication ...of PSC. The aim of this study was to identify specific serum miRNAs as diagnostic biomarkers for PSC and CC. The levels of 667 miRNAs were evaluated in 90 human serum samples (30 PSC, 30 CC and 30 control subjects) to identify disease‐associated candidate miRNAs (discovery phase). The deregulated miRNAs were validated in an independent cohort of 140 samples 40 PSC, 40 CC, 20 primary biliary cirrhosis (PBC) and 40 controls. Receiver operating characteristic (ROC) curves were established and only miRNAs with an area under the curve (AUC) > 0·70 were considered useful as biomarkers. In the discovery phase we identified the following: 21 miRNAs expressed differentially in PSC, 33 in CC and 26 in both in comparison to control subjects as well as 24 miRNAs expressed differentially between PSC and CC. After the validation phase, miR‐200c was found to be expressed differentially in PSC versus controls, whereas miR‐483‐5p and miR‐194 showed deregulated expression in CC compared with controls. We also demonstrate a difference in the expression of miR‐222 and miR‐483‐5p in CC versus PSC. Combination of these specific miRNAs further improved the specificity and accuracy of diagnosis. This study provides a basis for the use of miRNAs as biomarkers for the diagnosis of PSC and CC.
The Illumina Infinium 450 k DNA Methylation Beadchip is a prime candidate technology for Epigenome-Wide Association Studies (EWAS). However, a difficulty associated with these beadarrays is that ...probes come in two different designs, characterized by widely different DNA methylation distributions and dynamic range, which may bias downstream analyses. A key statistical issue is therefore how best to adjust for the two different probe designs.
Here we propose a novel model-based intra-array normalization strategy for 450 k data, called BMIQ (Beta MIxture Quantile dilation), to adjust the beta-values of type2 design probes into a statistical distribution characteristic of type1 probes. The strategy involves application of a three-state beta-mixture model to assign probes to methylation states, subsequent transformation of probabilities into quantiles and finally a methylation-dependent dilation transformation to preserve the monotonicity and continuity of the data. We validate our method on cell-line data, fresh frozen and paraffin-embedded tumour tissue samples and demonstrate that BMIQ compares favourably with two competing methods. Specifically, we show that BMIQ improves the robustness of the normalization procedure, reduces the technical variation and bias of type2 probe values and successfully eliminates the type1 enrichment bias caused by the lower dynamic range of type2 probes. BMIQ will be useful as a preprocessing step for any study using the Illumina Infinium 450 k platform.
BMIQ is freely available from http://code.google.com/p/bmiq/.
a.teschendorff@ucl.ac.uk
Supplementary data are available at Bioinformatics online.
The high-throughput analysis of microRNAs (miRNAs) circulating within the blood of healthy and diseased individuals is an active area of biomarker research. Whereas quantitative real-time reverse ...transcription polymerase chain reaction (qPCR)-based methods are widely used, it is yet unresolved how the data should be normalized. Here, we show that a combination of different algorithms results in the identification of candidate reference miRNAs that can be exploited as normalizers, in both discovery and validation phases. Using the methodology considered here, we identify normalizers that are able to reduce nonbiological variation in the data and we present several case studies, to illustrate the relevance in the context of physiological or pathological scenarios. In conclusion, the discovery of stable reference miRNAs from high-throughput studies allows appropriate normalization of focused qPCR assays.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Regular endurance exercise training induces beneficial functional and health effects in human skeletal muscle. The putative contribution to the training response of the epigenome as a mediator ...between genes and environment has not been clarified. Here we investigated the contribution of DNA methylation and associated transcriptomic changes in a well-controlled human intervention study. Training effects were mirrored by significant alterations in DNA methylation and gene expression in regions with a homogeneous muscle energetics and remodeling ontology. Moreover, a signature of DNA methylation and gene expression separated the samples based on training and gender. Differential DNA methylation was predominantly observed in enhancers, gene bodies and intergenic regions and less in CpG islands or promoters. We identified transcriptional regulator binding motifs of MRF, MEF2 and ETS proteins in the proximity of the changing sites. A transcriptional network analysis revealed modules harboring distinct ontologies and, interestingly, the overall direction of the changes of methylation within each module was inversely correlated to expression changes. In conclusion, we show that highly consistent and associated modifications in methylation and expression, concordant with observed health-enhancing phenotypic adaptations, are induced by a physiological stimulus.
Members of the transient receptor potential (TRP) channels are involved in mediating the electrical excitability and stimulus-secretion coupling in the pancreatic β-cells. The expression and the ...relative abundance of different TRP channels in the human β-cells are unknown. The objective of this study was to examine the expression of the TRP channels and their relative abundance in the human β-cell.
RNA sequencing data obtained from human islets, fluorescence-activated cell sorting-purified human β-cell and human pancreatic acinar cells were analyzed. Gene counts and fragments per kilobase per million mapped reads were obtained.
Among the TRPC family only the TRPC1 was expressed in the human β-cell. TRPV1 channels were not expressed in the human β-cells. Among the TRPM family, TRPM4, TRPM7, TRPM2, and TRPM3 were expressed in the human β-cell. Of the remaining TRP channels, TRPP2, TRPML1, and TRPML3 were expressed in these cells.
By analyzing the RNA sequencing data, we have detected for the first time the TRP channels that are expressed in the purified human β-cells, in comparison to the other relevant pancreatic cell types. Our study provides an opportunity to focus on these TRP channels for a better understanding of the electrophysiology and stimulus-secretion coupling in these cells.
The proper identification of differentially methylated CpGs is central in most epigenetic studies. The Illumina HumanMethylation450 BeadChip is widely used to quantify DNA methylation; nevertheless, ...the design of an appropriate analysis pipeline faces severe challenges due to the convolution of biological and technical variability and the presence of a signal bias between Infinium I and II probe design types. Despite recent attempts to investigate how to analyze DNA methylation data with such an array design, it has not been possible to perform a comprehensive comparison between different bioinformatics pipelines due to the lack of appropriate data sets having both large sample size and sufficient number of technical replicates. Here we perform such a comparative analysis, targeting the problems of reducing the technical variability, eliminating the probe design bias and reducing the batch effect by exploiting two unpublished data sets, which included technical replicates and were profiled for DNA methylation either on peripheral blood, monocytes or muscle biopsies. We evaluated the performance of different analysis pipelines and demonstrated that: (1) it is critical to correct for the probe design type, since the amplitude of the measured methylation change depends on the underlying chemistry; (2) the effect of different normalization schemes is mixed, and the most effective method in our hands were quantile normalization and Beta Mixture Quantile dilation (BMIQ); (3) it is beneficial to correct for batch effects. In conclusion, our comparative analysis using a comprehensive data set suggests an efficient pipeline for proper identification of differentially methylated CpGs using the Illumina 450K arrays.
Cigarette smoking is an established environmental risk factor for Multiple Sclerosis (MS), a chronic inflammatory and neurodegenerative disease, although a mechanistic basis remains largely unknown. ...We aimed at investigating how smoking affects blood DNA methylation in MS patients, by assaying genome-wide DNA methylation and comparing smokers, former smokers and never smokers in two Swedish cohorts, differing for known MS risk factors. Smoking affects DNA methylation genome-wide significantly, an exposure-response relationship exists and the time since smoking cessation affects methylation levels. The results also show that the changes were larger in the cohort bearing the major genetic risk factors for MS (female sex and HLA risk haplotypes). Furthermore, CpG sites mapping to genes with known genetic or functional role in the disease are differentially methylated by smoking. Modeling of the methylation levels for a CpG site in the AHRR gene indicates that MS modifies the effect of smoking on methylation changes, by significantly interacting with the effect of smoking load. Alongside, we report that the gene expression of AHRR increased in MS patients after smoking. Our results suggest that epigenetic modifications may reveal the link between a modifiable risk factor and the pathogenetic mechanisms.
Accumulating evidence links epigenetic age to diseases and age-related conditions, but little is known about its association with multiple sclerosis (MS).
We estimated epigenetic age acceleration ...measures using DNA methylation from blood or sorted cells of MS patients and controls.
In blood, sex (p = 4.39E-05) and MS (p = 2.99E-03) explained the variation in age acceleration, and isolated blood cell types showed different epigenetic age. Intrinsic epigenetic age acceleration and extrinsic epigenetic age acceleration were only associated with sex (p = 2.52E-03 and p = 1.58E-04, respectively), while PhenoAge Acceleration displayed positive association with MS (p = 3.40E-02).
Different age acceleration measures are distinctly influenced by phenotypic factors, and they might measure separate pathophysiological aspects of MS.
DNA methylation data can be accessed at Gene Expression Omnibus database under accession number GSE35069, GSE43976, GSE106648, GSE130029, GSE130030.
DNA methylation data from blood and sorted cells were employed to calculate several measures of age acceleration in multiple sclerosis patients and healthy controls.
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