Abstract
BackgroundAlthough the etiology of primary biliary cirrhosis (PBC) remains enigmatic, there are several pieces of data supporting the thesis that a strong genetic predisposition and environmental factors interact to produce a selective loss of tolerance. The striking female predominance of PBC has suggested that this sex predisposition may be secondary to epigenetic alterations on the X chromosome. In the present study, we rigorously defined the X chromosome methylation profile of CD4, CD8, and CD14 cells from 30 PBC patients and 30 controls. Genomic DNA from sorted CD4, CD8, and CD14 subpopulations was isolated, sonicated, and immunoprecipitated for analysis of methylation. All products were hybridized to a custom-tiled four-plex array containing 27,728 CpG islands annotated by UCSC and 22,532 well-characterized RefSeq promoter regions. Furthermore, bisulfite sequencing was then used for validation on a subsequent group of independent samples from PBC patients and controls. Thence, expression levels of selected X-linked genes were evaluated by quantitative real-time PCR with cDNA samples from all subjects.ResultsWe report herein that a total of 20, 15, and 19 distinct gene promoters reflected a significant difference in DNA methylation in CD4+ T, CD8+ T, and CD14+ cells in patients with PBC. Interestingly, there was hypermethylation of FUNDC2 in CD8+ T cells and a striking demethylation of CXCR3 in CD4+ T cells, which inversely correlated with CXCR3 expression levels in CD4+ T cells from early-stage PBC patients.ConclusionsOur data provides a set of genes with epigenetic alteration likely to be indicators of autoimmunity and emphasizes the role of CXCR3 in the natural history of PBC.Electronic supplementary materialThe online version of this article (doi:10.1186/s13148-015-0098-9) contains supplementary material, which is available to authorized users.
Highlights
The etiology of primary biliary cirrhosis (PBC) remains enigmatic, there are several pieces of data supporting the thesis that a strong genetic predisposition and environmental factors interact to produce a selective loss of tolerance
We report that the CXCR3 promoter is demethylated in CD4+ T cells from PBC patients and the expression level of CXCR3 in CD4+ T cells is significantly increased in comparison to controls and inversely correlated with the overall methylation levels of the CXCR3 promoter, indicating that promoter demethylation status contributed to CXCR3 overexpression in CD4+ T cells
DNA methylation Initially, X chromosome methylated DNA immunoprecipitation (MeDIP) arrays were performed on cell subsets from 10 PBC patients and 10 controls (Fig. 1)
Summary
The etiology of primary biliary cirrhosis (PBC) remains enigmatic, there are several pieces of data supporting the thesis that a strong genetic predisposition and environmental factors interact to produce a selective loss of tolerance. Genetic background confers susceptibility to disease onset but is not sufficient for disease development Epigenetic mechanisms such as DNA methylation and histone modifications regulate gene expression levels and provide an alternative mechanism for modulating gene function. We performed an extensive X chromosome methylation analysis in CD4, CD8, and CD14 cells from PBC patients and controls, coupled with gene expression profile from the same cells. The frequency of CXCR3-expressing cells in peripheral blood is known to be higher in PBC patients, and CXCR3positive cells are found in the portal areas of diseased livers, primarily on CD4+ T cells [27].
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