Whole-Genome DNA Methylation Analysis of Peripheral Blood Mononuclear Cells in Multiple Sclerosis Patients with Different Disease Courses

Abstract

Multiple sclerosis (MS) is a severe neurodegenerative disease of polygenic etiology affecting the central nervous system. In addition to genetic factors, epigenetic mechanisms, primarily DNA methylation, which regulate gene expression, play an important role in MS development and progression. In this study, we have performed the first whole-genome DNA methylation profiling of peripheral blood mononuclear cells in relapsing-remitting MS (RRMS) and primary-progressive MS (PPMS) patients and compared them to those of healthy individuals in order to identify the differentially methylated CpG-sites (DMSs) associated with these common clinical disease courses. In addition, we have performed a pairwise comparison of DNA methylation profiles in RRMS and PPMS patients. All three pairwise comparisons showed significant differences in methylation profiles. Hierarchical clustering of the identified DMS methylation levels and principal component analysis for data visualization demonstrated a clearly defined aggregation of DNA samples of the compared groups into separate clusters. Compared with the control, more DMSs were identified in PPMS patients than in RRMS patients (67 and 30, respectively). More than half of DMSs are located in genes, exceeding the expected number for random distribution of DMSs between probes. RRMS patients mostly have hypomethylated DMSs, while in PPMS patients DMSs are mostly hypermethylated. CpG-islands and CpG-shores contain 60% of DMSs, identified by pairwise comparison of RRMS and control groups, and 79% of those identified by pairwise comparison of PPMS and control groups. Pairwise comparison of patients with two clinical MS courses revealed 51 DMSs, 82% of which are hypermethylated in PPMS. Overall, it was demonstrated that there are more changes in the DNA methylation profiles in PPMS than in RRMS. The data confirm the role of DNA methylation in MS development. We have shown, for the first time, that DNA methylation as an epigenetic mechanism is involved in the formation of two distinct clinical courses of MS: namely, RRMS and PPMS.

Keywords: DNA methylation; epigenetics; multiple sclerosis; peripheral blood mononuclear cells.

Fig. 1

Heatmaps of differentially methylated sites of peripheral blood mononuclear cells in RRMS patients vs. the healthy group (A), PPMS patients vs. healthy group (B), and PPMS patients vs. RRMS patients (C). Top panel. Dendrogram showing the results of hierarchical sample clustering. Green color indicates DNA samples from RRMS patients; blue color, from PPMS patients; black color, from healthy individuals. Left panel. Dendrogram showing the results of hierarchical DMS clustering. The intensity of grey indicates DMS localization in the genome (black color, CpG-islands, dark grey, CpG-shores, light gray, CpG-shelves). Right panel. Samples labeling based on Human Methylation450BeadChip standard annotation. RRMS, relapsing-remitting multiple sclerosis; PPMS, primary progressive multiple sclerosis.

Fig. 2

2D (A–C) and 3D (D) samples clustering based on a principal component analysis (PCA) of differentially methylated sites (DMSs). The green dots indicate RRMS samples; blue dots PPMS samples; and black dots, healthy controls. The axes: the principal components PC1, PC2, and PC3; the proportion of explained variance of the data is indicated in brackets for each principal component.