Transciptome of B lymphocytes in multiple sclerosis

Objective: Over the past two decades B cells have increasingly moved into the spotlight in multiple sclerosis (MS) research. So far it has not been performed a detailed study of the B cells transcriptome in MS. This may be a prerequisite for identifying the elements that B cells use to carry out their pathogenic role. The primary aim of this project was to analyze the whole B cell-transcriptome (coding and non-coding RNA) in MS context. We studied the differences between MS patients and controls in B cell transcriptome: 1) mRNA including alternative splicing; 2) human microRNAs (miR). Methods: We studied peripheral CD19+ B cell subsets obtained from 10 MS patients, 10 healthy donors (HD), and 2 monozygotic twins discordant for MS. All affected individuals were treatment-naïve and all the investigations were performed at least three months after the last steroid therapy. Total RNA was extracted using miRNeasy Kit (Qiagen). Microarray data was performed using two Affymetrix’s platforms: GeneChip-Human Exon and GeneChip-miRNA. The statistical analysis was performed by means of two different software packages, whose output results have been compared: Partek Genomic Suite (version 6.6) and our own analysis pipe-line based on EasyExon tools. In order to discover and understand the connections among the genes obtained from the analysis, and their position in other biological processes, a pathway analysis was performed using Ingenuity Pathway Analysis (IPA). The miR Analysis was performed by IPA microRNA Target Filter. Results: Firstly, we found that EasyExon is much less permissive than Partek. So we focused on EasyExon results, in particular we selected 26 genes differentially expressed in MS compared HD (p=1.5). The list was then submitted to IPA and performed a pathway analysis that led us to choose seven, among the original 26 genes that were among those differentially expressed MS patients and HD. Furthermore, performing the miR analysis with IPA, it has emerged that 6 genes from our list are targeted by 9 miR differentially expressed in MS/HD groups. Conclusions: Peripheral B cell compartment seems to be promising for the identification of mRNA and miR signature. We have identified an MS-related pattern, in terms of miR and/or genes