A molecular characterization of meningeal inflammation in progressive MS brain

Abstract

Introduction: The presence of lymphoid-like immune cell aggregates in the leptomeninges is suggested to promote damage to the cerebral cortex and play a role in accumulating disability in multiple sclerosis (MS). Methods: Cryosections were cut from five cortical blocks per case from 55 SPMS, 12 PPMS and 14 control post-mortem brains. Meningeal tissue was dissected and RNA extracted. Affymetrix HTA 2.0 GeneChips were used to obtain the meningeal transcriptome and gene expression determined using R package Limma. Differentially expressed genes with FC>2 and FDR>0.05 were used to perform gene set enrichment analysis using WebGestalt and gene networks constructed using R package WGCNA. Results: When comparing controls with highly inflamed SPMS cases, alterations were mainly found in expression of homing chemokines and immune receptors and that enhance B cell survival, co-stimulation, proliferation and antibody and IFNγ production (SLAMF7, CXCR4, GZMK, CD27). Gene network analysis revealed networks associated with immune response, specially highlighting processes such as response to virus, chemokine signalling and antigen presentation. Functional pathway analysis for these network modules identified significant involvement in the MS meninges, of pathways associated with cell adhesion, TCR and BCR co-stimulation and activation, Th17, Th1/Th2 cell differentiation, haematopoietic and lymphoid organ development. The most highly altered genes were loaded into Ingenuity Pathway Analysis, yielding results that suggest a dysregulation of pathways critical for B-T cell interaction, disrupting antigen presentation and its subsequent outcomes. A panel of 55 inflammatory genes was chosen for validation of the microarray data using TaqMan OpenArrays. Highly significant, strong and moderately strong, correlations were found between the TaqMan and Affymetrix data for most individual genes, providing robustness to our meningeal transcriptomics data. Remarkably, we found a very different signature when analysing PPMS data pointing towards an involvement of the innate immune system, featuring alterations in the signalling axis involving chemokine receptors CXCR1, CXCR2, and their ligands CXCL8 and CXCL7. Besides, preliminary functional enrichment analysis mainly highlights the pathways granulocyte activation, platelet 2 degranulation, neutrophil mediated immunity, processes that did not appear as enriched in SPMS data. Altogether, suggesting that the immune response might be differently built or initiated in PPMS compared to SPMS. Conclusions: We have identified molecular cues that are likely to play a major role in mediating meningeal inflammation in MS, which suggest a dysregulation of pathways that are critical for B cell trafficking and recruitment into the CNS