Professor of Immune Regulation & Inflammation King's College London London, England, United Kingdom
Abstract Text: CD4+ T-cells are key players in the pathogenesis of rheumatoid arthritis (RA) through their production of pro-inflammatory mediators. Anti-TNF therapy has revolutionised the treatment of RA and we previously demonstrated that anti-TNF treatment in vitro promotes anti-inflammatory IL-10 expression in multiple subpopulations of CD4+ T-cells. Here we investigate the transcriptional mechanisms underlying the IL-10 induction by TNF-blockade in in vitro stimulated CD4+ T-cells.
CD4+ T-cells were isolated from PBMCs of healthy volunteers and stimulated polyclonally for 3 days in the absence or presence of anti-TNF. After culture, CD45RA+ cells were depleted. RNA was extracted for gene expression profiling by RNA-seq and cell nuclei were isolated for chromatin accessibility analysis by ATAC-seq.
Gene expression analysis of memory CD4+ T-cells showed a distinct gene signature of 183 genes (q-value < 0.05) conferred by anti-TNF treatment. Pathway enrichment analysis of the differentially expressed genes revealed multiple pathways related to regulation of cytokine signalling and production; in particular, IL10 was the most upregulated gene by anti-TNF, while the proinflammatory cytokines and chemokines IFNG, IL9, IL22 and CXCL10 were significantly downregulated (q-value < 0.05). Transcription factor (TF) motif analysis at the differentially open chromatin regions after anti-TNF treatment revealed 58 shared TF motifs enriched at the IL10 locus. We identified 7 TF candidates that were either differentially expressed or whose locus was differentially accessible upon anti-TNF stimulation. We postulate that this TF network, which includes MAF and PRMD1, drives transcriptional regulation of IL-10 in anti-TNF treated T-cells.
