Background/Purpose:  T cell DNA methylation defects play an important role in the pathogenesis of systemic lupus erythematosus. A CD4+CD28+ T cell subset characterized by overexpression of methylation sensitive genes, including CD11a and the KIR gene cluster, has been recently described in autoimmunity. The goal of this study was to characterize this T cell subset in vitro and in patients with SLE.

Methods:  The size of the CD4+CD28+KIR+CD11a^hi T cell subset was determined by flow cytometry in 49 female lupus patients of European descent. SLEDAI scores were calculated at the time of blood draw, and all patients were genotyped across 43 confirmed lupus susceptibility loci and a total genetic risk score for lupus was calculated. CD4+CD28+KIR+CD11a^hi and CD4+CD28+KIR-CD11a^low T cells were isolated from peripheral blood samples of normal healthy individuals after in vitro treatment with 5-azacytidine, and genome-wide DNA methylation and RNA sequencing was performed in both subsets. RNA sequences in the CDR3 region were used to assess the TCR repertoire.

Results:  The CD4+CD28+KIR+CD11a^hi T cell subset size correlated with disease activity in lupus patients as measured by SLEDAI score (r=0.36, P= 0.012). Linear regression models suggest that the subset size is a better predictor of disease activity when normalized for total genetic risk in each individual (r=0.42, P=0.003), suggesting that the relationship between KIR+CD11a^hi T cell subset size and disease activity in lupus is influenced by genetic risk. Genome-wide DNA methylation analysis identified a total of 31,019 differentially methylated sites in KIR+CD11a^hi compared to autologous KIR-CD11a^low T cells, with 30,736 methylation sites (99.1%) being hypomethylated. RNA sequencing analysis identified 1,620 overexpressed genes in the KIR+CD11a^hi T cell subset, including key pro-inflammatory cytokine genes, adhesion molecules, Fc-gamma receptor genes, Toll-like receptor genes, HLA molecules, and metalloproteinases. Of particular interest, and similar to what is known in lupus T cells, the experimentally derived demethylated KIR+CD11a^hi T cell subset demonstrates reduced IL-2 mRNA expression (3.3-fold). Bioinformatics analysis of 718 genes that are hypomethylated and overexpressed in KIR+CD11a^hi T cells revealed significant enrichment in pro-inflammatory gene ontologies, pathways, and gene meta-groups. Analysis of the TCR repertoire suggests that the KIR+CD11a^hi T cell subset is polyclonal, but with less diverse clonality compared to autologous KIR-CD11a^low T cells (mean number of TCR clones 673±72 versus 837±39; P= 0.001).

Conclusion:  CD4+CD28+KIR+CD11a^hi T cells are polyclonal and demethylated, and characterized by a pro-inflammatory transcriptional profile. These cells are expanded in lupus patients, and interact with total genetic risk to predict and possibly contribute to disease activity.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/demethylated-cd4cd28kircd11ahi-t-cells-are-characterized-by-a-pro-inflammatory-transcriptome-and-interact-with-genetic-risk-to-predict-disease-activity-in-lupus/