Upregulated IL-10 induced by E2F2-miR-17-5p circuitry in extrafollicular effector B cells contributes to autoantibody production in systemic lupus erythematosus

OBJECTIVE Elevated interleukin 10 (IL10) in SLE patients has B-cell promoting effects, contributing to autoantibody production and tissue damage. We aimed to characterize upregulated IL10+ B-cell subsets and dysregulated IL10 expression in SLE B cells for new therapeutic options. METHODS Proportions of Th10 and IL10+ B-cell subsets in PBMCs were assessed using flow cytometry. The IL10-3'UTR dual-luciferase vector was constructed and co-transfected with siRNA, miRNA mimics or inhibitors into RAJI cell line. Transcript levels were quantified using Taqman assays. RESULTS Culture conditions that induced IL10+ Breg in healthy controls (HC) resulted in expansion of IL10+ DN2 (IgD- CD27- CD21- CD11c+ ) B-cells in SLE PBMCs. Proportions of IL10+ DN2, but not those of IL10- DN2, correlated with disease activity, levels of antibodies to dsDNA, and associated with high levels or seropositivity of anti-Smith and anti-cardiolipin IgG in SLE patients from two cohorts of mainly African Americans and Asians, respectively. Proportions of Th10 (CD45RA- CXCR5- CXCR3+ PD1hi CD4+ ) cells correlated with IL10+ DN2 frequencies, ANA titers and proteinuria levels in SLE patients. Screening predicted IL10 3'UTR-targeting miRNAs in SLE B cells identified miR-17-5p and miR-20a-5p with their levels inversely correlated with IL10 and transcription factor E2F2. In RAJI cells, knockdown of E2F2 expression resulted in increased levels of miR-17-5p and miR-20a-5p and decreased IL10 mRNA and protein levels, and overexpression and inhibition of miR-17-5p down-regulated and up-regulated IL10 mRNA levels respectively; suggesting regulation of IL10 expression by E2F2-miR-17-5p loop. CONCLUSION IL10 promotes extrafollicular autoimmune responses in active SLE patients, which might be dampened by targeting the E2F2-miR-17-5p circuitry.