Up‐Regulated Interleukin‐10 Induced by E2F Transcription Factor 2–MicroRNA‐17‐5p Circuitry in Extrafollicular Effector B Cells Contributes to Autoantibody Production in Systemic Lupus Erythematosus

• Published in: Arthritis & rheumatology (Hoboken, N.J.) Vol. 74; no. 3; pp. 496 - 507
• Main Authors: Xu, Lingxiao, Wang, Lei, Shi, Yumeng, Deng, Yun, Oates, Jim C., Kamen, Diane L., Gilkeson, Gary S., Wang, Fang, Zhang, Miaojia, Tan, Wenfeng, Tsao, Betty P.
• Format: Journal Article
• Language: English
• Published: Boston, USA Wiley Periodicals, Inc 01.03.2022; Wiley Subscription Services, Inc
• Subjects: 3' Untranslated regions; Adolescent; Adult; Aged; Antibodies; Antinuclear antibodies; Autoantibodies; Autoimmune diseases; B-Lymphocyte Subsets - metabolism; Cardiolipin; CD11c antigen; CD27 antigen; CD45RA antigen; Cell culture; Chronic conditions; Circuits; Correlation; CXCR3 protein; CXCR5 protein; Cytokines; Deoxyribonucleic acid; DNA; E2F protein; E2F Transcription Factors - metabolism; Effector cells; Female; Flow cytometry; Gene expression; Humans; Immunoglobulin D; Immunoglobulin G; Interleukin 1; Interleukin 10; Interleukin-10 - genetics; Interleukin-10 - metabolism; Interleukins; Leukocytes (mononuclear); Leukocytes, Mononuclear - metabolism; Lupus; Lupus Erythematosus, Systemic - genetics; Lupus Erythematosus, Systemic - metabolism; Lymphocytes B; Male; MicroRNAs; MicroRNAs - genetics; MicroRNAs - metabolism; Middle Aged; miRNA; Peripheral blood mononuclear cells; Proteinuria; Ribonucleic acid; RNA; siRNA; Systemic lupus erythematosus; Transcription factors; Up-Regulation; Young Adult
• DOI: 10.1002/art.41987
• ISSN: 2326-5191; 2326-5205

Objective Elevated interleukin‐10 (IL‐10) levels in patients with systemic lupus erythematosus (SLE) have B cell–promoting effects, contributing to autoantibody production and tissue damage. We aimed to characterize up‐regulated IL‐10+ B cell subsets and dysregulated IL10 expression in SLE B cells for new therapeutic options. Methods Proportions of Th10 and IL‐10+ B cell subsets in peripheral blood mononuclear cells (PBMCs) were assessed using flow cytometry. The IL10 3′‐untranslated region (3′‐UTR) dual‐luciferase vector was constructed and cotransfected with small interfering RNA (siRNA), microRNA (miRNA) mimics, or miRNA inhibitors into Raji cells. Transcript levels were quantified using TaqMan assays. Results Culture conditions that induced IL‐10+ Breg cells in healthy controls resulted in expansion of IL‐10+ double‐negative 2 (DN2; IgD−CD27−CD21−CD11c+) B cells in SLE PBMCs. Proportions of IL‐10+ DN2, but not those of IL‐10− DN2, correlated with disease activity and levels of antibodies to double‐stranded DNA (dsDNA) (r = 0.60, P = 0.03 for cohort 1; r = 0.38, P = 0.03 for cohort 2), and were associated with high levels or seropositivity of anti‐Sm (P = 0.03 for cohort 1; P = 0.01 for cohort 2) and IgG anticardiolipin (P < 0.01 for cohort 1; P = 0.02 for cohort 2) in SLE patients from 2 cohorts, of mainly African American subjects (cohort 1) and of Asian subjects (cohort 2). Proportions of Th10 (CD45RA−CXCR5−CXCR3+PD‐1highCD4+) cells correlated with IL‐10+ DN2 frequencies (r = 0.60, P < 0.01 for cohort 2), antinuclear antibody titers (r = 0.52, P = 0.01 for cohort 2), and proteinuria levels (r = 0.72, P < 0.01 for cohort 2) in SLE patients. Screening of predicted IL10 3′‐UTR–targeting miRNAs in SLE B cells identified miRNA‐17‐5p (miR‐17‐5p) and miR‐20a‐5p, with their levels inversely correlated with IL10 (r = −0.47, P < 0.01 for miR‐17‐5p; r = −0.37, P = 0.03 for miR‐20‐5p) and transcription factor E2F2 (r = −0.48, P = 0.04 for miR‐17‐5p; r = −0.45, P = 0.05 for miR‐20‐5p). In Raji cells, knockdown of E2F2 expression resulted in increased levels of miR‐17‐5p and miR‐20a‐5p and decreased IL10 messenger RNA (mRNA) and protein levels, and overexpression and inhibition of miR‐17‐5p down‐regulated and up‐regulated, respectively, IL10 mRNA levels, suggesting regulation of IL10 expression by an E2F2‐miR‐17‐5p loop. Conclusion IL‐10 promotes extrafollicular autoimmune responses in patients with active SLE, which might be dampened by targeting the E2F2–miR‐17‐5p circuitry.