Background/Purpose: Aberrant macrophage polarization is generally present in autoimmune diseases. Overwhelming M1 macrophage induces the continuous progression of inflammation, which is one of the vital reasons for the development of autoimmune diseases. However, the underlying mechanism is still unclear. This study mainly explores the role of regulation factor X1 (RFX1) on macrophage polarization in systemic lupus erythematosus (SLE), and elucidates the mechanism of RFX1 regulating abnormal macrophage polarization.

Methods: Western blot (WB) was used to detect RFX1 expression in CD14⁺ monocyte-derived macrophage (hMDMs) and mouse peritoneal macrophages (PMAs). The conditional knockout mice-Rfx1^f/fLyz2-Cre (CKO) was constructed to inhibit RFX1 expression specifically in macrophages. The RNA-seq was used to detect the differently expressed genes in RFX1-overexpressed or knockout PMAs. The expression levels of macrophage markers and pro-inflammatory cytokines in macrophages were detected by RT-qPCR, Flow cytometry or ELISA. The CKO and WTmice with colitis or lupus-like symptom were induced by dextran sulfate sodium (DSS) or IMQ respectively. The damage of intestinal or kidney tissue was examined by H&E staining. The infiltrations of CD45⁺, neutrophils, macrophages, CD4⁺ and CD8⁺ cells in colon or kidney were detected by flow cytometry, as well as markers of M1 and M2 subtypes. ChIP-seq was used to screen the target genes regulated by RFX1.

Results: The protein expression of RFX1 was increased significantly in LPS induced M1 macrophage. Compared with control group, the expressions of M1-related genes were increased in RFX1-overexpressed PMAs, while the expressions were significantly reduced in Rfx1-deficient PMAs. We also found that the relative mRNA and protein expression of RFX1 in CD14⁺ monocyte from the peripheral blood of SLE patients was significantly increased compared with healthy control (HC).

In addition, deficiency of Rfx1 protect the integrity of the intestinal structure in colitis mice and inhibited the damage to kidney from lupus-like mice. The infiltration of immune cells was obviously reduced in colon or kidney form CKO mice. The M1-related gene expression such CD86 or MHCII was decreased in macrophages from CKO mice. Besides, the concentrations of pro-inflammatory cytokines including Il-6 and Tnf-a and Il-1b were lessened significantly in serum from CKO mice.

Dual luciferase reporter and ChIP-qPCR assay indicated that APOBEC3A transcription was regulated by RFX1 directly. And the methylation of IL6 and TNF promoter in macrophage was decreased by APOBEC3A or RFX1 overexpression. Besides, the methylation of IL6 and TNF promoter were also decreased in monocyte and macrophage from SLE patients.

Conclusion: Both in vitro and in vivo experiments show that RFX1 promotes M1 polarization by APOBEC3A-mediated demethylation. And RFX1 may promoted the autoimmune inflammation in SLE.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/transcription-factor-rfx1-promotes-m1-macrophage-polarization-in-systemic-lupus-erythematosus-via-regulating-apobec3a/