Unveiling the miR-26b-3p/XAF1 Axis: A New Frontier in Systemic Lupus Erythematosus Inflammation Running Head: miR-26b-3p–XAF1 regulation in lupus

Background Systemic lupus erythematosus (SLE) is a complex autoimmune disorder characterized by dysregulated inflammatory responses, yet the molecular mechanisms underlying inflammation regulation remain incompletely understood. XAF1, traditionally recognized as a tumor suppressor, and miR-26b-3p, a microRNA with emerging immunoregulatory functions, have not been previously investigated in SLE pathogenesis. Aims To investigate the regulatory relationship between miR-26b-3p and XAF1 and determine their functional significance in modulating inflammatory responses associated with SLE. Methods THP-1 monocytes were differentiated into macrophages and stimulated with lipopolysaccharide (LPS) to establish an inflammatory model. Lentiviral vectors were employed to generate XAF1 overexpression and knockdown cell lines. Peripheral blood mononuclear cells (PBMCs) from SLE patients (n = 26) and healthy controls (n = 30) were analyzed. The miR-26b-3p-XAF1 interaction was validated using quantitative PCR, western blotting, dual-luciferase reporter assays, and gain/loss-of-function studies. Pro-inflammatory cytokine levels (IL-6, IL-15, TNF-α) were quantified by ELISA. Results LPS stimulation significantly upregulated XAF1 expression (2.7-fold mRNA, 2.4-fold protein, P < 0.05) alongside marked elevation of pro-inflammatory cytokines IL-6, IL-15, and TNF-α (2.9-4.2-fold increases, P < 0.01) in THP-1 cells. SLE patient PBMCs demonstrated significantly reduced miR-26b-3p expression (0.42-fold, P < 0.01) with reciprocal XAF1 upregulation, showing strong inverse correlation (r=-0.68, P < 0.001). Dual-luciferase reporter assays confirmed direct targeting of XAF1 3'UTR by miR-26b-3p (0.42-fold luciferase activity, P < 0.01). Functional studies revealed that XAF1 overexpression enhanced cytokine production (2.4-3.2-fold, P < 0.01), while miR-26b-3p mimics effectively suppressed both XAF1 expression and inflammatory cytokine secretion (40–55% reduction, P < 0.05). Conclusion Our findings establish the miR-26b-3p/XAF1 axis as a novel regulatory mechanism in SLE inflammation, with miR-26b-3p functioning as a negative regulator of XAF1-mediated inflammatory responses. This discovery highlights the therapeutic potential of targeting this axis for SLE treatment and advances our understanding of microRNA-mediated regulation in autoimmune inflammation.