Synovial mesenchymal stem cells derived exosomes circRNF20 aggravated rheumatoid arthritis via TLR4/NETs/IL-6 pathway

Rheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by persistent synovial inflammation and progressive joint destruction. Neutrophil extracellular traps (NETs) and toll-like receptor 4 (TLR4) signaling have been implicated in RA pathogenesis, primarily through the formation of inflammatory amplification loops. The present study was designed to investigate the regulatory mechanism by which exosomes circRNF20 derived from synovial mesenchymal stem cells (SMSCs) modulates the TLR4/NETs/IL-6 axis in RA. Exosomes carrying circRNF20 were isolated from LPS-stimulated SMSCs and characterized by electron microscopy and Western blot. Fibroblast-like synoviocytes (FLS) were co-cultured with exosomes, and transfections with si-circRNF20 or microRNA-921 (miR-921) mimic were performed. A collagen-induced arthritis (CIA) rat model was established. Expression of TLR4, NETs markers (MPO, Cit H3, PAD4), and cytokines (IL-6, TNF-α, IL-1β) was measured by Western blot, ELISA, and immunofluorescence. Cell proliferation and apoptosis were assessed via CCK8 and TUNEL assays. Inflammatory stimulation increased circRNF20 abundance in SMSC-derived exosome preparations, which increased circRNF20 and TLR4 expression in FLSs, NET-related markers in neutrophils, IL-6 production, and proliferative and inflammatory responses. CircRNF20 silencing in donor SMSCs attenuated these changes. In the human recipient-cell system, hsa-miR-921 overexpression reduced EV-associated increases in TLR4, NET-related markers, and IL-6. DNase I reduced TLR4 expression and IL-6 production, supporting a contribution of NET-associated structures without establishing a feedback loop. In CIA rats, exosomes from circRNF20-silenced rSMSCs were associated with reduced synovial inflammation and histopathological damage. SMSC EV-associated circRNF20 may contribute to inflammatory responses in RA. Not applicable.