Abstract

Objectives: To investigate the mechanism by which fibroblast-derived extracellular vesicles (EVs) carrying miR-25-3p alleviate knee osteoarthritis (KOA) with proof-of-concept (POC) in a murine KOA model through targeting TAF15 to inhibit NF-κB signaling pathway activation. Methods: miR-25-3p mimic/inhibitor-transfected murine fibroblast EVs were co-cultured with ATDC5 chondrocytes. Chondrocyte proliferation, migration, and apoptosis were assessed via CCK-8, Transwell, and TUNEL assays. Inflammatory cytokines (TNF-α, IL-1β, IL-6) were measured by ELISA and qPCR. miR-25-3p/TAF15 binding was verified via dual-luciferase assay, with TAF15 expression and NF-κB subunit (p65/IκBα) interactions analyzed by Western blot and co-immunoprecipitation (Co-IP). In vivo, monosodium iodoacetate (MIA)-induced KOA mice received intra-articular miR-25-3p-loaded EVs, with therapeutic effects evaluated by ELISA, H&E staining, and immunohistochemistry. Results: Fibroblast-derived EVs carrying miR-25-3p promoted chondrocyte proliferation and migration, inhibited apoptosis, and reduced inflammatory cytokine secretion in vitro. Mechanistically, miR-25-3p directly targeted TAF15, downregulating its expression and disrupting interactions between TAF15 and p65/IκBα, thereby suppressing NF-κB nuclear translocation and transcriptional activity. In the KOA mouse model, intra-articular administration of miR-25-3p-loaded EVs alleviated cartilage degradation, synovial inflammation, and pain sensitivity, thus confirming the POC of this EV-based strategy in murine KOA accompanied by decreased NF-κB-mediated pro-inflammatory gene expression. Conclusion: Fibroblast-derived EVs delivering miR-25-3p mitigate KOA progression in a murine model by targeting TAF15 to inhibit NF-κB signaling as verified by POC in a murine KOA model, highlighting a novel EV-based therapeutic strategy for experimental KOA.