Synovial Fluid Extracellular Vesicles from Patients with Severe Osteoarthritis Differentially Promote a Pro-Catabolic, Inflammatory Chondrocyte Phenotype

Synovial inflammation is recognised as a pathological driver of osteoarthritis (OA), a degenerative joint disease involving cartilage degradation and joint pain. Since extracellular vesicles (EVs) have emerged as key mediators of cellular cross-talk, this study characterised synovial fluid EVs (SFEVs) in OA patients with varying disease severity and determined their functional effects on OA articular chondrocytes. Synovial fluid and articular cartilage were collected from patients undergoing knee surgery. SFEVs were isolated via ultracentrifugation and characterised by nanoparticle tracking analysis, ExoView, and Luminex analysis of protein cargo. Patients were stratified into mild/moderate- and severe-OA groups based on Oxford Knee Score and EQ5D. Chondrocytes were treated with SFEVs, and transcriptomic and secretome responses were analysed using RNA sequencing, Luminex, and ELISA. SFEVs from patients with severe OA were more abundant, smaller and exhibited increased tetraspanin expression. Synovial fluid and SFEVs induced distinct transcriptomic changes in chondrocytes. SFEVs from patients with severe OA promoted a pro-inflammatory and catabolic chondrocyte phenotype, with upregulation of <i>CRTAC1</i>, <i>COL6A3</i>, <i>TNC</i>, and <i>CXCL5</i>, greater secretion of IL-6, MMP1, MMP3 and MMP13, and pro-nociceptive mediators (NGF and Substance P). These findings suggest that SFEVs may contribute to OA progression by exacerbating cartilage damage and promoting pain sensitisation.