Deep digital flexor tendon (DDFT) and navicular bone fibrocartilage (NBF) undergo degeneration in navicular disease and have limited innate healing capacity. Extracellular vesicles (EV) from mesenchymal stem cells (MSC) may provide therapeutic benefits of cell-based therapies with ‘off-the-shelf’ convenience. We assess equine bone marrow MSC (BM-MSC)-derived extracellular vesicles (BM-EV) for anti-inflammatory cytokines and anabolic growth factors. Then we evaluate the effects of BM-EV on DDFT-NBF co-cultures with IL- hypothesizing that BM-EV mitigate inflammation. BM-EV isolated from BM-MSCs (n=5) via ultracentrifugation were analyzed, including protein estimation, cytokine/growth factor quantification, flow cytometry, and mass spectrometry. Effects on tenocyte proliferation (MTT assay) and migration (Scratch assay) were assessed. DDFT-NBF transwell co-cultures (n=7) with IL- were treated with BM-EV. Culture media MMP-3, MMP-13, and IL-6 were quantified using ELISA. DMMB was performed for GAG content. T-test, one-way ANOVA or the non-parametric equivalent were performed (p<0.05). Mean concentrations of 119.7±56.44ng/mL, 0.009±0.03pg/mL, and 0.02±0.04ng/mL were measured for IL-1ra, IL-6, and IL-10, respectively; VEGF, TGF, and PDGF were 38.4±110.1pg/mL, 5.055±1.664pg/mL, and 2.107±2.261pg/mL, respectively. BM-EV increased tenocyte proliferation; migration was unaffected. MMP-3 was decreased in BM-MV control and treatment groups relative to media and IL-1β controls. MMP-13 was increased in IL-1β control and treatment groups relative to media control. In DDFT, GAG content was higher in the BM-MV control relative to IL-1β control. Equine BM-EV possess anti-inflammatory cytokines and anabolic growth factors. MMP-3 and DDFT GAG data suggest anti-inflammatory and matrix-preserving potential of BM-MVs, respectively. In vivo studies with naturally occurring disease are needed to determine clinical applications.
