Platelet-derived mitochondrial transplantation expands respiratory reserve and extracellular matrix anabolism in primary osteoarthritic chondrocytes

Osteoarthritis (OA) is a leading cause of disability worldwide, driven by the complex interplay of aging, obesity, and mechanical stress, which converge on progressive deterioration of cartilage, subchondral bone, and synovial tissue. Current treatments alleviate symptoms but fail to halt the degenerative process, making joint replacement surgery the only definitive option for advanced disease. A central mechanism underlying OA pathogenesis is mitochondrial dysfunction in chondrocytes, in which the balance between oxidative phosphorylation (OXPHOS) and glycolysis is disrupted, leading to impaired OXPHOS, accumulation of reactive oxygen species (ROS), and sustained overproduction of matrix-degrading enzymes and pro-inflammatory cytokines. Mitochondrial transplantation has emerged as a promising strategy to restore cellular bioenergetics, and platelets, which contain functional, respiration-competent mitochondria, represent an accessible allogenic donor source. Mitochondria isolated from platelets of healthy donors were structurally and functionally characterized and transplanted into primary human OA chondrocytes by mitoception at increasing chondrocyte-to-mitochondria ratios. Internalization was assessed by flow cytometry and confocal colocalization analysis. Bioenergetic effects were quantified by Seahorse extracellular flux analysis; mitochondrial membrane potential and superoxide production were measured by flow cytometry; extracellular matrix protein expression was evaluated by immunofluorescence and RT-qPCR; and the inflammatory secretome was profiled by Luminex multiplex assay. Isolated platelet mitochondria preserved outer membrane integrity, membrane potential, and ADP-coupled respiratory activity prior to transplantation. They were efficiently internalized by OA chondrocytes in a dose-dependent manner, with optimal uptake observed at chondrocyte-to-platelet-equivalent mitochondria dose ratios of 1:100. Transplantation increased maximal respiratory capacity and spare respiratory reserve without altering basal respiration, was accompanied by elevated type II collagen and aggrecan protein levels, and selectively reduced secreted IL-1β and osteopontin while leaving IL-6, TNF-α, IL-10, and IL-1RA unchanged. Mitochondrial superoxide production was modestly increased and was prevented by the mitochondria-targeted antioxidant MitoTEMPO, while mitochondrial membrane potential remained unchanged. Platelet-derived mitochondrial transplantation expands the respiratory reserve and modulates the catabolic-anabolic balance of human OA chondrocytes without inducing mitochondrial depolarization, providing a mechanistic basis for allogenic, blood-derived mitochondrial therapies in joint disease.