Changes on the expression of ectopic inhibitor protein subunit of f1-f0 atp synthase (if1) in osteoarthritis

Abstract

Purpose: Osteoarthritis (OA) is a disease characterized by the cartilage degradation, and involves the entire joint, including the subchondral bone, synovial membrane, menisci and periarticular structures. The inflammation and angiogenesis occurs at the osteochondral junction as well as within the osteoarthritic synovium providing a potential mechanism for the genesis of pain in knee OA. Menisci play a role in the OA disease while is injured and there is a strong association between meniscal damage/degenerative tears and cartilage loss with pathological changes extracellular matrix degeneration with secretion of cytokines and growth factors. A few studies have investigated on meniscal cells in the process of OA and angiogenesis. Previous studies have demonstrated the ectopic localization of mitochondrial F1-F0 ATP synthase as a target for anti-angiogenesis intervention. It has been shown that β subunits of ATP synthase bind angiostatin on cell surface of endothelial cells (EC) and mediate down-regulation of endothelial cell proliferation and migration. Some data suggest that angiostatin inhibits vascularization by suppression of endothelial ATP metabolism regulating vascular physiology. Angiostatin blocks IF1 subunit binding at the same binding site on ATP synthase and abolishes its ability to conserve ATP. We demonstrated the distribution of IF1 subunit on OA knee joint and evaluate the regulation of the ectopic expression of IF1 by IL-1β and TNF-α. Methods: The identification of the IF1 subunit of ATP synthase was using flow cytometry and confocal microscopy using rabbit polyclonal antibodies on cultured human menisci cells. The distribution was detected by immunofluorescence using the same antibodies. Results: The fluorescence microscopy results showed a superficial cartilage and meniscus marker distribution of IF1 in control whereas the IF1 signal was localized on subchondral bone and hypertrofic chondrocytes in OA samples. Human menisci cells were cultivated after digestion with collagenase 2 and incubated with polyclonal antibody against IF1 subunit to show a plasma membrane location in control and different distribution on OA samples. Histograms of flow cytometry show changes on the IF1 membrane expression under TNF-α and IL-1β. There is low expression of ectopic IF-1 under IL-1β effect. Conclusions: We showed that the distribution of the IF1 subunit of ATP synthase on joint knee change under OA and this expression is regulated by IL-1β and TNF-α.