Regulation of the nitric oxide production resulting from the glucocorticoid-insensitive expression of iNOS in human osteoarthritic cartilage

Objective: Nitric oxide (NO) produced by cartilage and synovial membranes is implicated in the pathogenesis of osteoarthritis (OA) and inhibitors of NO synthesis may have indications in the treatment or prevention of joint destruction in OA. Because the signaling mechanisms as well as the NOS isoform involved in induction of NO production in human cartilage remain in many parts unclear, the present study was designed to investigate the regulation of inducible NO synthesis in human intact CA cartilage. Methods: Cartilage specimens were collected from CA patients undergoing knee replacement surgery and studied for iNOS expression and NO production in organ culture to allow intact chondrocyte-matrix interactions. J774 macrophages were used for comparison as a well-documented source of iNOS. Results: CA cartilage expressed iNOS and produced NO in the absence of exogenous cytokines. Addition of interleukin-1 beta (IL-1 beta), tumor necrosis factor alpha (TNF alpha) or lipopolysaccharide (LPS) into the culture medium enhanced NO production in a dose-and time-dependent manner. Various NOS inhibitors suppressed NO production in the following order of potency: 1400W (novel selective iNOS inhibitor)= L-NIO >L-NMMA >L-NAME. Cycloheximide (an inhibitor of protein synthesis), pyrrolidine dithiocarbamate (PDTC; an NF-kappaB inhibitor) and genistein (an inhibitor of tyrosine protein kinases) inhibited cytokine-induced NO production, while dexamethasone, diaminohydroxypyrimidine (DAHP; an inhibitor of tetrahydrobiopterin synthesis) and PID 98059 (p42/44 MAP kinase inhibitor) had no effect. Conclusions: The results suggest that NO synthesis in human osteoarthritic cartilage derives from the glucocorticoid-insensitive expression of iNOS. Very similar mechanisms appear to regulate inducible NO synthesis in human ostecarthritic cartilage and J774 macrophages with the exception that dexamethasone inhibited NO production in J774 cells but not in osteoarthritic cartilage. (C) 2001 OsteoArthritis Research Society International.