Nitric oxide synthesis and its regulation by rabbit synoviocytes

Objective: To determine whether rabbit synovial fibroblasts can synthesize nitric oxide (NO) and, if so, how production is regulated by cytokines. Methods. Primary cultures of synovial fibroblasts (type B synoviocytes) were established from synovia excised from the knee joints of New Zealand white rabbits. Synthesis of NO was measured as nitrite accumulation in conditioned media in the presence or absence of various cytokines and other activators. Results: Resting cultures of synoviocytes normally produced little or no NO: However, production of this free radical was induced by interleukin 1 (IL-1), tumor necrosis factor α (TNF-α or the phagocytosis of latex beads; in some cultures, the synthesis of NO occurred spontaneously. In each case, NO synthesis began approximately 9 h after the addition of cytokines, suggesting the involvement of an inducible form of NO synthase. Antagonists of this phenomenon included interferon feron γ (IFN-γ), which weakly inhibited NO production, and transforming growth factor β (TGF-β), a very strong inhibitor. Platelet derived growth,factor (PDGF) inhibited NO synthesis by cells stimulated with IL-1, but not by cells stimulated with TNF-α. Synovial autocrine factors (CAF) modestly induced NO synthesis, but inhibited synthesis by IL-1; TGF-β was identified as an inhibitory component of CAF. Phorbol myristate acetate (PMA) dad only a small inductive effect,:and inhibited induction by IL-1. However, the protein kinase inhibitor staurosporin Was a strong inducer. Modulators of cyclic nucleotides, in contrast, had relatively modest effects on NO synthesis. Inhibition of NO biosynthesis by N(G)-monomethyl-L-arginine (NMA) had no effect upon the increase in the production of prostaglandin E, (PGE₂), matrix metalloproteinases (MMP) or lactate by synoviocytes responding to IL-1. The rabbit synaviocyte cell line, HIG-82, did not synthesize detectable NO under any of the culture conditions tested. Conclusion: Synoviocytes area potential source of NO in arthritic joints.