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 alpha (TNF-alpha) 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 gamma (IFN-gamma), which weakly inhibited NO production, and transforming growth factor beta (TGF-beta), 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-alpha. Synovial autocrine factors (CAF) modestly induced NO synthesis, but inhibited synthesis by IL-1; TGF-beta was identified as an inhibitory component of CAF. Phorbol myristate acetate (PMA) had 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 NG-monomethyl-L-arginine (NMA) had no effect upon the increase in the production of prostaglandin E2 (PGE2), matrix metalloproteinases (MMP) or lactate by synoviocytes responding to IL-1. The rabbit synoviocyte cell line, HIG-82, did not synthesize detectable NO under any of the culture conditions tested.


CONCLUSIONS
Synoviocytes are a potential source of NO in arthritic joints.
Ferguson Laboratory, Department of Orthopaedic Surgery, University of Pittsburgh School of Medicine, PA.
M
M
Stefanovic-Racic
J
J
Stadler
H I
HI
Georgescu
C H
CH
Evans
D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology / Q000502:physiology*; D015232:Dinoprostone / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D005609:Free Radicals; D007371:Interferon-gamma / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D008666:Metalloendopeptidases / Q000378:metabolism; D009569:Nitric Oxide / Q000096:biosynthesis*; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011817:Rabbits; D019311:Staurosporine; D013583:Synovial Membrane / Q000166:cytology* / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D000470:Alkaloids; D016207:Cytokines; D005609:Free Radicals; D007375:Interleukin-1; D010982:Platelet-Derived Growth Factor; D016212:Transforming Growth Factor beta; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide; D007371:Interferon-gamma; D011493:Protein Kinase C; D008666:Metalloendopeptidases; D019311:Staurosporine; D015232:Dinoprostone; D013755:Tetradecanoylphorbol Acetate