Abstract
Objectives Interferon gamma (IFNγ) is found to be elevated in the synovial fluid of patients with rheumatoid arthritis and osteoarthritis, suggesting its implication in joint disease pathogenesis. In this study, we investigated the effects of IFNγ on the production of cytokines (IL-6, IL-8, IL-10), prostaglandin E2(PGE2), proteoglycans (PG), nitric oxide (NO), interleukin-1 receptor antagonist (IL-1ra) and stromelysin by non-stimulated and IL-1β-treated human chondrocytes. The role played by NO in the responses of chondrocytes to IFNγ was also examined by incubation of chondrocytes with NG-monomethyl-L-arginine (L-NMMA), a competitive inhibitor of NO synthase.
Methods Enzymatically isolated human chondrocytes were cultured for 48h in the absence or presence of IL-1β, IFNγ or NG-monomethyl-L-arginine (L-NMMA) added solely or in combination. The productions of IL-6, IL-8, IL-10, IL-1ra and stromelysin were measured by enzyme amplified sensitivity immunoassays (EASIA). PG and PGE2were quantified by specific radioimmunoassays (RIA). Nitrite concentrations in the culture supernatants were determined by a spectrophotometric method based upon the Griess reaction.
Results As expected, IL-1β highly stimulated NO, IL-6, IL-8, IL-10, IL-1ra, PGE2and stromelysin synthesis, but dramatically decreased PG production. NO, IL-6, IL-1ra and PGE2production by non-stimulated chondrocytes was dose-dependently increased by IFNγ while PG production was inhibited. In the absence of IL-1β, IL-10 was undetectable in the culture supernatants. At the doses of 10 and 100U/ml, IFNγ markedly inhibited the constitutive and IL-1β-stimulated IL-8, IL-10 and stromelysin productions. Interestingly, IFNγ synergized with IL-1β to increase NO, IL-6, IL-1ra and to depress PG production. As previously reported, the inhibition of NO synthesis by the competitive inhibitor L-NMMA led to enhancement of IL-6, IL-8 and PGE2production by IL-1β treated chondrocytes, but did not significantly modify IL-10, PG and MMP-3 productions. Inhibition of NO synthase significantly inhibited the stimulating effect of IFNγ on IL-6 and IL-1ra but did not affect the inhibitory effect of IFNγ on IL-8, PG or stromelysin production.
ConclusionsThese findings suggest that IFNγ and IL-1 synergistically stimulate the production of IL-6, IL-1ra, NO and PGE2and inhibit PG synthesis. By contrast, IL-1β and IFNγ have opposite effects on IL-8, IL-10 and stromelysin productions. These effects are not reversed by L-NMMA, suggesting that NO is not the principal mediator involved in responses of chondrocytes to IFNγ.