Activation of the peroxisome proliferator-activated receptor a pathway potentiates interleukin-1 receptor antagonist production in cytokine-treated chondrocytes

Objective. To determine whether peroxisome proliferator-activated receptor alpha (PPAR alpha) agonists protect chondrocytes against the effects of interleukin-1 beta (IL-1 beta). Methods. PPAR alpha expression and function in cultured rabbit articular chondrocytes were studied by Northern blotting, electrophoretic mobility shift assay, and transient expression of a luciferase reporter construct bearing the human IL-1 receptor antagonist (II-1Ra) gene promoter. Chondrocytes were incubated in vitro with IL-1 beta alone or in combination with CloFibrate (CloF) or other PPAR ligands. Proteoglycans were evaluated by S-35-sulfate incorporation, matrix metalloproteinase (NIMP) levels were assessed by zymography and enzyme-linked immunosorbent assay (ELISA), and VIMP messenger RNA (mRNA) levels were measured by Northern blotting and real-time reverse transcriptase-polymerase chain reaction. IL-1 beta and IL-1Ra soluble contents were measured by ELISA. Results. CloF counteracted IL-1 beta-induced S-35-proteoglycan degradation, gelatinolytic activity, and MMP-1, -3, and -13 mRNA expression. CloF also maximized IL-1 beta-induced endogenous production of soluble IL-1Ra (sIL-1Ra). This stimulating effect on IL-1Ra gene expression was shown, by transient expression assay, to be transcriptional. Inhibition of sIL-1Ra expression by a specific small interfering RNA suppressed the effect of CloF on IL-1 beta-induced NIMP expression. The stimulatory effect of CloF was enhanced by cotransfection with wild-type PPAR alpha and abolished by a dominant-negative PPAR alpha mutant. Fenofibrate and WY-14643 displayed a similar stimulating effect on the IL-1Ra promoter, while rosiglitazone did not. Two PPAR response elements, an NF-kappa B-binding site, and a CCAAT/enhancer binding protein-binding site were identified in the IL-1Ra promoter. All 4 sites were necessary for mediation of the effects of CloF. Conclusion. Our findings support the notion that there is a PPAR alpha-dependent mechanism that inhibits IL-1 beta function in chondrocytes, which operates via an increase in sIL-1Ra production.