Activation of the peroxisome proliferator-activated receptor alpha pathway potentiates interleukin-1 receptor antagonist production in cytokine-treated chondrocytes.
OBJECTIVE
To determine whether peroxisome proliferator-activated receptor alpha (PPARalpha) agonists protect chondrocytes against the effects of interleukin-1beta (IL-1beta).


METHODS
PPARalpha 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 (Il-1Ra) gene promoter. Chondrocytes were incubated in vitro with IL-1beta alone or in combination with CloFibrate (CloF) or other PPAR ligands. Proteoglycans were evaluated by 35S-sulfate incorporation, matrix metalloproteinase (MMP) levels were assessed by zymography and enzyme-linked immunosorbent assay (ELISA), and MMP messenger RNA (mRNA) levels were measured by Northern blotting and real-time reverse transcriptase-polymerase chain reaction. IL-1beta and IL-1Ra soluble contents were measured by ELISA.


RESULTS
CloF counteracted IL-1beta-induced 35S-proteoglycan degradation, gelatinolytic activity, and MMP-1, -3, and -13 mRNA expression. CloF also maximized IL-1beta-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-1beta-induced MMP expression. The stimulatory effect of CloF was enhanced by cotransfection with wild-type PPARalpha and abolished by a dominant-negative PPARalpha 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-kappaB-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.


CONCLUSIONS
Our findings support the notion that there is a PPARalpha-dependent mechanism that inhibits IL-1beta function in chondrocytes, which operates via an increase in sIL-1Ra production.
François
Mathias
M
INSERM UMR-S-530-747, University Paris Descartes, Paris, France.
Richette
Pascal
P
Tsagris
Lydia
L
Fitting
Catherine
C
Lemay
Cedric
C
Benallaoua
Mourad
M
Tahiri
Khadija
K
Corvol
Marie-Therese
MT
D000818:Animals; D064236:Cartilage Oligomeric Matrix Protein; D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D002994:Clofibrate / Q000494:pharmacology; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism; D053590:Interleukin 1 Receptor Antagonist Protein; D007375:Interleukin-1 / Q000037:antagonists & inhibitors / Q000502:physiology; D064235:Matrilin Proteins; D047493:PPAR alpha / Q000502:physiology*; D011817:Rabbits; D012795:Sialoglycoproteins / Q000096:biosynthesis*
D064236:Cartilage Oligomeric Matrix Protein; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; C507763:IL1RN protein, human; D053590:Interleukin 1 Receptor Antagonist Protein; D007375:Interleukin-1; D064235:Matrilin Proteins; D047493:PPAR alpha; D012795:Sialoglycoproteins; C577553:TSP5 protein, human; D002994:Clofibrate