Peroxisome proliferator-activated receptor-gamma down-regulates chondrocyte matrix metalloproteinase-1 via a novel composite element.
Interleukin-1beta (IL-1beta) induces degradation via hyperexpression of an array of genes, including metalloproteinases (MMP), in cartilage cells during articular degenerative diseases. In contrast, natural ligands for peroxisome proliferator-activated receptors (PPARs) display protective anti-cytokine effects in these cells. We used the PPAR agonist rosiglitazone (Rtz) to investigate PPAR-gamma isotype on IL-1beta-target genes. Immunocytochemistry, electrophoretic mobility shift, and transient transfection assays revealed a functional PPAR-gamma in chondrocytes in vitro. Rtz displayed significant inhibition of IL-1beta effects in chondrocytes. Low Rtz concentrations (close to K(d) values for PPAR-gamma, 0.1 to 1 microm) inhibited the effects of IL-1beta on (35)S-sulfated proteoglycan production and gelatinolytic activities and downregulated MMP1 expression at mRNA and protein levels. We have investigated the mechanism of action of Rtz against IL-1beta-mediated MMP1 gene hyperexpression. Rtz effect occurs at the transcriptional level of the MMP1 promoter, as observed in transiently transfected cells with pMMP1-luciferase vector. Transient expression of wild type PPAR-gamma enhanced Rtz inhibitory effect in chondrocytes, whereas a mutated dominant negative PPAR-gamma abolished it, supporting the role of PPAR-gamma in this effect. MMP1 gene promoter analysis revealed the involvement of a cis-acting element located at -83 to -77, shown to be a composite PPRE/AP1 site. Gel mobility and supershift assays demonstrated that PPAR-gamma and c-Fos/c-Jun proteins bind this cis-acting element in a mutually exclusive way. Our data highlight a new PPAR-gamma-dependent inhibitory mechanism on IL-1beta-mediated cartilage degradation occurring through DNA binding competition on the composite PPRE/AP1 site in the MMP1 promoter.
François
Mathias
M
INSERM UMR-S-530, Université Paris 5, UFR Biomédicale, 45 Rue des Saints Pères, 75006 Paris, France.
Richette
Pascal
P
Tsagris
Lydia
L
Raymondjean
Michel
M
Fulchignoni-Lataud
Marie-Claude
MC
Forest
Claude
C
Savouret
Jean-François
JF
Corvol
Marie-Thérèse
MT
D000818:Animals; D001665:Binding Sites; D015152:Blotting, Northern; D015153:Blotting, Western; D002356:Cartilage / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D005343:Fibrinolytic Agents / Q000494:pharmacology; D005799:Genes, Dominant; D006801:Humans; D007150:Immunohistochemistry; D007375:Interleukin-1 / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D008156:Luciferases / Q000378:metabolism; D020781:Matrix Metalloproteinase 1 / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D016328:NF-kappa B / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011509:Proteoglycans / Q000378:metabolism; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000077154:Rosiglitazone; D013431:Sulfates / Q000378:metabolism; D045162:Thiazolidinediones / Q000494:pharmacology; D013997:Time Factors; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection
D018076:DNA, Complementary; D005343:Fibrinolytic Agents; D007375:Interleukin-1; D008024:Ligands; D016328:NF-kappa B; D011509:Proteoglycans; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D013431:Sulfates; D045162:Thiazolidinediones; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D000077154:Rosiglitazone; D012313:RNA; D008156:Luciferases; D020781:Matrix Metalloproteinase 1