CCAAT/enhancer binding protein beta regulates expression of matrix metalloproteinase-3 in arthritis.

摘要

OBJECTIVES: To investigate whether CCAAT/enhancer binding protein beta (C/EBPbeta) mediates the expression of matrix metalloproteinase-3 (MMP-3) and aggrecanases in arthritis. METHODS: Localisation of C/EBPbeta and MMP-3 in synovium and cartilage from patients with rheumatoid arthritis and osteoarthritis was determined by immunohistochemistry. Cell lines SW982, C28/I2 and human fibroblast-like synoviocytes stimulated by interleukin 1beta (IL-1beta) were subjected to western blotting and quantitative PCR. Overexpression of C/EBPbeta by adenovirus was performed in cells and organ culture of normal cartilage. Knockdown of C/EBPbeta by small interference RNA was performed in cells. Activity of the human MMP-3 and aggrecanase-2 ADAMTS-5 (a disintegrin and metalloproteinase with thrombospondin motifs) promoters was analysed by a luciferase assay. To determine whether C/EBPbeta directly binds to the MMP-3 or ADAMTS-5 promoter,a chromatin immunoprecipitation assay was performed. RESULTS: Immunohistochemistry showed that C/EBPbeta and MMP-3 were co-localised in arthritic synovium and cartilage. Western blots revealed increased C/EBPbeta expression in cells treated with IL-1beta. Expression of MMP-3, MMP-13 and ADAMTS-5 mRNA was significantly increased by the overexpression of C/EBPbeta. C/EBPbeta stimulated MMP-3 expression and induced matrix degradation in cartilage explants. C/EBPbeta knockdown reduced MMP-3 and ADAMTS-5 expression. C/EBPbeta stimulated the 2011 bp MMP-3 promoter and the 1768 bp ADAMTS-5 promoter in a dose-dependent manner. Deletion and mutation analysis of the MMP-3 promoter showed that the C/EBPbeta core responsive element was located between -108 bp and -100 bp. The chromatin immunoprecipitation assay showed that C/EBPbeta was directly bound to MMP-3 and ADAMTS-5 promoters. CONCLUSIONS: These data demonstrate that C/EBPbeta is involved in expression of MMP-3 and ADAMTS-5 in arthritic synovium and cartilage.