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

Objectives To investigate whether CCAAT/enhancer binding protein beta (C/EBP beta) mediates the expression of matrix metalloproteinase-3 (MMP-3) and aggrecanases in arthritis. Methods Localisation of C/EBP beta 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 1 beta (IL-1 beta) were subjected to western blotting and quantitative PCR. Overexpression of C/EBP beta by adenovirus was performed in cells and organ culture of normal cartilage. Knockdown of C/EBP beta 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/EBP beta directly binds to the MMP-3 or ADAMTS-5 promoter, a chromatin immunoprecipitation assay was performed. Results Immunohistochemistry showed that C/EBP beta and MMP-3 were co-localised in arthritic synovium and cartilage. Western blots revealed increased C/EBP beta expression in cells treated with IL-1 beta. Expression of MMP-3, MMP-13 and ADAMTS-5 mRNA was significantly increased by the overexpression of C/EBP beta. C/EBP beta stimulated MMP-3 expression and induced matrix degradation in cartilage explants. C/EBP beta knockdown reduced MMP-3 and ADAMTS-5 expression. C/EBP beta 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/EBP beta core responsive element was located between -108 bp and -100 bp. The chromatin immunoprecipitation assay showed that C/EBP beta was directly bound to MMP-3 and ADAMTS-5 promoters. Conclusions These data demonstrate that C/EBP beta is involved in expression of MMP-3 and ADAMTS-5 in arthritic synovium and cartilage.