title: CCAAT/enhancer binding protein β regulates expression of matrix metalloproteinase-3 in arthritis.
abstract: OBJECTIVE
  To investigate whether CCAAT/enhancer binding protein β (C/EBPβ) mediates the expression of matrix metalloproteinase-3 (MMP-3) and aggrecanases in arthritis.
  
  
  METHODS
  Localisation of C/EBPβ 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 1β (IL-1β) were subjected to western blotting and quantitative PCR. Overexpression of C/EBPβ by adenovirus was performed in cells and organ culture of normal cartilage. Knockdown of C/EBPβ 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/EBPβ directly binds to the MMP-3 or ADAMTS-5 promoter,a chromatin immunoprecipitation assay was performed.
  
  
  RESULTS
  Immunohistochemistry showed that C/EBPβ and MMP-3 were co-localised in arthritic synovium and cartilage. Western blots revealed increased C/EBPβ expression in cells treated with IL-1β. Expression of MMP-3, MMP-13 and ADAMTS-5 mRNA was significantly increased by the overexpression of C/EBPβ. C/EBPβ stimulated MMP-3 expression and induced matrix degradation in cartilage explants. C/EBPβ knockdown reduced MMP-3 and ADAMTS-5 expression. C/EBPβ 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/EBPβ core responsive element was located between -108 bp and -100 bp. The chromatin immunoprecipitation assay showed that C/EBPβ was directly bound to MMP-3 and ADAMTS-5 promoters.
  
  
  CONCLUSIONS
  These data demonstrate that C/EBPβ is involved in expression of MMP-3 and ADAMTS-5 in arthritic synovium and cartilage.
authors.affiliation: Department of Orthopaedic Surgery, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Hidetoshi
authors.initials: H
authors.lastname: Tsushima
authors.forename: Ken
authors.initials: K
authors.lastname: Okazaki
authors.forename: Mitsumasa
authors.initials: M
authors.lastname: Hayashida
authors.forename: Takahiro
authors.initials: T
authors.lastname: Ushijima
authors.forename: Yukihide
authors.initials: Y
authors.lastname: Iwamoto
mesh_terms: D051722:ADAM Proteins / Q000378:metabolism; D000071121:ADAMTS4 Protein; D000071116:ADAMTS5 Protein; D000368:Aged; D000369:Aged, 80 and over; D001172:Arthritis, Rheumatoid / Q000378:metabolism*; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D002358:Cartilage, Articular / Q000201:enzymology / Q000378:metabolism; D002460:Cell Line; D005786:Gene Expression Regulation / Q000187:drug effects; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D055785:Gene Knockdown Techniques; D006801:Humans; D053583:Interleukin-1beta / Q000494:pharmacology; D053509:Matrix Metalloproteinase 13 / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D008875:Middle Aged; D020370:Osteoarthritis, Knee / Q000378:metabolism*; D011348:Procollagen N-Endopeptidase / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D013583:Synovial Membrane / Q000378:metabolism; D046509:Tissue Culture Techniques; D055815:Young Adult
chemical_list: D022782:CCAAT-Enhancer-Binding Protein-beta; D053583:Interleukin-1beta; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D051722:ADAM Proteins; D000071116:ADAMTS5 Protein; C121532:ADAMTS5 protein, human; D053509:Matrix Metalloproteinase 13; D011348:Procollagen N-Endopeptidase; D019278:Matrix Metalloproteinase 3; D000071121:ADAMTS4 Protein