title: CCN1 (Cyr61) Is Overexpressed in Human Osteoarthritic Cartilage and Inhibits ADAMTS-4 (Aggrecanase 1) Activity.
abstract: OBJECTIVE
  ADAMTS-4, also called aggrecanase 1, is considered to play a key role in aggrecan degradation in human osteoarthritic (OA) cartilage, but information about regulators of ADAMTS-4 aggrecanase activity remains limited. We undertook this study to search for molecules that modulate ADAMTS-4 activity.
  
  
  METHODS
  Molecules copurified with ADAMTS-4 from ADAMTS-4-transfected chondrocytic cells were sequenced by nanoscale liquid chromatography tandem mass spectrometry. Binding activity was determined by immunoprecipitation and solid-phase binding assay. Effects on ADAMTS-4 activity were examined by aggrecan digestion assay. Expression of the binding molecule in OA cartilage and chondrocytes was examined by immunohistochemistry and reverse transcription-polymerase chain reaction.
  
  
  RESULTS
  We identified CCN1 (Cyr61) as an ADAMTS-4-binding protein and showed specific binding to the ADAMTS-4 cysteine-rich domain. Aggrecanase activity of ADAMTS-4 was inhibited by interaction with CCN1. Expression of messenger RNA for CCN1 was significantly higher in human OA cartilage than in normal cartilage. CCN1 was immunolocalized to chondrocytes in OA cartilage, showing direct correlations of immunoreactivity with the Mankin score of cartilage lesions and chondrocyte cloning. CCN1 and ADAMTS-4 were commonly coexpressed in clustered chondrocytes. CCN1 expression in OA chondrocytes was down-regulated by interleukin-1α (IL-1α) and up-regulated by transforming growth factor β (TGFβ). ADAMTS-4 expression was induced by treatment with IL-1α or TGFβ, but aggrecanase activity was detected only under stimulation with IL-1α. TGFβ-treated chondrocytes exhibited aggrecanase activity when CCN1 expression was knocked down.
  
  
  CONCLUSIONS
  Our findings provide the first evidence that CCN1 suppresses ADAMTS-4 activity and that CCN1 overexpression is directly correlated with chondrocyte cloning in OA cartilage. Our results suggest that the TGFβ/CCN1 axis plays a role in chondrocyte cluster formation through inhibition of ADAMTS-4.
authors.affiliation: Keio University School of Medicine, Tokyo, Japan.
authors.forename: Miyuki
authors.initials: M
authors.lastname: Chijiiwa
authors.affiliation: Keio University School of Medicine, Tokyo, Japan.
authors.forename: Satsuki
authors.initials: S
authors.lastname: Mochizuki
authors.affiliation: Yamaguchi University Graduate School of Medicine, Ube, Japan.
authors.forename: Tokuhiro
authors.initials: T
authors.lastname: Kimura
authors.affiliation: Keio University School of Medicine, Tokyo, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Abe
authors.affiliation: Fukui University School of Medicine, Fukui, Japan.
authors.forename: Yukie
authors.initials: Y
authors.lastname: Tanaka
authors.affiliation: Fukui University School of Medicine, Fukui, Japan.
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Fujii
authors.affiliation: Keio University School of Medicine, Tokyo, Japan.
authors.forename: Hidenori
authors.initials: H
authors.lastname: Shimizu
authors.affiliation: Keio University School of Medicine, Tokyo, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Enomoto
authors.affiliation: Keio University School of Medicine, Tokyo, Japan.
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Toyama
authors.affiliation: Keio University School of Medicine, Tokyo, Japan.
authors.forename: Yasunori
authors.initials: Y
authors.lastname: Okada
mesh_terms: D051722:ADAM Proteins / Q000378:metabolism*; D000071121:ADAMTS4 Protein; D000368:Aged; D000369:Aged, 80 and over; D002358:Cartilage, Articular / Q000378:metabolism*; D016022:Case-Control Studies; D019902:Chondrocytes / Q000201:enzymology / Q000378:metabolism*; D055515:Cysteine-Rich Protein 61 / Q000235:genetics* / Q000378:metabolism; D020869:Gene Expression Profiling; D006801:Humans; D047468:Immunoprecipitation; D008875:Middle Aged; D010003:Osteoarthritis / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D011348:Procollagen N-Endopeptidase / Q000378:metabolism*; D011485:Protein Binding; D012333:RNA, Messenger / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016212:Transforming Growth Factor beta / Q000378:metabolism
chemical_list: C501848:CCN1 protein, human; D055515:Cysteine-Rich Protein 61; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D051722:ADAM Proteins; D011348:Procollagen N-Endopeptidase; D000071121:ADAMTS4 Protein; C000603918:ADAMTS4 protein, human