title: Modulation of collagenase 3 in human osteoarthritic cartilage by activation of extracellular transforming growth factor beta: role of furin convertase.
abstract: OBJECTIVE
  Treatment of normal cartilage with transforming growth factor beta (TGFbeta) can increase the synthesis of collagenase 3 by chondrocytes and mimic the in situ distribution of this enzyme in osteoarthritic (OA) cartilage, which occurs predominantly in the deep zone. In this study, we examined the elements of the TGFbeta system that are potentially relevant to this effect.
  
  
  METHODS
  TGFbeta1 and TGFbeta2 levels in cultured cartilage explants were determined by enzyme-linked immunosorbent assay (ELISA). OA cartilage explants were treated with small latent TGFbeta1 complex in the presence of various inhibitors, and collagenase 3 levels were determined by ELISA. The inhibitors were against serine proteases, plasmin, cathepsins, furin, and a neutralizing antibody against the mannose-6 phosphate/ insulin-like growth factor 2 receptor (M6P/IGF-2R). Small latent TGFbeta1, TGFbeta receptor types I, II, and III (TGFbetaRI, RII, and RIII), M6P/IGF-2R, and furin were immunolocalized in cartilage.
  
  
  RESULTS
  Our data showed that latent TGFbeta1 is the major isoform that is synthesized; levels of 17.2 +/-1.7 pg/mg and 1.1 +/- 0.3 pg/mg tissue wet weight (mean +/- SEM) were found for total TGFbeta1 and TGFbeta2, respectively, in OA cartilage. A general serine protease inhibitor abrogated activation of both endogenous and exogenous small latent TGFbeta1. Plasmin and furin inhibitors and anti-M6P/IGF-2R reduced the levels of exogenous small latent TGFbeta1 complex-induced collagenase 3 by 33%, 95%, and 76%, respectively, but the cathepsin inhibitor had no effect. Immunolocalization of the small latent TGFbeta1 complex as well as of TGFbetaRI and RII revealed a statistically significant increase in the chondrocyte score in only the deep zone of OA cartilage. The M6P/IGF-2R level was significantly higher in OA cartilage in both the superficial and deep zones. Furin was found in normal cartilage exclusively in the superficial zone, whereas in OA cartilage, a level similar to that in normal cartilage was found in the superficial zone, but a significantly higher cell score (mean +/- SEM 23.6 +/- 4.7%) was registered in the deep zone.
  
  
  CONCLUSIONS
  The mechanisms of TGFbeta activation/ activity with regard to collagenase 3 modulation in cartilage appear to be controlled by furin convertase with or without M6P/IGF-2R. These factors and the small latent TGFbeta complex are increased in the deep zone of OA cartilage, corresponding to the preferential site of collagenase 3 production.
authors.affiliation: Centre hospitalier de l'Université de Montréal, Hôpital Notre-Dame, Quebec, Canada.
authors.forename: F
authors.initials: F
authors.lastname: Moldovan
authors.forename: J P
authors.initials: JP
authors.lastname: Pelletier
authors.forename: F
authors.initials: F
authors.lastname: Mineau
authors.forename: M
authors.initials: M
authors.lastname: Dupuis
authors.forename: J M
authors.initials: JM
authors.lastname: Cloutier
authors.forename: J
authors.initials: J
authors.lastname: Martel-Pelletier
mesh_terms: D000368:Aged; D002358:Cartilage, Articular / Q000737:chemistry*; D017364:Collagenases / Q000187:drug effects* / Q000378:metabolism; D005260:Female; D045683:Furin; D006801:Humans; D008297:Male; D053509:Matrix Metalloproteinase 13; D020370:Osteoarthritis, Knee / Q000378:metabolism*; D013381:Subtilisins / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000378:metabolism*
chemical_list: D016212:Transforming Growth Factor beta; D013381:Subtilisins; D045683:Furin; D017364:Collagenases; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13