title: Prothrombin gene expression in articular cartilage with a putative role in cartilage degeneration secondary to joint immobility.
abstract: OBJECTIVE
  To test the hypothesis that thrombin is expressed by chondrocytes from human and animal articular cartilage and to monitor its levels of expression during cartilage degeneration induced by joint immobility in a rat model.
  
  
  METHODS
  Rat knees were immobilized for periods of 2 or 4 weeks, after which the articular cartilage was harvested, total RNA extracted, and the differential display (ddPCR) protocol applied to identify differentially expressed genes. One differentially expressed fragment showed 100% homology with the prothrombin gene. Results were verified by RT-PCR, Northern and Western blot analysis, and immunohistochemistry in human, rat, and rabbit articular cartilage.
  
  
  RESULTS
  In our rat model of cartilage degeneration induced by joint immobilization, increases in the levels of prothrombin mRNA, thrombin protein, and fibrin deposition were observed. Expression of the prothrombin gene by chondrocytes was confirmed by ddPCR (rat), RT-PCR (rat and human), and by Northern blot analysis (rabbit). In addition, thrombin-like immunoreactivity was increased in chondrocytes after a 4 week immobilization period compared with rat knees receiving sham surgery. Thrombin activity was reflected by the presence of fibrin immunoreactivity in operated rat knee joints.
  
  
  CONCLUSIONS
  Articular chondrocytes express the prothrombin gene and its local expression in joints is translated into thrombin protein. Prothrombin expression is increased in response to joint immobility. Our results support generation of thrombin locally in joints and an upregulation of thrombin expression in cartilage degeneration secondary to immobility. These results may provide information on the source of increased thrombin activity in various animal models and in clinical forms of arthritis.
authors.affiliation: Bone and Joint Research Laboratory, Department of Medicine, University of Ottawa, Ontario, Canada.
authors.forename: Guy
authors.initials: G
authors.lastname: Trudel
authors.forename: Hans K
authors.initials: HK
authors.lastname: Uhthoff
authors.forename: Odette
authors.initials: O
authors.lastname: Laneuville
mesh_terms: D000818:Animals; D002358:Cartilage, Articular / Q000473:pathology* / Q000502:physiology*; D019902:Chondrocytes / Q000473:pathology / Q000502:physiology; D003286:Contracture / Q000473:pathology* / Q000503:physiopathology*; D003600:Cytosol / Q000378:metabolism; D004195:Disease Models, Animal; D005109:Extracellular Matrix / Q000378:metabolism; D015870:Gene Expression; D007103:Immobilization / Q000009:adverse effects; D008297:Male; D011516:Prothrombin / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D012333:RNA, Messenger; D011516:Prothrombin