title: Downregulation of inhibitor of apoptosis proteins in apoptotic human chondrocytes treated with tumor necrosis factor-alpha and actinomycin D.
abstract: OBJECTIVE
  Apoptosis of chondrocytes plays a pivotal role in cartilage degeneration. Tumor necrosis factor-alpha (TNF-alpha) is a proinflammatory cytokine and has been assumed to cause the degradation of human cartilage. To investigate the mechanisms of TNF-alpha-mediated apoptosis of human chondrocytes from a point of view of the balance between the caspase-cascade and the expression of inhibitor of apoptosis proteins (IAPs), although both of them are induced with TNF-signals.
  
  
  METHODS
  The expression of TNF-receptors (TNF-Rs) in normal human articular chondrocyte (NHAC-kn) was examined with immunocytochemistry. Subconfluent cultures of NHAC-kn were tested with TNF-alpha and/or actinomycin D (actD), and the induction of apoptosis was evaluated by the frequency of apoptotic cells visualized with nuclear staining using Hoechst 33342. The activation of caspases and the expression of IAPs were examined with Western blot analyses.
  
  
  RESULTS
  NHAC-kn expressed TNF-R1 and -R2. When NHAC-kn was treated with TNF-alpha (10 ng/ml) and actD (0.2 microg/ml) for 24 h, the frequency of apoptotic cells increased to more than 25%. TNF-alpha alone, however, induced the apoptosis insufficiently (up to 8.3%), even when used at the concentration of 100 ng/ml for 48 h. In apoptotic human chondrocytes induced with TNF-alpha (10 ng/ml) and actD (0.2 microg/ml), the caspase-3, -8, and -9 were activated and the protein expression of XIAP and c-IAP1 decreased.
  
  
  CONCLUSIONS
  In apoptotic human chondrocytes induced with TNF-alpha and actD, the balance between caspase activation and IAPs' expression lay with the executioner caspase (caspase-3) and led to decreased expression of XIAP and c-IAP1.
authors.affiliation: Department of Pathology, Iwate Medical University School of Medicine, 19-1 Uchimaru, Morioka 020-8505, Japan.
authors.forename: F
authors.initials: F
authors.lastname: Yoshimura
authors.forename: H
authors.initials: H
authors.lastname: Kanno
authors.forename: M
authors.initials: M
authors.lastname: Uzuki
authors.forename: K
authors.initials: K
authors.lastname: Tajima
authors.forename: T
authors.initials: T
authors.lastname: Shimamura
authors.forename: T
authors.initials: T
authors.lastname: Sawai
mesh_terms: D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western / Q000379:methods; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects*; D003609:Dactinomycin / Q000494:pharmacology*; D015536:Down-Regulation / Q000502:physiology; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D018124:Receptors, Tumor Necrosis Factor / Q000032:analysis; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D011500:Protein Synthesis Inhibitors; D018124:Receptors, Tumor Necrosis Factor; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D003609:Dactinomycin; D020169:Caspases