title: Regulation of autophagy in human and murine cartilage: hypoxia-inducible factor 2 suppresses chondrocyte autophagy.
abstract: OBJECTIVE
  We have previously demonstrated that the transcription factor hypoxia-inducible factor 1 (HIF-1) promotes the onset of autophagy in chondrocytes. The overall goal of this study was to test the hypothesis that another HIF family transcription factor, HIF-2, modulates the induction of autophagy by chondrocytes.
  
  
  METHODS
  Expression of HIF-1, HIF-2, and light chain 3 (LC3) in human and murine articular cartilage was visualized by immunohistochemistry. Suppression of HIF-2 was achieved using small interfering RNA technology. Assessments of autophagic flux and lysosomal activity, as well as ultrastructural analysis, were performed in chondrocytes in cell culture.
  
  
  RESULTS
  HIF-2 was expressed abundantly by cells in human and murine articular cartilage and in the cartilage of mineralizing vertebrae from neonatal mice. Protein levels were reduced in articular cartilage from older mice, in end-plate cartilage from mice, and in chondrocytes from human osteoarthritic (OA) cartilage. HIF-2 was robustly expressed in the prehypertrophic cells of mouse growth cartilage. When HIF-2alpha was silenced, the generation of reactive oxygen species was found to be elevated, with a concomitant decrease in catalase and superoxide dismutase activity. Suppression of HIF-2 was associated with decreased Akt-1 and mammalian target of rapamycin activities, reduced Bcl-xL expression, and a robust autophagic response, even under nutrient-replete conditions. In these silenced chondrocytes, HIF-1 expression was elevated. Decreased HIF-2 expression was associated with autophagy in OA tissues and aging cartilage samples. The autophagic response of chondrocytes in HIF-2alpha-knockout mouse growth plate showed an elevated autophagic response throughout the plate.
  
  
  CONCLUSIONS
  Based on these observations, we conclude that HIF-2 is a potent regulator of autophagy in maturing chondrocytes. Our data suggest that this protein acts as a brake on the autophagy-accelerator function of HIF-1.
authors.affiliation: Department of Orthopaedic Surgery, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors.forename: Jolene
authors.initials: J
authors.lastname: Bohensky
authors.forename: Shawn P
authors.initials: SP
authors.lastname: Terkhorn
authors.forename: Theresa A
authors.initials: TA
authors.lastname: Freeman
authors.forename: Christopher S
authors.initials: CS
authors.lastname: Adams
authors.forename: Joseph A
authors.initials: JA
authors.lastname: Garcia
authors.forename: Irving M
authors.initials: IM
authors.lastname: Shapiro
authors.forename: Vickram
authors.initials: V
authors.lastname: Srinivas
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D001343:Autophagy / Q000502:physiology*; D051792:Basic Helix-Loop-Helix Proteins / Q000032:analysis / Q000037:antagonists & inhibitors / Q000502:physiology*; D002374:Catalase / Q000032:analysis; D002478:Cells, Cultured; D019902:Chondrocytes / Q000502:physiology* / Q000648:ultrastructure; D006801:Humans; D051793:Hypoxia-Inducible Factor 1 / Q000032:analysis / Q000502:physiology; D007150:Immunohistochemistry; D051379:Mice; D018345:Mice, Knockout; D008869:Microtubule-Associated Proteins / Q000032:analysis; D013482:Superoxide Dismutase / Q000032:analysis; D000099162:Endothelial PAS Domain-Containing Protein 1
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D002374:Catalase; D051793:Hypoxia-Inducible Factor 1; D008869:Microtubule-Associated Proteins; D013482:Superoxide Dismutase; D000099162:Endothelial PAS Domain-Containing Protein 1; C477012:MAP1LC3A protein, human