title: Upregulated hypoxia-inducible factor-1 DNA binding activity to the vascular endothelial growth factor-A promoter mediates increased vascular permeability in donor lung grafts.
abstract: BACKGROUND
  Transplantation-induced hypoxia results in enhanced vascular permeability and tissue vascular endothelial growth factor (VEGF) and endothelin-1 (ET-1) overexpression in donor lung grafts. Promoter studies have uncovered a hypoxia-inducible factor (HIF)-1 binding site (HBS) in 5'-flanking region of VEGF gene that regulates the hypoxia-induced expression of VEGF; and ET-1 potently stimulates VEGF-A production. We hypothesized that HIF-1 regulates VEGF-mediated vascular permeability in lung grafts.
  
  
  METHODS
  We studied the mRNA and protein expression of HIF-1 and its protein-binding capacity to the HBS of the VEGF gene in biopsies of preserved donor and control lungs, using real-time reverse transcription-polymerase chain reaction, Western blotting, and electrophoretic mobility shift assay. Wet-to-dry lung weight ratio was measured in donor and control lungs.
  
  
  RESULTS
  While HIF-1 alpha mRNA expression was unchanged, HIF-1 beta was downregulated (p < 0.05) in donor versus control lungs. Protein expression of both, HIF-1 alpha and -beta was significantly upregulated in donor lung grafts. HIF-1 binding to the HBS of the VEGF promoter as well as tissue fluid content were increased in donor lung biopsies versus controls (p < 0.05).
  
  
  CONCLUSIONS
  These data indicate that upregulated HIF-1 DNA binding activity to the HBS of VEGF-A most likely contributes to elevated VEGF levels in preserved lung grafts. Unchanged HIF-1 alpha mRNA expression did not affect HIF-1 alpha protein levels. Endothelin-1 increases HIF-1 alpha accumulation and activates HIF-1 transcription complex in vitro. Therefore, ET-1-mediated increased HIF-1 alpha protein stability most likely leads to transcriptional activation of VEGF during lung graft preservation. Targeting HIF might be of benefit to counteract edema formation in preserved lung grafts.
authors.affiliation: Laboratory for Cardiovascular Research, Department of Anatomy, University of Vienna, Vienna, Austria.
authors.forename: Dietmar
authors.initials: D
authors.lastname: Abraham
authors.forename: Katharina
authors.initials: K
authors.lastname: Krenn
authors.forename: Gernot
authors.initials: G
authors.lastname: Seebacher
authors.forename: Patrick
authors.initials: P
authors.lastname: Paulus
authors.forename: Walter
authors.initials: W
authors.lastname: Klepetko
authors.forename: Seyedhossein
authors.initials: S
authors.lastname: Aharinejad
mesh_terms: D051784:Aryl Hydrocarbon Receptor Nuclear Translocator; D001665:Binding Sites; D015153:Blotting, Western; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology*; D024202:Electrophoretic Mobility Shift Assay; D019332:Endothelin-1 / Q000502:physiology; D006801:Humans; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D016040:Lung Transplantation / Q000502:physiology*; D009687:Nuclear Proteins / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000032:analysis; D018336:Receptors, Aryl Hydrocarbon / Q000235:genetics / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology; D015854:Up-Regulation; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000502:physiology*
chemical_list: C497396:ARNT protein, human; D004268:DNA-Binding Proteins; D019332:Endothelin-1; C497442:HIF1A protein, human; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009687:Nuclear Proteins; D012333:RNA, Messenger; D018336:Receptors, Aryl Hydrocarbon; D014157:Transcription Factors; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D051784:Aryl Hydrocarbon Receptor Nuclear Translocator