title: Abrupt reoxygenation of microvascular endothelial cells after hypoxia activates ERK1/2 and JNK1, leading to NADPH oxidase-dependent oxidant production.
abstract: OBJECTIVE
  Mitogen-activated protein kinases (MAPK) in microvascular endothelial cells (EC) may participate in organ pathophysiology following hypoxia/reoxygenation (H/R). The authors aimed to determine the role of MAPK in H/R-induced reactive oxygen species (ROS) generation in mouse microvascular EC.
  
  
  METHODS
  Cultured EC derived from skeletal muscle of male wild-type (WT), gp91phox-/- or p47phox-/- mice were subjected to hypoxia (0.1% O2, 1 h) followed by abrupt reoxygenation, H/RA (hypoxic medium quickly replaced by normoxic medium), or slow reoxygenation, H/RS (O2 diffused to cells through hypoxic medium). Cells were analyzed for ERK, JNK, and p38 MAPK phosphorylation, NADPH oxidase activation, and ROS generation.
  
  
  RESULTS
  In WT cells, H/RA but not H/RS rapidly phosphorylated ERK1/2 and JNK1 and subsequently increased ROS production. H/RA did not affect p38. MAPK phosphorylation persisted despite inhibition of NADPH oxidase, mitochondrial respiration, protein tyrosine kinase, or PKC. ROS increase during H/RA was prevented by deletion of gp91phox or p47phox, or MAPK inhibition.
  
  
  CONCLUSIONS
  Abrupt reoxygenation after hypoxia activates ERK1/2 and JNK1 in mouse microvascular endothelial cells via a tyrosine kinase-, PKC-, and NADPH oxidase-insensitive mechanism, leading to increased NADPH oxidase-dependent ROS production. The results suggest that MAPK activation in the microvascular endothelium is O2-sensitive, contributing critically to tissue pathophysiology after H/R.
authors: The Centre for Critical Illness Research, Lawson Health Research Institute, University of Western Ontario, London, Canada.
authors: Guo
authors: G
authors: Yu
authors: Tianqing
authors: T
authors: Peng
authors: Qingping
authors: Q
authors: Feng
authors: Karel
authors: K
authors: Tyml
mesh_terms: D000818:Animals; D042783:Endothelial Cells / Q000378:metabolism*; D000860:Hypoxia / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology*; D008297:Male; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008817:Mice, Mutant Strains; D008833:Microcirculation / Q000502:physiology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D048055:Mitogen-Activated Protein Kinase 8 / Q000378:metabolism; D018482:Muscle, Skeletal / Q000098:blood supply / Q000378:metabolism; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000378:metabolism; D018384:Oxidative Stress / Q000502:physiology*; D010100:Oxygen / Q000378:metabolism; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D008562:Membrane Glycoproteins; D047428:Protein Kinase Inhibitors; D017382:Reactive Oxygen Species; C488842:Cybb protein, mouse; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D048055:Mitogen-Activated Protein Kinase 8; D010100:Oxygen