title: Transient and reversible deoxyribonucleic acid damage in human left ventricle under controlled ischemia and reperfusion.
abstract: OBJECTIVE
  We sought to describe the sequence of molecular events during ischemia and reperfusion of the human heart and to determine the activation of stress kinases and deoxyribonucleic acid (DNA) damage response elements on apoptosis in ischemia or reperfusion of the human heart.
  
  
  BACKGROUND
  Brief ischemia is tolerated by cardiac myocytes, but it determines immediate metabolic changes and block of contraction. Prompt restoration of coronary blood flow is inexorably associated with a slow recovery of myocardial contractile function. The prolonged, postischemic contractile dysfunction in the viable tissue is called myocardial stunning. The molecular mechanisms underlying myocardial stunning and ischemia-reperfusion injury are still poorly understood. Their elucidation would be valuable in order to identify novel therapeutic strategies.
  
  
  METHODS
  We examined human left ventricular samples taken from 20 patients undergoing elective valve surgery before aortic cross-clamping, 20 +/- 2 min (brief ischemia), 58 +/- 5 min after the cross-clamping period (prolonged ischemia), and 21 +/- 4 min after reconstitution of coronary blood flow (reperfusion). Stress kinases and DNA damage sensor proteins (ATM, H2AX, p53) were determined by immunoblotting with specific antibodies. Electron microscopy analysis was carried out on ischemic and reperfused samples. ATP content, reactive oxygen species (ROS) levels, and cytochrome oxidase activity were determined by biochemical assays.
  
  
  RESULTS
  Ischemia caused accumulation of ROS, reduction of cytochrome C oxidase and ATP, and activation of stress kinases p38 and Jun terminal kinase. Electron microscopy showed significant mitochondrial swelling in the majority of cells, but no appreciable apoptosis of cardiomyocytes. During ischemia, myocytes were intensely stained by TUNEL, and many cells showed proliferative cell nuclear antigen-positive nuclei. Finally, we found in ischemic tissues increased p53/p21(WAF) levels and phosphorylation of histone H2AX, a substrate of ATM kinase, which marks double-strand DNA breaks. Reperfusion caused a robust extracellular signal-regulated kinase-1/2 activation, a marked reduction of TUNEL staining, and persistent activation of ATM checkpoint.
  
  
  CONCLUSIONS
  These data indicate that ischemia induces extensive DNA damage and activation of ATM checkpoint. Reperfusion allows the repair of the DNA lesions and salvage of ischemic cells.
authors.affiliation: Division of Anesthesiology, University of Cagliari, Cagliari, Italy.
authors.forename: Gian G
authors.initials: GG
authors.lastname: Corbucci
authors.forename: Cinzia
authors.initials: C
authors.lastname: Perrino
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Donato
authors.forename: Antonio
authors.initials: A
authors.lastname: Ricchi
authors.forename: Biagio
authors.initials: B
authors.lastname: Lettieri
authors.forename: Giancarlo
authors.initials: G
authors.lastname: Troncone
authors.forename: Ciro
authors.initials: C
authors.lastname: Indolfi
authors.forename: Massimo
authors.initials: M
authors.lastname: Chiariello
authors.forename: Enrico V
authors.initials: EV
authors.lastname: Avvedimento
mesh_terms: D000255:Adenosine Triphosphate / Q000235:genetics / Q000378:metabolism; D000328:Adult; D000368:Aged; D015153:Blotting, Western; D004249:DNA Damage; D003576:Electron Transport Complex IV / Q000235:genetics / Q000378:metabolism; D005260:Female; D006352:Heart Ventricles / Q000473:pathology* / Q000648:ultrastructure; D006801:Humans; D020287:In Situ Nick-End Labeling; D008297:Male; D008875:Middle Aged; D020928:Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017202:Myocardial Ischemia / Q000473:pathology*; D015428:Myocardial Reperfusion Injury / Q000473:pathology*; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D017382:Reactive Oxygen Species; D000255:Adenosine Triphosphate; D003576:Electron Transport Complex IV; D020928:Mitogen-Activated Protein Kinases