title: Apoptosis of endothelial cells precedes myocyte cell apoptosis in ischemia/reperfusion injury.
abstract: BACKGROUND
  Apoptosis contributes to cell loss after ischemia/reperfusion injury in the heart. This study describes the time course and level of apoptosis in different cell types in the intact heart during ischemia/reperfusion injury.
  
  
  RESULTS
  Isolated Langendorff-perfused rat hearts were subjected to perfusion alone (control) or to 35 minutes of regional ischemia, either alone or followed by 5, 60, or 120 minutes of reperfusion. Sections were stained by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) and propidium iodide and with anti-von Willebrand factor, anti-desmin, or anti-active caspase 3 antibodies; they were then visualized by confocal microscopy. Sections were also examined by electron microscopy. No TUNEL-positive cells were seen in control hearts or hearts exposed to ischemia alone. Early in reperfusion, TUNEL staining was colocalized with endothelial cells from small coronary vessels. Endothelial apoptosis peaked at 1 hour of reperfusion and, at this time, there was clear perivascular localization of apoptotic cardiac myocytes, whose number was inversely proportional to their distance from a positive vessel. After 2 hours of reperfusion, apoptotic cardiac myocytes assumed a more homogeneous distribution. Active caspase 3 labeling was seen independent of DNA fragmentation during ischemia alone, but it colocalized with TUNEL staining over the 3 time points of reperfusion. Immunocytochemical findings were confirmed by electron microscopy and Western blotting.
  
  
  CONCLUSIONS
  In the very early stages of reperfusion, apoptosis is first seen in the endothelial cells from small coronary vessels. The radial spread of apoptosis to surrounding cardiac myocytes suggests that reperfusion induces the release of soluble pro-apoptotic mediators from endothelial cells that promote myocyte apoptosis.
authors.affiliation: Medical Molecular Biology Unit, Institute of Child Health, University College London, UK.
authors.forename: T
authors.initials: T
authors.lastname: Scarabelli
authors.forename: A
authors.initials: A
authors.lastname: Stephanou
authors.forename: N
authors.initials: N
authors.lastname: Rayment
authors.forename: E
authors.initials: E
authors.lastname: Pasini
authors.forename: L
authors.initials: L
authors.lastname: Comini
authors.forename: S
authors.initials: S
authors.lastname: Curello
authors.forename: R
authors.initials: R
authors.lastname: Ferrari
authors.forename: R
authors.initials: R
authors.lastname: Knight
authors.forename: D
authors.initials: D
authors.lastname: Latchman
mesh_terms: D000818:Animals; D017209:Apoptosis; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002452:Cell Count; D004730:Endothelium, Vascular / Q000201:enzymology / Q000473:pathology*; D020287:In Situ Nick-End Labeling; D066298:In Vitro Techniques; D008297:Male; D017202:Myocardial Ischemia / Q000201:enzymology / Q000473:pathology*; D015425:Myocardial Reperfusion; D015428:Myocardial Reperfusion Injury / Q000201:enzymology / Q000473:pathology*; D009206:Myocardium / Q000201:enzymology / Q000473:pathology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013997:Time Factors
chemical_list: C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases