APOPTOTIC AND NECROTIC MYOCYTE CELL DEATHS ARE INDEPENDENT CONTRIBUTING VARIABLES OF INFARCT SIZE IN RATS

Programmed cell death in the myocardium has been linked to ischemia re perfusion injury as well as to excessive mechanical forces associated with increases in ventricular loading. Moreover, hypoxia activates the suicide program of cardiac myocytes in vitro. Because the supplied po rtion of the ventricular wall is ischemic and subjected to high levels of systolic and diastolic stresses (acutely after coronary artery occ lusion), apoptosis and necrosis may contribute independently to myocyt e cell death after infarction. Therefore, myocardial infarction was pr oduced in rats, and, after the determination of ventricular hemodynami cs, the contribution of apoptotic and/or necrotic myocyte cell death t o infarct size was measured quantitatively from 20 minutes to 7 days a fter coronary artery occlusion. Programmed cell death was assessed by the terminal deoxynucleotidyl transferase assay and by the electrophor etic detection of DNA laddering. Myocyte necrosis was evaluated by myo sin monoclonal Ab labeling. Moreover, the expression of Bcl-2, Pax, an d Fas proteins in myocytes was examined by immunocytochemistry. Myocyt e cell death by apoptosis and necrosis comprised nearly 3 million myoc ytes at 2 hours. Apoptotic cell death involved 2.8 million cells and n ecrotic cell death only 90,000 myocytes. Apoptosis continued to repres ent the major independent form of myocyte cell death, affecting 6.6 mi llion myocytes at 4.5 hours. Myocyte necrosis peaked at 1 day, includi ng 1.1 million myocytes. DNA electrophoretic analysis confirmed these observations by showing nucleosomal ladders at 2-3 hours, 4.5 hours, 1 day, and 2 days after coronary artery occlusion. Myocytes showing bot h DNA strand breaks and myosin labeling were a prominent aspect of myo cardial damage only after 6 hours. Finally, the expression of Bcl-2 an d Fas in myocytes increased 18-fold and 131-fold, respectively. In con clusion, programmed myocyte cell death is the major form of myocardial damage produced by occlusion of a major epicardial coronary artery, w hereas necrotic myocyte cell death follows apoptosis and contributes t o the progressive loss of cells with time after infarction. The enhanc ed expression of Fas may be implicated in the activation of apoptosis in spite of the increase in Bcl-2, which tends to preserve cell surviv al.