Coronary reperfusion following ischemia: Different expression of Bcl-2 and bax proteins, and cardiomyocyte apoptosis

Abstract

The aim of this work was to examine factors that could be involved in the occurrence of apoptosis in rat hearts subjected to coronary occlusion followed by reperfusion. To this end, we studied the expression of the pro- and anti-apoptotic factors, bax and bcl-2, respectively, in reperfused ischemic hearts and in hearts injected with bFGF or saline. In anesthetized rats the left coronary artery was occluded for 45 min, the anesthesia withdrawn and the occlusion removed to allow reperfusion; in sham-operated rats the occlusion was omitted. After 4 hours the rats were decapitated and the heart excised. Sections from the left ventricle were stained with anti-bcl-2-antibody and anti-bax-antibody using the TUNEL method which detects apoptosis. Fragmentation of DNA isolated from reperfused ventricles was examined by agarose electrophoresis. In reperfused hearts no bcl-2 staining was observed in the discrete area in which many cardiomyocyte nuclei were stained by the TUNEL method; outside this area staining for bcl-2 was more marked than in sham-operated rats. Sections from reperfused hearts were stained for bax protein over a wide area including the apoptotic region; sham-operated hearts showed little reaction. Staining for bcl-2 was demonstrable in some nuclei in hearts from saline-injected rats; the numbers were unaffected by i. v. bFGF. Ischemia/reperfusion increases the overall expression of both bcl-2 and bax proteins, but bcl-2 is lost from the reperfused area as indicated by TUNEL staining. Accordingly, the ratio of bcl-2 to bax was reduced in the reperfused area, indicating a pro-apoptotic trend. The marked increase in bcl-2 outside the reperfused area could be a mechanism with which to salvage surviving cardiomyocytes.