Non-specific caspase inhibition reduces infarct size and improves post-ischaemic recovery in isolated ischaemic/reperfused rat hearts

Abstract.

Myocardial ischaemia and reperfusion lead to myocardial cell death due, at least in part, to apoptotic mechanisms. Although cysteinyl aspartate-specific proteinase (caspase) activation is a major event and the most-cited culprit in the development of apoptosis, its potential contribution to ischaemic myocardial cell death is largely unknown. To study the role of caspase activation, isolated rat hearts (n=6 per group) were subjected to 30 min coronary artery occlusion followed by 120 min reperfusion. A non-selective [0.1 or 0.5 µM acetyl-Tyr-Val-Ala-Asp chloromethylketone (YVAD-cmk)] or selective caspase inhibitors [0.07 or 0.2 µM acetyl-Asp-Glu-Val-Asp-cmk (Ac-DEVD-cmk, caspase-3 inhibitor); 0.07 or 0.2 µM benzoxycarbonyl-Leu-Glu-OMe-His-Asp(OMe)-fluoromethylketone (z-LEHD-fmk, caspase-9 inhibitor)] were added to the perfusate at the start of reperfusion. Non-selective caspase inhibition with 0.1 or 0.5 µM YVAD-cmk limited infarct size: (21±4%, P<0.05; 17±3%, P<0.05, respectively) compared with the ischaemic/reperfused control (32±5%). In hearts treated with 0.1 or 0.5 µM caspase II non-selective inhibitor, the fraction of terminal-deoxynucleotidyl-transferase deoxyuridine nick end labelling (TUNEL)-positive myocyte nuclei in the infarcted zone was reduced from the ischaemic/reperfused non-treated control of 11.2±2.1% to 6.2±1.6% (P<0.05) and 1.2±0.2% (P<0.05), respectively. The recovery of post-ischaemic cardiac function (coronary flow, aortic flow and left-ventricular developed pressure) improved significantly with the application of the non-selective caspase inhibitor as well. In hearts perfused with specific caspase inhibitors (caspase-3 and caspase-9) there was no significant reduction in the infarct size, no improvement in post-ischaemic cardiac function and no reduction of apoptotic cell death. We conclude that non-specific inhibition of caspases may be therapeutically beneficial in myocardial ischaemia/reperfusion-induced damage, while selective caspase inhibitors may fail to prevent such reperfusion-induced injury in our model system.