Ischemia/reperfusion of heart causes contractile dysfunction,
necrosis and/or apoptosis and is a major cause of human death, but the molecular mechanisms are unclear. We show that
ischemia alone (without reperfusion) is sufficient to induce apoptosis and
mitochondrial dysfunction, and we have investigated the mechanism responsible; 30 and 60 min stop-flow
ischemia in Langendorff-perfused rat hearts induced progressive (a). release of
cytochrome c from mitochondria to cytosol, (b). inhibition of the mitochondrial respiratory functions, (c). activation of caspase-3-like
protease activity and (d).
DNA strand breaks (however, only 2% of myocyte nuclei were TUNEL positive at 60 min). Fifteen minutes pre-perfusion of hearts with
cyclosporin A, an inhibitor of mitochondrial-permeability transition (MPT), largely prevented all these ischemic changes. Pre-perfusion of hearts with
FK506, an inhibitor of
calcineurin, caused no protection. Pre-perfusion with
DEVD-CHO, an inhibitor of caspase-3-like
proteases, completely prevented
ischemia-induced
DNA strand breaks, but only partially blocked
cytochrome c release and mitochondrial respiratory inhibition. Reperfusion of hearts after 30 min
ischemia further stimulated
caspase activity and nuclear apoptosis. We conclude that
ischemia-induced MPT causes release of
cytochrome c, which then activates the
caspases that execute apoptosis and feedback to cause further
cytochrome c release. The MPT-induced
cytochrome c release is also largely responsible for the ischemic respiratory inhibition, which might contribute to contractile dysfunction or
necrosis at reperfusion.