Optimal timing of
therapeutic hypothermia for cardiac
ischemia is unknown. Our prior work suggests that
ischemia with rapid reperfusion (I/R) in cardiomyocytes can be more damaging than prolonged
ischemia alone. Also, these cardiomyocytes demonstrate
protein kinase C (PKC) activation and
nitric oxide (NO) signaling that confer protection against I/R injury. Thus we hypothesized that
hypothermia will protect most using extended
ischemia and early reperfusion cooling and is mediated via PKC and
NO synthase (NOS). Chick cardiomyocytes were exposed to an established model of 1-h
ischemia/3-h reperfusion, and the same field of initially contracting cells was monitored for viability and NO generation. Normothermic I/R resulted in 49.7 +/- 3.4% cell death.
Hypothermia induction to 25 degrees C was most protective (14.3 +/- 0.6% death, P < 0.001
vs. I/R control) when instituted during extended
ischemia and early reperfusion, compared with induction after reperfusion (22.4 +/- 2.9% death). Protection was completely lost if onset of cooling was delayed by 15 min of reperfusion (45.0 +/- 8.2% death). Extended
ischemia/early reperfusion cooling was associated with increased and sustained NO generation at reperfusion and decreased
caspase-3 activation. The NOS inhibitor
N(omega)-nitro-L-arginine methyl ester (200 microM) reversed these changes and abrogated
hypothermia protection. In addition, the PKCepsilon inhibitor myr-PKCepsilon v1-2 (5 microM) also reversed NO production and
hypothermia protection. In conclusion,
therapeutic hypothermia initiated during extended
ischemia/early reperfusion optimally protects cardiomyocytes from I/R injury. Such protection appears to be mediated by increased NO generation via activation of
protein kinase Cepsilon;
nitric oxide synthase.