Abstract 9004: Creatine Kinase Over-Expression Improves ATP Kinetics and Contractile Function in the Post-Ischemic Myocardium

Abstract

ATP is the biochemical fuel for myocardial contraction but it is depleted during prolonged ischemia and its depletion may limit post-ischemic mechanical function. Because it has been difficult to augment ATP synthesis during and after ischemia, we generated mice that overexpress the myofibrillar isoform of Creatine Kinase (CKM), the primary energy reserve reaction of the heart, in cardiac-specific, conditional fashion to test the hypothesis that enhanced CKM expression increases ATP delivery via Creatine Kinase (CK) in ischemic-reperfused hearts and improves functional recovery. CKM mice were generated using a tet-off system. Gene induction was achieved by switching from a doxycycline diet (650mg/kg diet) to regular diet 4 weeks prior to the study. Isolated, retrograde-perfused hearts from control and CKM mice were subjected to 25 minutes of global ischemia and 40 minutes of reperfusion while cardiac function (rate pressure product, RPP) was monitored using a left ventricular balloon connected to a pressure transducer. A combination of 31P nuclear magnetic resonance magnetization transfer experiments at 11.7T and biochemical assays were used to measure the CK pseudo first-order rate constant (kf), the rate of ATP synthesis via CK (CK flux), intracellular pH and in vitro CK activity. Pre-ischemic kf and CK flux were three-fold higher in CKM than control hearts with no difference in RPP (Table). During ischemia, intracellular pH fell more in control than CKM hearts (6.10±0.29 vs. 6.46±0.08, p<0.05) with no difference in PCr and ATP levels between the two groups. Post-ischemic kf, PCr, CK flux and functional recovery were significantly higher in CKM vs. control hearts as was CK activity (21.6±7.23 vs 7.59±1.42 IU/mg protein, p<0.01). These observations demonstrate that augmenting CKM expression attenuates ischemic acidosis and improves not only [PCr] and rate of ATP synthesis in the post-ischemic myocardium but also recovery of contractile function.