| Myocardial damage due to ischemic tissue injury is the single leading cause of death worldwide. Efforts made to redress this cost have uncovered a large number of stimuli, which if applied prior to an ischemic insult, can limit myocardial ischemia-reperfusion (I/R) injury. Collectively, these stimuli have been termed myocardial preconditioners, and they elicit a myocardial phenotype that is commonly referred to as being cardioprotected. While most preconditioners are of limited clinical application because they educe pathologic responses after repeated application, a small number have been shown to remain chronically protective. We have attempted to understand the mechanisms underlying two such sustainable forms of cardioprotection: (1) Protection characteristic of the female myocardium - sex-specific cardioprotection (SSC), and (2) Protection conferred by life-long caloric restriction.;At the level of the cardiac myocyte, separate lines of evidence indicate that SSC depends upon function of the sarcolemmal ATP-sensitive potassium channel (SarcKATP), and protein kinase C (PKC). While PKC has been shown to activate SarcKATP in other forms of inducible cardioprotection, it is not known whether their interaction is a component of the mechanisms underlying SSC. Accordingly, we pursued the hypothesis that they do interact, either directly or diffusely, to contribute to the I/R resistance exhibited by the female heart. Using the excised hearts of adult Sprague-Dawley rats in a Langendorff model of I/R, we found that the I/R resistance exhibited by females was susceptible to individual blockade of either PKC or sarcK ATP. Simultaneous blockade of both PKC and SarcKATP did not combine to further expand infarction in female hearts, suggesting that these two proteins act in the same pathway underlying SSC.;SarcKATP is expressed at a higher level in the female myocardium than that of males, and as such, we pursued a second hypothesis that the interaction of PKC and sarcKATP in SSC may involve trafficking of sarcK ATP from an intracellular depot to the cell surface. To investigate this hypothesis we assessed the sarcolemmal concentrations of PKC and sarcK ATP in male and female hearts. These experiments were performed in the presence and absence of ischemia, and catalytic blockade of PKC. We found the female sarcolemma to be basally enriched with sarcKATP, and that this enrichment was susceptible to PKC blockade, suggesting that PKC is involved in enriching the female sarcolemma with sarcKATP. This enrichment was not due to sex-differences in the sarcolemmal concentration of the epsilon isoform of PKC (PKC&egr;) or of PKC&egr; phosphorylation. However, that phosphorylation was dramatically lowered by pharmacologic blockade in both sexes, suggesting that our blockade effectively disrupted sarcolemmal PKC activity. In concert these data indicate that PKC plays a permissive role in SSC, which may involve trafficking of sarcKATP between the sarcolemma and intracellular depots of the female myocyte.;Short-term caloric restriction (CR) enhances myocardial I/R resistance in a manner that does not appear to rely upon sarcKATP, but is known to involve AMP-activated protein kinase (AMPK). The potential for this AMPK-dependent mechanism to remain intact over a longer-term CR intervention is unknown. We hypothesized that, as for short-term CR, AMPK activation underlies protection afforded by life-long CR. Relative to ad libitum controls (AL), we observed pronounced infarct-resistance in a mouse model of lifelong CR (LCR) after global ischemia/reperfusion challenge by the Langendorff method. Post-ischemic functional recovery was also markedly enhanced by LCR, and together these findings indicate that life-long CR is among the most potent forms of chronic cardioprotection. Importantly, LCR-associated protection measured by both infarct size and post-ischemic ventricular function was dependent upon AMPK, as assessed by pre-ischemic pharmacologic blockade (LCR+AraA). This AMPK-dependent protection was associated with a modest increase in total AMPKalpha in LCR hearts, and greatly enhanced AMPKalpha phosphorylation relative to AL. We conclude that the AMPK-dependent protection characteristic of short-term CR does not decay with a life-long intervention, but remains intact at a level that is pronounced compared to other chronically effective stimuli. This enhanced protection involves a modest enhancement of AMPKalpha expression, but much greater AMPK activation as indicated by phosphorylation at Thr172 on the alpha subunit.;Together these two investigations suggest that activation of the PKC-SarcK ATP axis, and AMPK, underlie distinct forms of chronic cardioprotection. As such, each of these targets may offer independent and synergistic clinical potential that should be tested by pharmacologic or genetic activation. |
