Metabolic enzyme activity mediating myocardial stunning

描述（由申请人提供）：已知通过丙酮酸脱氢酶（PDH）激活刺激碳水化合物氧化可改善缺血后心肌的收缩恢复。我们已经确定激活PDH的益处既不依赖于糖酵解也不依赖于线粒体能量产生。相反，细胞溶质氧化还原状态的PDH依赖性变化影响缺血后心脏的恢复。我们的研究结果表明，在清醒的猪体内心脏再灌注的第一个小时内，PDH激活逆转了早期再灌注损伤，导致收缩力的持续改善。令人兴奋的新数据表明，刺激复氧心肌细胞中的丙酮酸氧化可改善对钙的收缩反应，消除钙超载，并改善收缩舒张速率。因此，本研究探讨了PDH激活通过细胞溶质氧化还原平衡的有利变化消除早期再灌注损伤的假设，细胞溶质氧化还原平衡改善钙稳态和线粒体功能，由于消除钙超载和潜在的肌丝的完整性。这一假设将在单一的成年大鼠心肌细胞和在隔离，灌注大鼠心脏使用一种新的组合技术，包括光学显微镜和13 C NMR光谱测试。具体目标是：2）a）评估SERCA 2a活性对增强的碳水化合物氧化的响应，作为再灌注心脏中改善的钙处理的机制; B）如果在再灌注心脏中通过PDH活化提高SERCA 2a活性，则确定在体内大鼠心脏中SERCA 2a过表达是否产生类似的有利于收缩性恢复; 3）确定在再灌注心肌中在增加的碳水化合物氧化期间影响收缩功能的细胞溶质氧化还原状态的钙和诱导的转变之间的关系; 4）阐明钙激活的线粒体脱氢酶α-酮戊二酸脱氢酶与细胞溶质还原当量转运活性之间的关系/竞争（苹果酸-天冬氨酸穿梭）; 5）检查增强的碳水化合物氧化通过减少肌丝蛋白降解和改善钙敏感性来逆转顿抑的潜力。

DESCRIPTION (provided by applicant): Stimulating carbohydrate oxidation via pyruvate dehydrogenase (PDH) activation is known to irmprove contractile recovery of postischemic myocardium. We have determined that the benefits of activating PDH rely neither on glycolytic nor mitochondrial energy production. Instead, PDH-dependent changes in cytosolic redox state influence the recovery of the postischemic heart. Our results on PDH activation during the first hour of reperfusion in the in vivo heart of conscious pigs demonstrate reversal of early reperfusion injury that results in sustained improvement in contractility. Exciting new data show that stimulating pyruvate oxidation in reoxygenated cardiomyocytes improves contractile response to calcium, eliminates calcium overload, and improves contractile relaxation rate. Therefore, this study explores the hypothesis that PDH activation eliminates early reperfusion injury via favorable shifts in cytosolic redox balance that improve calcium homeostasis and mitochondrial function due to elimination of calcium overload and potentially, the integrity of the myofilaments. This hypothesis will be tested in both the single adult rat cardiomyocyte and in the isolated, perfused rat heart using a novel combination of techniques that include optical microscopy and 13C NMR spectroscopy. Specific aims are: 1) Determine the effects of cytosolic redox state on intracellular calcium levels and contractile response to calcium in reoxygenated cardiomyocytes; 2) a) Evaluate the response of SERCA2a activity to augmented carbohydrate oxidation as a mechanism of improved calcium handling in reperfused hearts; b) if SERCA2a activity is improved by PDH activation in reperfused hearts then determine whether SERCA2a over expression in vivo rat hearts produces similar benefits to contractile recovery; 3) Determine the relationship between calcium and induced shifts in cytosolic redox state in affecting contractile function during augmented carbohydrate oxidation in reperfused myocardium; 4) Elucidate the relationship/competition between the calcium activated mitochondrial dehydrogenase, alpha-ketoglutarate dehydrogenase, versus activity of cytosolic reducing equivalent transport (malate-aspartate shuttle) in response to calcium load in the reperfused myocardium; 5) Examine the potential for augmented carbohydrate oxidation to reverse stunning via reduced myofilament protein degradation and improved calcium sensitivity.