Comprehensive and translational analysis of cardioprotective signal transduction in response to conditioning

缺血后处理，一个简单的机械动作在再灌注开始，减少梗死面积缺血/再灌注后，在所有物种测试到目前为止，包括人，其潜在的信号转导尚不清楚。我们已经证明，在猪中，与啮齿动物不同，再灌注损伤补救激酶（RISK）系统的激活不是后处理保护的原因。因此，目前的建议集中在替代信号通路：肿瘤坏死因子α（TNFα）在猪中具有心脏保护作用，鞘氨醇和STAT 3都在TNFα的下游信号级联中。因此，我们将集中在鞘氨醇和STAT 3的作用相结合的蛋白质印迹法，梗死面积定量，和药理学阻断剂的方法。第二个重点是线粒体通透性转换孔（mPTP）在心肌保护中的作用。再灌注时mPTP的关闭似乎是心肌细胞存活的决定性因素。我们想研究在猪的实验模型中，后处理是否也涉及再灌注时mPTP的关闭，特别是酸中毒在这种mPTP关闭中的作用。因此，我们将测量和操纵心肌pH值和分析mPTP开放后处理的猪心脏分离线粒体。环孢菌素A（CsA）可能会延迟mPTP开放，并减少梗死面积时，在再灌注，即使在人，然而，CsA也抑制蛋白磷酸酶。因此，我们将分析CsA对mPTP开放和蛋白磷酸酶活性的影响，并将其与梗死面积缩小相关。

Ischemic postconditioning, a simple mechanical maneuver at the onset of reperfusion, reduces infarct size after ischemia/reperfusion in all species tested so far, including man; its underlying signal transduction is not yet understood. We have demonstrated that in pigs, different from rodents, activation of the reperfusion injury salvage kinase (RISK) system is not causal for postconditioning s protection. The present proposal therefore focuses on alternative signaling pathways: tumor necrosis factor α (TNFα) is cardioprotective in pigs, and both sphingosine and STAT3 are in TNFα s downstream signaling cascade. Therefore we will focus on the role of sphingosine and STAT3 by a combination of Western blotting, infarct size quantification, and pharmacological blocker approaches. The second focus will be on the role of the mitochondrial permeability transition pore (mPTP) in myocardial protection. Closure of the mPTP at reperfusion appears decisive for cardiomyocyte survival. We want to investigate whether postconditioning in our experimental model in pigs also involves mPTP closure at reperfusion, notably the role of acidosis in such mPTP closure. We will therefore measure and manipulate myocardial pH and analyze mPTP opening in isolated mitochondria from postconditioned pig hearts. Cyclosporine A (CsA) presumably delays mPTP opening and reduces infarct size when given at reperfusion, even in man; however, CsA also inhibits protein phosphatases. We will therefore analyze the effects of CsA on mPTP opening and on protein phosphatase activity and correlate them to infarct size reduction.