Roles of Endothelial and Smooth Muscle KATP Channels in Myocardial Ischemic Injury

摘要 肌膜上的ATP敏感性钾通道(KATP)在心脏中大量表达。有几个组织已经 现在确定了这些通道在介导对缺血损伤的心脏保护中的关键作用，以及它们的 参与缺血预适应的保护机制。在心里有几种不同的 KATP通道的亚型，对其在缺血和再灌注过程中的作用知之甚少。特别的 人们感兴趣的是存在于冠状动脉平滑肌(SM)和冠状动脉内皮细胞中的KATP通道亚型 细胞(EC)。越来越多的人关注这些冠脉通道作为血流调节和 然而，人们对它们的了解相对较少。SM和EC KATP通道不同于 心室肌KATP通道，它们也是不同的。我们理解的一个主要障碍 它们在心肌缺血等复杂事件中各自的作用是目前可用的不足 专门研究这两种渠道子类型的资源。我们已经产生了新的遗传小鼠模型 它可以区分KATP通道这些子类型，并用这些EC KATP通道之一显示 强烈参与缺血/再灌流期间的心肌保护。拟议研究的目标是 系统研究EC和SM KATP通道亚型在冠状动脉调节中的作用(S) 缺血时的血液流动、保护和对缺血预适应的保护反应。我们 EC和SM KATP通道亚型均参与冠脉血液调节的假设 流动和心脏保护，但通过截然不同的机制。使用新颖且经过验证的 条件性基因敲除小鼠，我们将专门针对EC或SM KATP通道亚型。建议进行的研究 有三个目标。在目标1中，我们将研究这两种冠脉KATP通道亚型在血液中的作用 在缺血期间流动。我们将使用隔离、加压的微血管和隔离、灌流的心脏 正常、缺氧和缺血状态。此外，我们还将研究EC和SM KATP通道的作用 在心肌中出现“无复流”现象。目标2将研究EC和SM KATP通道在 利用在体小鼠I/R模型进行心肌保护及研究心肌梗死的调节途径 发展。目标3将研究EC和SM KATP通道亚型在缺血时的作用 使用体内小鼠I/R模型和细胞分析进行预适应。我们还将审查这些药物的贩运情况 KATP通道亚型作为潜在的保护机制及分子信号通路研究 牵涉其中。这项由多名调查员参与的提案结合了三位资深调查员的专业知识； Lefer在活体心脏缺血/再灌注模型方面的丰富专业知识，以及Coetzee博士在 用电生理、生化和分子方法研究KATP通道和Tinker博士的 擅长研究血管KATP通道中的分子信号通路。拟议的研究将 为冠脉KATP通道亚型的独特功能提供重要的分子见解 病理生理条件。

SUMMARY Sarcolemmal ATP-sensitive K+ (KATP) channels are abundantly expressed in the heart. Several groups have now identified a key role for these channels in mediating cardioprotection against ischemic injury and their participation in the protective mechanism of ischemic preconditioning. In the heart there several different subtypes of KATP channels and little is known about the roles during ischemia and reperfusion. Of particular interest are the KATP channel subtypes present in the coronary smooth muscle (SM) and coronary endothelial cells (EC). There is increasing focus on these coronary channels as a target for blood flow regulation and cardioprotection, yet they are relatively poorly understood. The SM and EC KATP channels are distinct from ventricular KATP channels and they are also distinct from each other. A major barrier to our understanding of their respective roles during a complex event such as myocardial ischemia is the lack of currently available resources specifically to study these two channel subtypes. We have generated novel genetic mouse models that can distinguish these subtypes of KATP channels and show with one of these that EC KATP channels strongly participate in myocardial protection during ischemia/reperfusion. The goal of the proposed studies is systematically to examine the role(s) of the EC and SM KATP channel subtypes in the regulation of coronary blood flow, protection during ischemia and the protective response to ischemic preconditioning. We hypothesize that both EC and SM KATP channel subtypes contribute to the regulation of coronary blood flow and cardioprotection, but through distinctly different mechanisms. Using novel and validated conditional knockout mice, we will specifically target EC or SM KATP channel subtypes. The proposed studies have three Aims. In Aim 1, we will investigate the roles of these two coronary KATP channel subtypes in blood flow during ischemia. We will use isolated, pressurized microvessels and isolated, perfused hearts under normal, hypoxic and ischemic conditions. We will additionally investigate the role of EC and SM KATP channels in the myocardial “no-reflow” phenomenon. Aim 2 will investigate the roles of EC and SM KATP channels in myocardial protection using an in vivo murine I/R model and investigate pathways that regulate infarct development. Aim 3 will investigate the contribution of EC and SM KATP channel subtypes during ischemic preconditioning using an in vivo murine I/R model and cellular assays. We will also examine trafficking of these KATP channel subtypes as a potential protective mechanism and investigate molecular signaling pathways involved. This multi-investigator proposal combines the expertise of three highly established investigators; Dr. Lefer’s extensive expertise with in vivo cardiac ischemia/reperfusion models, and Dr. Coetzee’s track record of studying KATP channels with electrophysiological, biochemical and molecular approaches and Dr. Tinker’s expertise in studying molecular signaling pathways in vascular KATP channels. The proposed studies will provide important molecular insights into the unique functions of coronary KATP channel subtypes under pathophysiological conditions.