Mechanisms of Ethanol-Induced Cardiac Protection

描述（由申请人提供）：十年前，我们发现在心脏缺血前短暂暴露于10-50 mM乙醇可使梗死面积减少约70%，这一过程依赖于epsilon蛋白激酶C （5PKC）的激活。在过去的资助期内，我们发现线粒体酶醛脱氢酶2 （ALDH2）的激活似乎是乙醇诱导的心肌缺血保护所必需和充分的。线粒体ALDH2对人类健康的重要性还体现在40%携带ALDH2基因失活突变的东亚人患各种与氧化应激和由此产生的有毒醛积累相关的慢性疾病（包括心肌梗死）的倾向增加。我们计划利用转基因小鼠（AIM 1A）确定乙醇诱导的细胞保护是否需要5PKC和ALDH2活性。接下来，我们将确定乙醇诱导胞质5PKC进入ALDH2所在的线粒体的机制（AIM 1B）。然后，我们将确定乙醇和其他活化剂激活ALDH2是否会减少醛合物负荷，从而降低细胞毒性（AIM 1C）。我们将确定在乙醇处理中积累的乙醛是否有助于乙醇诱导的心脏保护（AIM 1D），并确定乙醇诱导和5pkc介导的ALDH2磷酸化是否保护ALDH2活性免受长链醛的失活，以及这种作用是否与ALDH2的新小分子激活剂Alda （AIM 1E）一起增加。在AIM 2中，我们将研究硝酸甘油（NTG）抑制ALDH2导致乙醇诱导的心脏保护丧失。我们将确定抑制NTG诱导的ALDH2失活（NTG耐受性）的小分子（AIM 2A， B），并将确定NTG耐受性抑制剂对乙醇诱导的5PKC介导的急性缺血性损伤的心脏保护的作用（AIM 2C）。总之，这些研究将阐明与野生型和失活（ALDH2*2）形式的ALDH2动物的细胞保护相关的基本过程，以及适度的乙醇消耗如何影响它们。我们的研究还将提供新的工具，并通过动物模型测试其在心脏缺血治疗中的应用。公共卫生相关性：急性暴露于中等水平的乙醇可保护心脏免受缺血性损伤，如冠状动脉搭桥手术期间发生的损伤。我们最近发现，一种主要的酶参与代谢在氧化应激过程中积累的毒素，对这种乙醇诱导的心脏保护至关重要。由于乙醇的潜在毒性和成瘾性作用，我们寻找了直接激活这种解毒酶的新化合物，并计划在心脏缺血动物模型中测试它们的功效。该研究可能最终为诱导缺血性损伤的细胞保护提供新的手段。

DESCRIPTION (provided by applicant): Ten years ago, we found that a brief exposure to 10-50 mM ethanol prior to cardiac ischemia reduces infarct size by ~70% in a process that is dependent on activation of epsilon protein kinase C, 5PKC. In the past funding period, we found that activation of the mitochondrial enzyme, aldehyde dehydrogenase 2, ALDH2, appears to be required and sufficient for ethanol-induced cardiac protection from ischemia. The importance of mitochondrial ALDH2 in human health is also suggested by the increased propensity of 40% of East Asians that carry an inactivating mutation in the Aldh2 gene, Aldh2*2, to have a variety of chronic diseases associated with oxidative stress and the resulting accumulation of toxic aldehydes, including myocardial infarction. We plan to determine whether ethanol-induced cytoprotection requires 5PKC and ALDH2 activity, using genetically manipulated mice (AIM 1A). We will next identify the mechanisms that enable ethanol-induced entry of the cytosolic 5PKC into the mitochondria, where ALDH2 is found, (AIM 1B). We will then determine whether ALDH2 activation by ethanol and other activators reduces aldehydic adduct loads to reduce cytotoxicity (AIM 1C). We will determine whether acetaldehyde, which accumulates on ethanol treatment, contributes to ethanol-induced cardioprotection (AIM 1D) and determine whether ethanol- induced and 5PKC-mediated phosphorylation of ALDH2 protects ALDH2 activity from inactivation by long chain aldehydes and whether the effect is additive with new small molecule activators of ALDH2, called Alda (AIM 1E). In AIM 2, we will study the loss of ethanol-induced cardioprotection due to ALDH2 inhibition by nitroglycerine (NTG). We will identify small molecule that inhibit NTG-induced ALDH2 inactivation (NTG tolerance) (AIM 2A, B) and will determine the effect of NTG tolerance inhibitors on ethanol-induced 5PKC- mediated cardioprotection from acute ischemic damage ex vivo and in animal models (AIM 2C). Together, these studies will elucidate fundamental processes associated with cytoprotection in animals with wildtype and inactive (ALDH2*2) form of ALDH2 and how moderate ethanol consumption affects them. Our studies will also provide new tools and test their application as treatment for cardiac ischemia using animal models. PUBLIC HEALTH RELEVANCE: Acute exposure to moderate levels of ethanol protects the heart from ischemic injury, such as that occurring during coronary bypass surgery. We recently found that a major enzyme involved in metabolizing toxins that accumulate during oxidative stress is critical for this ethanol-induced cardioprotection. Because of the potential toxic and addictive effects of ethanol, we searched for new compounds that directly activate this detoxifying enzyme and plan to test their efficacy in animal models of cardiac ischemia. This study may eventually provide new means to induce cytoprotection in humans subjected to ischemic insult.