Mechanisms of Ethanol-Induced Cardiac Protection

描述（由申请人提供）：10年前，我们发现在心肌缺血前短暂暴露于10-50 mM乙醇可使梗死面积减少约70%，这一过程依赖于β蛋白激酶C（5 PKC）的激活。在过去的资助期间，我们发现，激活线粒体酶，醛脱氢酶2，ALDH 2，似乎是必要的，足以乙醇诱导的心肌缺血保护。线粒体ALDH 2在人类健康中的重要性也通过40%的东亚人携带Aldh 2基因中的失活突变Aldh 2 *2的增加的倾向来表明，这些东亚人具有与氧化应激相关的各种慢性疾病以及由此产生的有毒醛的积累，包括心肌梗死。我们计划确定乙醇诱导的细胞保护是否需要5 PKC和ALDH 2活性，使用基因操作小鼠（AIM 1A）。我们接下来将确定乙醇诱导的胞质5 PKC进入线粒体的机制，在线粒体中发现ALDH 2（AIM 1B）。然后，我们将确定乙醇和其他活化剂对ALDH 2的活化是否减少了细胞毒性加合物负荷（AIM 1C）。我们将确定乙醇处理时积累的乙醛是否有助于乙醇诱导的心脏保护（AIM 1D），并确定乙醇诱导的和5 PKC介导的ALDH 2磷酸化是否保护ALDH 2活性免受长链醛的失活，以及该作用是否与新的ALDH 2小分子激活剂Alda（AIM 1 E）相加。在AIM 2中，我们将研究由于硝酸甘油（NTG）抑制ALDH 2而导致的乙醇诱导的心脏保护作用的丧失。我们将鉴定抑制NTG诱导的ALDH 2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5 PKC介导的心脏保护免受离体和动物模型中的急性缺血性损伤的作用（AIM 2C）。总之，这些研究将阐明与野生型和非活性（ALDH 2 *2）形式的ALDH 2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，并使用动物模型测试它们作为心肌缺血治疗的应用。 公共卫生相关性：急性暴露于中等水平的乙醇可以保护心脏免受缺血性损伤，例如冠状动脉搭桥手术期间发生的损伤。我们最近发现，参与代谢氧化应激过程中积累的毒素的主要酶对乙醇诱导的心脏保护作用至关重要。由于乙醇的潜在毒性和成瘾作用，我们寻找直接激活这种解毒酶的新化合物，并计划在心脏缺血的动物模型中测试其功效。这项研究可能最终提供新的手段，诱导细胞保护人类受到缺血性损伤。

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.