4-hydroxy-2-nonenal in mitochondrial DNA damage and contractile dysfunction in diabetic heart: a role for aldehyde dehydrogenase 2

项目摘要： 糖尿病（DM）困扰着美国2600万人。大约65%的糖尿病患者死于 心血管并发症我们和其他人已经发现，DM增加活性氧（ROS）- 介导的醛类如4-羟基-2-壬烯醛（4 HNE）水平。4 HNE与大分子形成共价键 称为加合物，导致细胞损伤和心脏功能下降。醛脱氢酶 ALDH 2是一种心脏线粒体酶，在心脏中大大解毒4 HNE。我们和其他人已经 报道，在链脲霉素诱导的高血糖模型中，4 HNE蛋白加合物增加， 在心肌中ALDH 2活性与心肌病相关。尽管我们认为这会导致心脏功能障碍， 确切的机制尚不清楚。然而，大多数糖尿病患者患有2型糖尿病。因此，必须 研究心肌细胞中线粒体4 HNE增加和ALDH 2活性降低是否有助于 2型糖尿病模型中的心功能障碍。我们最近证明，高糖应激或4 HNE 给药降低了线粒体呼吸，增加了线粒体DNA（mtDNA）损伤， 培养的心肌细胞在我们对2型糖尿病小鼠心脏的初步研究中，我们发现， 线粒体水平的8-羟基鸟嘌呤（8 OHG），一种氧化的mtDNA产物，主要由8-羟基鸟嘌呤修复。 氧代鸟嘌呤糖基化酶（OGG）-1。接下来，我们发现OGG-1上4 HNE加合物的形成增加，而OGG-1上4 HNE加合物的形成减少。 心脏OGG-1水平。这些数据表明，4 HNE对OGG-1的加合降低了其水平和活性，从而 提高未代谢的8 OHG水平。因此，我们推测4 HNE介导的mtDNA损伤是线粒体损伤的一部分。 ALDH 2降低导致线粒体呼吸功能障碍从而导致心脏收缩的机制 功能障碍为了测试我们的想法，我们将在野生型C57 BL/6中使用高脂肪饮食诱导的2型DM模型， ALDH 2 *2突变小鼠。这种突变模仿东亚人的E487 K变体（ALDH 2 *2）， ALDH 2活性降低。我们将在心肌原位过表达ALDH 2和OGG-1基因，或者治疗我们的 用Alda-1治疗糖尿病小鼠，Alda-1是唯一可用于改善野生型和 突变ALDH 2。我们提出以下两个具体目标： 目标1.为了确定增加的4 HNE对mtOGG-1的加合是否导致mtDNA损伤， 线粒体呼吸和2型糖尿病心肌细胞收缩力受损。 目标2.为了确定在心脏病发作后减少4 HNE介导的mtDNA损伤是否是一种有效的方法， 2型糖尿病的功能障碍减弱了心肌病的发病机制。 这项研究将确定ALDH 2在2型糖尿病介导的心功能不全中的新作用，并建立 ALDH 2可能是2型糖尿病患者恢复心功能的治疗靶点。

Project Abstract: Diabetes mellitus (DM) afflicts 26 million people in the US. Around 65% of these diabetic patients die of cardiovascular complications. We and others have found that DM increases reactive oxygen species (ROS)- mediated aldehydes like 4-hydroxy-2-nonenal (4HNE) levels. 4HNE forms covalent bonds with macromolecules known as adducts, which lead to cellular damage and decreased cardiac function. Aldehyde dehydrogenase (ALDH2) is a cardiac mitochondrial enzyme that detoxifies 4HNE greatly in the heart. We and others have reported that in streptozotocin-induced hyperglycemic models increase in 4HNE protein adducts and decrease in myocardial ALDH2 activity correlate with cardiomyopathy. Although we think this causes cardiac dysfunction, the exact mechanism is unclear. However, most diabetic patients have type-2 DM. Thus, it is imperative to investigate whether increased mitochondrial 4HNE and lower ALDH2 activity in the cardiomyocytes contribute to cardiac dysfunction in type-2 DM models. We recently demonstrated that high glucose stress or 4HNE administration decreased mitochondrial respiration with increased mitochondrial DNA (mtDNA) damage in cultured cardiomyocytes. In our preliminary study using type-2 diabetic mouse heart, we found an increase in mitochondrial levels of 8-hydroxyguanine (8OHG), an oxidized mtDNA product, which is primarily repaired by 8- oxoguanine glycosylase (OGG)-1. Next, we found increased 4HNE adduct formation on OGG-1 and reduced cardiac OGG-1 levels. These data suggest that 4HNE adduction on OGG-1 reduces its level and activity thereby raising the unmetabolized 8OHG level. Thus, we postulate that 4HNE-mediated mtDNA damage is part of the mechanism by which lower ALDH2 causes mitochondrial respiratory dysfunction and thus cardiac contractile dysfunction. To test our idea, we will use a high-fat diet induced type-2 DM model in wild type C57BL/6 and ALDH2*2 mutant mice. This mutation mimics East Asians with the E487K variant (ALDH2*2), which exhibits lower ALDH2 activity. We will overexpress ALDH2 and OGG-1 genes in the myocardium in situ or treat our diabetic mice with Alda-1, the only specific drug available to improve the catalytic activity of both wild type and mutant ALDH2. We propose following two specific aims: Aim 1. To determine whether increased 4HNE adduction on mtOGG-1 causes mtDNA damage, poor mitochondrial respiration, and impaired cardiomyocyte contractility in type-2 DM. Aim 2. To determine whether decreasing 4HNE-mediated mtDNA damage after the onset of cardiac dysfunction in type2-DM attenuates pathogenesis of cardiomyopathy. This study will identify a novel role of ALDH2 in type-2 DM mediated cardiac dysfunction and establish that ALDH2 could be a therapeutic target for restoring cardiac function in type-2 diabetic patients.