Regulation of lipid metabolism in diabetic myocardium

糖尿病的心脏后遗症是糖尿病人群发病率和死亡率的最大原因。在I型和II型糖尿病中，对脂肪酸底物的依赖增加，导致内源性心肌病变的发生，并增加糖尿病心肌对缺血/缺氧影响的敏感性。在目前的授予间隔期间，我们已经确定了一种新的钙非依赖性磷脂酶A2(IPA2Gamma)的完整基因组组织和蛋白质序列，我们认为它调节心肌细胞的能量储存(脂肪积累)和耗散(通过产生热量来去除FA)。因此，在特定的目标1中，我们将首先将iPA2γ纯化到均一状态，确定其底物特异性和转酰化过氧化体隔室中显著脂类的能力。在特定目标2中，将通过以心肌细胞特有的方式表征过度表达iPA2Gamma的小鼠和通过产生iPA2Gamma基因缺失的小鼠来检查心肌iPLA2 Gamma活性改变的生化和病理生理后遗症。I型和II型糖尿病对iPLA2-γ过度表达和缺失的转基因小鼠的影响将通过关注糖尿病状态下转基因依赖的血流动力学功能和脂流量的变化来检验。值得注意的是，编码iPLA2伽马的基因共分离到一个通过位置克隆确定的基因，该基因是皮马印度人II型糖尿病的多基因决定因素之一。因此，我们将首先在皮马印度人群体中鉴定iPLA2ganma基因的常见等位变异，并在Sf9细胞中表达这些等位变异，以确定其生化后遗症。最后，我们证明了以心肌细胞特有的方式过度表达iPLA2β的小鼠会出现缺血诱导的恶性室性心律失常。因此，在特定的目标4中，我们将通过这些转基因小鼠详细的血流动力学、电生理学和生化特征来研究iPLA2β过度表达在I型和II型糖尿病模型中的功能后遗症。糖尿病状态引起的iPLA2β的共价变化将被识别和表征。总体而言，项目1代表了一项多学科的、高度协同的、有针对性的建议，以确定心肌细胞脂质储存和利用的改变作为糖尿病患者过度心血管死亡率和发病率的主要决定因素的重要性。

The cardiac sequelae of diabetes represents the largest cause of morbidity and mortality in the diabetic population. In both Type I and Type II diabetes, an increased reliance on fatty acid substrate is present which leads to the development of an intrinsic cardiomyopathy and an increased susceptibility of diabetic myocardium to the effects of ischemia/hypoxia. During the current grant interval, we have defined the complete genomic organization and protein sequence of a novel calcium- independent phospholipase A2 (iPA2gamma), which we propose modulates energy storage (lipid accumulation) and dissipation (FA removal through heat production) in the cardiac myocyte. Accordingly, in Specific Aim 1 we will first purify iPA2gamma to homogeneity, determine its substrate specificity and ability to transacylate salient lipids in the peroxisomal compartment. In Specific Aim 2, the biochemical and pathophysiologic sequelae of alterations in myocardial iPLA2gamma activity will be examined both by characterizing mice over-expressing iPA2gamma in a cardiac myocyte specific fashion and by generating mice null for the iPA2gamma gene. The effects of both Type I and Type II diabetes on transgenic mice over-expressing and null for iPLA2gamma will be examined by focusing on the transgene-dependent alterations in hemodynamic function and lipid flux manifest in the diabetic state. Remarkably, the gene encoding iPLA2gamma cosegregates to a locus identified by positional cloning as one of the multigenic determinants of Type II diabetes in the Pima Indian population. Accordingly, we will first identify common allelic variants of the iPLA2ganma gene in the Pima Indian population and express those allelic variants in Sf9 cells to identify their biochemical sequelae. Finally, we have demonstrated that mice over-expressing iPLA2beta in a cardiac myocyte specific fashion have ischemia-induce malignant ventricular arrhythmias. Accordingly, in Specific Aim 4 we will examine the functional sequelae of iPLA2beta over-expression in both Type I and Type II models of diabetes through detailed hemodynamic electrophysiologic and biochemical characterization of these transgene mice. Covalent alterations in iPLA2beta induced by the diabetic state will be identified and characterized. Collectively, Project 1 represents a multi-disciplinary, highly synergistic, targeted proposal to identify the importance of alterations in cardiac myocyte lipid storage and utilization as a primary determinant of the excessive cardiovascular mortality and morbidity present in the diabetic patient.