LIPID SECOND MESSENGERS IN DIABETIC CARDIAC DYSFUNCTION

该计划项目的统一假设是改变了糖酵解 糖尿病中的流通量导致钙调节功能障碍- 心脏和血管中的独立磷脂酶A2。这 该项目研究了功能失调的调节机制的作用 钙离子作用下糖酵解偶联产生脂质第二信使 非依赖性磷脂酶A2在肌萎缩症发病机制中的作用 糖尿病心肌功能障碍与糖尿病易感性增加 糖尿病心肌到急性缺血。我们已经证明了 糖尿病心肌表现出三个病理改变特征 在脂代谢方面包括：1)花生四烯酸急剧耗竭 其内源磷脂储存库中的酸含量；2) 钙非依赖性磷脂酶A2水平显著升高 活性；以及3)花生四烯酸流入和流出的异常流量 肌细胞磷脂池。因此，具体目标1将审查 糖酵解通量与花生四烯酸偶联的重要性 糖尿病心肌的释放和溶血磷脂的产生。角色 钙非依赖性磷脂酶A2在这一过程中的作用 通过利用基于机制的 抑制剂(E)-6-(bromomethylene)-3-(1-naphthalenyl)-2-H-tetrahydropyran- 2-1(BEL)。其次，通过以下方式加速膜的水解作用 钙非依赖性磷脂酶A2在介导血管紧张素转换酶升高中的作用 糖尿病心肌对缺血性损伤的易感性将是 探索过了。在特定目标2中，花生四烯酸的异质性 糖尿病患者心肌亚细胞膜室的耗竭 心肌检查将通过开发的力量和敏感性 电喷雾电离质谱仪。大鼠肝组织中脂质通量的变化 糖尿病状态下的特定亚细胞膜基因座 利用定量电子显微镜放射自显影进行鉴定 而钙非依赖性磷脂酶A2在这一过程中的作用将 用BEL测定。在具体目标3中，机制 钙非依赖性磷脂酶A2显著升高的原因 糖尿病心肌的活性将通过分析 钙非依赖性磷脂酶A2质量、异构体的变化 糖尿病患者的成分和/或翻译后修饰 州政府。总的来说，这些研究代表了一个多学科的 旨在确定糖酵解偶联重要性的方法 脂质第二信使生成与心功能不全的关系 糖尿病状态。

The unifying hypothesis of the program project is that altered glycolytic flux in diabetes results in the dysfunctional regulation of calcium- independent phospholipase A2 in the heart and blood vessels. This project examines the role of the dysfunctional regulation of the glycolytically-coupled generation of lipid second messengers by calcium- independent phospholipase A2 in the pathogenesis of contractile dysfunction in diabetic myocardium and the increased susceptibility of diabetic myocardium to acute ischemia. We have demonstrated that diabetic myocardium exhibits three hallmarks of pathologic alterations in lipid metabolism including: 1) dramatic depletion of the arachidonic acid content present in its endogenous phospholipid storage depots; 2) markedly increased levels of calcium-independent phospholipase A2 activity; and 3) abnormal fluxes of arachidonic acid into and out of myocytic phospholipid pools. Accordingly, Specific Aim 1 will examine the importance of the coupling of glycolytic flux with arachidonic acid release and lysophospholipid generation in diabetic myocardium. The role of calcium-independent phospholipase A2 in this process will be determined by exploiting the specificity inherent in the mechanism-based inhibitor (E)-6-(bromomethylene)-3-(1-naphthalenyl)-2-H-tetrahydropyran- 2-one(BEL). Next, the role of accelerated membrane hydrolysis by calcium-independent phospholipase A2 in mediating the increased susceptibility of diabetic myocardium to ischemic injury will be explored. In Specific Aim 2, the heterogeneity of arachidonic acid depletion in myocardial subcellular membrane compartments in diabetic myocardium will be examined by exploiting the power and sensitivity of electrospray ionization mass spectrometry. Alterations of lipid flux in specific subcellular membrane loci in the diabetic state will be identified utilizing quantitative electron microscopic autoradiography and the role of calcium-independent phospholipase A2 in this process will be determined utilizing BEL. In Specific Aim 3, the mechanisms underlying the marked increase in calcium-independent phospholipase A2 activity in diabetic myocardium will be identified by analysis of alterations in calcium-independent phospholipase A2 mass, isoform composition and/or post-translational modifications in the diabetic state. Collectively, these studies represent a multidisciplinary approach targeted at identifying the importance of glycolytic coupling of lipid second messenger generation to cardiac dysfunction in the diabetic state.