Mechanisms of Myocardial Ischemic Injury in Diabetes

描述(申请人提供)：这项建议的总体目标是阐明高血糖增加心肌缺血和再灌注损伤的机制，目的是降低糖尿病和高血糖患者的心血管发病率和死亡率。一氧化氮受损(No.)信号转导被认为在糖尿病和高血糖时心血管疾病的发病机制中起关键作用，而内皮型一氧化氮合酶(ENOS)功能是一个关键因素。目前的提议将检验总体假设，即高血糖通过涉及活性氧、氮物种和四氢生物蝶呤(BH4)的途径减弱热休克(HSP)90/eNOS相互作用，从而损害心脏的心脏保护信号转导机制。在具体目标1中，我们将评估高血糖剂量依赖性地损害eNOS偶联并导致eNOS依赖性NO减少的假说。并通过调节HSP90/eNOS伴侣和BH4辅助因子在体内和体外内皮细胞和心肌细胞中的可用性而增加超氧阴离子(O2.-)。我们将提出这样的假设，即高血糖诱导的过氧亚硝酸盐(ONOO-)的形成通过降低磷酸酪氨酸-HSP90和增加硝基酪氨酸-HSP90来损害HSP90/eNOS，并通过减少BH4来实现。最后，我们将评估高血糖通过减弱HSP90/eNOS相互作用和降低BH4在体内的可获得性来阻断缺血预适应(IPC)的假说。这些实验将提供有关HSP90和高血糖通过蛋白质-蛋白质相互作用和改变自由基形成调节心脏保护作用的新机制信息，使用细胞、分子和药理学的综合方法。在具体目标2中，我们将解决以下假设：用磷酸酪氨酸磷酸酶抑制剂增加HSP90上的磷酸酪氨酸；用外源性BH4或其代谢前体sepiapterin增加BH4的利用率；用新的apoA-1模拟D4-F减少氧化应激，将增强HSP90/eNOS的结合；在体外和体内高血糖时恢复eNOS偶联；以及通过HSP90介导的途径恢复对高血糖下IPC产生的心肌梗死的保护。这些实验将阐明HSP90在心脏保护中的新作用，并将确认潜在的治疗糖尿病和高血糖的新靶点。简单描述：糖尿病患者的血糖升高会增加心脏病发作和死亡的风险。这项拟议的研究将评估热休克蛋白(HSP)90在促进心脏损伤抵抗方面的作用；将确定血糖对削弱HSP90影响的不利影响：并将确定潜在的糖尿病新治疗策略。

DESCRIPTION (provided by applicant): The overall objective of this proposal is to elucidate the mechanisms whereby hyperglycemia increases myocardial ischemia and reperfusion injury with the goal of reducing cardiovascular morbidity and mortality in patients with diabetes and hyperglycemia. Impaired nitric oxide (NO.) signaling is believed to play a key role in the pathogenesis of cardiovascular disease during diabetes and hyperglycemia and endothelial nitric oxide synthase (eNOS) function is a critical factor. The current proposal will test the overall hypothesis that hyperglycemia impairs cardioprotective signal transduction mechanisms in the heart by attenuating heat shock (HSP)90/eNOS interactions through a pathway involving reactive oxygen and nitrogen species and tetrahydrobiopterin (BH4). During Specific Aim 1, we will evaluate the hypotheses that hyperglycemia dose-dependently impairs eNOS coupling and results in eNOS-dependent decreases in NO. and increases in superoxide anion (O2.-) by modulation of HSP90/eNOS chaperone and BH4 co-factor availability in rabbits in vivo and in endothelial cells and cardiomyocytes in vitro. We will address the hypotheses that hyperglycemia-induced formation of peroxynitrite (ONOO-) impairs HSP90/eNOS by decreasing phosphotyrosine-HSP90 and increasing nitrotyrosine-HSP90; and by decreasing BH4. Finally, we will evaluate the hypothesis that hyperglycemia blocks ischemic preconditioning (IPC) by attenuating HSP90/eNOS interactions and by decreasing the availability of BH4 in vivo. These experiments will provide novel mechanistic information on the role of HSP90 and hyperglycemia to modulate cardioprotection through protein-protein interactions and altered radical formation using an integrated cellular, molecular and pharmacological approach. During Specific Aim 2, we will address the hypotheses that increasing phosphotyrosine on HSP90 with a phosphotyrosine phosphatase inhibitor; increasing BH4 availability with exogenous BH4 or its metabolic precursor sepiapterin; and decreasing oxidant stress with a novel apo A-1 mimetic D4-F will enhance HSP90 /eNOS association; restore eNOS coupling during hyperglycemia in vitro and in vivo; and restore protection against myocardial infarction produced by IPC in the presence of hyperglycemia through an HSP90-mediated pathway. These experiments will elucidate a novel role for HSP90 during cardioprotection and will confirm potential new therapeutic targets for the treatment of diabetes and hyperglycemia. Lay description: Increases in blood sugar that occur in individuals with diabetes increase the risk of heart attack and death. The proposed research will evaluate the role of heat shock protein (HSP)90 to promote resistance to heart injury; will determine the adverse effects of blood sugar to impair HSP90 effects: and will identify potential new treatment strategies for diabetes.