AMPK, Metabolic and Inflammatory Stress and the Endothelial Cell

说明应用程序的广泛、长期目标和具体目标，并提及与健康相关的 ；项目(即与该机构使命的相关性)。简明扼要地描述实现这些目标的研究设计和方法。描述 V你将用来追求这些目标的基本原理和技术。 此外，我还用两三句话，通俗易懂地描述了这项研究与公共卫生的相关性。如果应用程序得到资助，则此 我的描述，按原样，将成为公开信息。因此，不包括专有/机密信息。不要超过空格 ：提供。 代谢综合征的临床定义是一种以血脂异常为特征的疾病， 高血压、中心性肥胖和过早动脉粥样硬化性心血管疾病的易感性以及 2型糖尿病。它还与肌肉中的毛细血管稀疏和血管生成障碍有关。在 在授权期之前，我们证明了AMPK在培养的血管内皮细胞中是有效的。 ‘(HUVEC，HAEC)，它的激活保护它们免受细胞凋亡、炎症和线粒体的攻击 (在富含葡萄糖、棕榈酸酯或肿瘤坏死因子-α的培养液中孵育引起的功能障碍。此外，我们 证明了Sirt1/LKB1/AMPK信号机制在各种培养细胞和大鼠中的存在 我们已经获得了在培养的内皮细胞中存在它的初步证据。在项目1中， 我们将通过开展以下研究来扩展这项研究：1)确定 Sirt1和LKB1在糖剥夺等因素介导AMPK激活中的作用 2)测定高血糖对血管内皮细胞的影响 诱导胰岛素抵抗和细胞凋亡，增强肿瘤坏死因子-α诱导的黏附分子 通过引起AMPK失调(活性降低或受损)在内皮细胞中表达 激活)。如果如预期的那样，我们将确定这种失调是否涉及抑制 Pf Sirt1/LKB1机制[可能通过SirT1的翻译后氧化修饰(项目2)]， 和/或蛋白磷酸酶的激活；以及3)检查AMPK介导的事件是否在 内皮细胞特异性SirT1或SirT1基因缺失的小鼠体内血管系统受损 LKB1。研究将在沃尔什博士(核心B)产生的cre/lox小鼠身上进行，他将利用这些小鼠 研究缺血诱导的血管生成(项目3)和科恩博士(项目2)的动脉粥样硬化形成。我们 将与Nathan LeBrasseur博士一起描述运动对这些小鼠和对照组小鼠的影响 啮齿动物运动生理学家。特别是，我们将确定SirT1和LKB1下调调控如何影响 通常发生的肌肉毛细血管密度增加和动脉粥样硬化改变的减少 对具有代谢综合征表型的啮齿类动物的运动反应。 J-？ 这些研究将提供有关SirT1/LKB1/AMPK信号功能的第一批信息 血管系统中的机制。他们还应该提供新的见解，即它的监管失调是否会 是与代谢综合征相关的内皮细胞功能障碍的原因之一，也是其靶点 心理治疗。 ，

State the application's broad, long-term objectives and specific aims, making reference to the health relatedness of ; the project (i.e., relevance to the mission of the agency). Describe concisely the research design and methods for achieving these goals. Describe v the rationale and techniques you will use to pursue these goals. i In addition, in two or three sentences, describe in plain, lay language the relevance of this research to public health. If the application is funded, this i description, as is, will become public information. Therefore, do not include proprietary/confidential information. DO NOT EXCEED THE SPACE : PROVIDED. The metabolic syndrome has been defined clinically as a disorder characterized by dyslipidemia, hypertension, central obesity and a predisposition to premature atherosclerotic cardiovascular disease and type 2 diabetes. It is also associated with capillary rarefaction and impaired angiogenesis in muscle. In the preceding grant periods, we demonstrated that AMPK is operative in cultured vascular endothelial cells '(HUVEC, HAEC) and that its activation protects them against the apoptosis, inflammation and mitochondrial (dysfunction caused by incubation in media enriched in glucose, palmitate or TNF-alpha. In addition, we demonstrated the existence of a Sirt1/LKB1/AMPK signaling mechanism in various cultured cells and in rat liver in vivo and we have obtained preliminary evidence for its presence in cultured endothelium. In project 1, we will extend this research by carrying out studies with the following objectives: 1) To characterize the roles of Sirtl and LKB1 in mediating AMPK activation by such factors as glucose deprivation, phenformin, and resveratrol in cultured endothelial cells; 2) To determine whether hyperglycemia induces insulin resistance and apoptosis and enhances TNF-alpha - induced adhesion molecule expression in endothelial cells by causing AMPK dysregulation (decreased activity or impaired activation). If as anticipated it does, we would then determine whether this dysregulation involves inhibition pf the Sirt1/LKB1 mechanism [possibly by post-translational oxidative modification of SirT1 (with project 2)], and/or activation of protein phosphatases; and 3) To examine whether AMPK-mediated events in the vasculature are impaired in vivo in mice with endothelial-cell specific deletions of either SirT1 or LKB1. Studies will be done in cre/lox mice that will be generated by Dr. Walsh (Core B), which he will use to study ischemia-induced angiogenesis (Project 3), and Dr. Cohen (Project 2) atherogenesis in the aorta. We will characterize the effects of exercise in these and in control mice together with Dr. Nathan LeBrasseur, a rodent exercise physiologist. In particular, we will determine how SirT1 and LKB1 downregulation affect the increase in muscle capillary density and the decrease in atherogenic changes in the aorta that typically occur in response to exercise in rodents with a metabolic syndrome phenotype. j - ¿ ^These studies will provide the first information about the function of the SirT1/LKB1/AMPK signaling mechanism in the vasculature. They should also provide novel insights as to whether its dysregulation could be a cause of the endothelial cell dysfunction associated with the metabolic syndrome and a target for its therapy. ,