OKHSC COBRE: ACTIVATION OF AMPK PROTECTS AGAINST DIABETIC VASCULAR COMPLICATIONS

OKHSC COBRE：激活 AMPK 可预防糖尿病血管并发症

• 批准号: 8360279
• 负责人: Zhonglin Xie
• 金额: $ 21.25万
• 依托单位: UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2012-09-09
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8360279
• 关键词: Adverse effects; Animal Model; Antioxidants; Arterial Fatty Streak; Biological Assay; Cell Adhesion Molecules; Cells; Centers of Research Excellence; Diabetes Mellitus; Diabetic Angiopathies; Endothelial Cells; Functional disorder; Funding; Glucose; Grant; Hyperglycemia; Incubated; Ischemic Preconditioning; Lead; Lipid Peroxidation; Measurement; Mentors; Monitor; Mus; National Center for Research Resources; Nitric Oxide; Nonesterified Fatty Acids; Oklahoma; Pathway interactions; Peroxonitrite; Prevention; Principal Investigator; Prostacyclin synthase; Research; Research Infrastructure; Resources; Source; Superoxide Dismutase; Time; Tyrosine; United States National Institutes of Health; Vascular Diseases; Work; cost; diabetic; in vivo; insight; macrovascular disease; nitration; overexpression; oxidant stress; prevent

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. It has been well established that diabetes leads to microvascular complications, and accelerates macrovascular diseases. One feature of diabetes is excessive oxidant stress, which impairs nitric oxide (NO) bioactivity, increase adhesion molecule expression, and promote atherosclerotic lesion formation. Preliminary studies demonstrate that hyperglycemia and free fatty acids (FFA) impart an oxidant stress in endothelial cells, resulting in lipid peroxidation, tyrosine nitration of prostacyclin synthase (PGIS), and endothelial dysfunction; Treatment of cells with either AICAR or with adenoviruse overexpressing constitutively active AMPK prevents the increased O2.-, inactivation of NO, PGIS nitration and endothelial dysfunction; Activation of AMPK by ischemic preconditioning (IPC) effectively blocked the markers of oxidant stress, likely via over expression of UCP-2. The most conclusive evidence that AMPK reduced oxidant stress is that (IPC) failed to alter both the markers of oxidant stress and UCP-2 expression in the AMPK-KO mice. We hypothesize that AMPK activation could protect the endothelial cell against the adverse effects of hyperglycemia and FFA by increasing antioxidant potentials (UCP-2 and/or superoxide dismutase) that lead to a decrease in oxidant stress and increase in NO bioactivity. This hypothesis will be pursued in two specific aims. 1) Determine if activation of AMPK reduces oxidant stress and endothelial dysfunction induced by hyperglycemia and FFA, and evaluate how it works. HAEC will be incubated with glucose and FFA, the markers of oxidant stress such as (O2.-), peroxynitrite, UCP2 expression, NO bioactivity and expression of adhesion molecules will be monitored under conditions in which AMPK activity is altered by using AICAR or adenoviral constructs. 2) Determine if AMPK-dependent reduction in oxidant stress and endothelial dysfunction is operating in vivo. Wild type and AMPK alpha 1 and alpha 2 KO mice will be used to make diabetic animal model. Aortic rings will be assayed or assayed after being incubated for various time in hyperglycemia/FFA media by measurements of functional parameters UCP-2 expression and selected parameters of ONOOprostacyclin synthase pathway. The proposed studies will provide insight into the mechanism by which diabetes leads to vascular diseases and may lead to use activation of AMPK as an important target for the prevention and treatment of diabetic vascular complications.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的主要研究者可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 众所周知，糖尿病会导致微血管并发症，并加速大血管疾病。糖尿病的一个特征是过度的氧化应激，这会损害一氧化氮（NO）的生物活性，增加粘附分子的表达，并促进动脉粥样硬化病变的形成。 初步研究表明，高血糖和游离脂肪酸（FFA）会给内皮细胞带来氧化应激，导致脂质过氧化、前列环素合酶（PGIS）酪氨酸硝化和内皮功能障碍；用 AICAR 或过表达组成型活性 AMPK 的腺病毒处理细胞可防止 O2.- 增加、NO 失活、PGIS 硝化和内皮功能障碍；缺血预处理 (IPC) 激活 AMPK 可以有效阻断氧化应激标志物，这可能是通过 UCP-2 的过度表达实现的。 AMPK 减少氧化应激的最确凿证据是 (IPC) 未能改变 AMPK-KO 小鼠中氧化应激标记物和 UCP-2 表达。 我们假设 AMPK 激活可以通过增加抗氧化潜力（UCP-2 和/或超氧化物歧化酶）来保护内皮细胞免受高血糖和 FFA 的不利影响，从而导致氧化应激减少和 NO 生物活性增加。这一假设将有两个具体目标。 1) 确定 AMPK 的激活是否会减少氧化应激和内皮功能障碍引起的 高血糖和 FFA，并评估其工作原理。 HAEC 将与葡萄糖和 FFA 一起孵育，在使用 AICAR 或改变 AMPK 活性的条件下，将监测氧化应激标志物，例如 (O2.-)、过氧亚硝酸盐、UCP2 表达、NO 生物活性和粘附分子表达。 腺病毒构建体。 2) 确定 AMPK 依赖性氧化应激和内皮功能障碍的减少是否在体内发挥作用。野生型和AMPK α1和α2 KO小鼠将用于制作糖尿病动物模型。主动脉环将通过测量UCP-2表达的功能参数和ONOO前列环素合酶途径的选定参数来测定或在高血糖/FFA培养基中温育不同时间后进行测定。拟议的研究将深入了解糖尿病导致血管疾病的机制，并可能导致使用 AMPK 的激活作为预防和治疗糖尿病血管并发症的重要靶点。