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.

该副本是利用资源的众多研究子项目之一 由NIH/NCRR资助的中心赠款提供。对该子弹的主要支持 而且，副投影的主要研究员可能是其他来源提供的 包括其他NIH来源。 列出的总费用可能 代表subproject使用的中心基础架构的估计量， NCRR赠款不直接向子弹或副本人员提供的直接资金。 已经很好地确定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氧化应激，会损害一氧化氮（NO）生物活性，增加粘附分子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）在内皮细胞中赋予氧化剂应激，从而导致脂质过氧化，前列环素合酶（PGIS）的酪氨酸硝化和内皮功能障碍；用AICAR或腺病毒过表达的组成性AMPK治疗细胞，可防止O2.-的增加，灭活NO，PGIS硝化和内皮功能障碍；通过缺血性预处理（IPC）激活AMPK，有效地阻断了氧化应激的标记，这可能是通过UCP-2的过度表达来阻断的。 AMPK减少氧化剂应激的最终证据是（IPC）未能改变AMPK-KO小鼠中氧化应激和UCP-2表达的标记。 我们假设AMPK激活可以通过增加抗氧化电位（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，从而导致氧化应激的降低并无生物活性增加。该假设将在两个具体目标中提出。 1）确定AMPK的激活是否会减少由 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，氧化应激的标志物，例如（O2.-），过氧亚硝酸盐，UCP2表达，在使用AICAR或使用AICAR或使用AICK活性改变的条件下，将监测粘附分子的生物活性和粘附分子的表达 腺病毒构建体。 2）确定氧化应激和内皮功能障碍的AMPK依赖性降低是否在体内工作。野生型和AMPK Alpha 1和Alpha 2 KO小鼠将用于制造糖尿病动物模型。通过测量功能参数UCP-2表达和Onoo Prostacyclin合酶途径的选定参数，将在高血糖/FFA培养基中孵育多个时间后，将分析或测定主动脉环。拟议的研究将洞悉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标。