AMPK as a redox sensor and modulator

AMPK 作为氧化还原传感器和调制器

• 批准号: 7849800
• 负责人: MING-HUI ZOU
• 金额: $ 36.63万
• 依托单位: UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7849800
• 关键词: 3-nitrotyrosine; 5' -AMP-activated protein kinase; Abbreviations; Acetylcholine; Actins; Address; Adenoviruses; Adipocytes; Adverse effects; Aging; Antibodies; Antioxidants; Aorta; Apolipoprotein E; Arachidonic Acids; Arterial Fatty Streak; Atherosclerosis; Biological Assay; Biology; Blood; Blood Vessels; CaM kinase I activator; Cardiovascular Diseases; Cardiovascular system; Catalytic Domain; Cell Adhesion Molecules; Cells; Cerebrum; Cholesterol; Chronic; Coronary; Cyclic GMP; Data; Detection; Development; Diabetes Mellitus; Dimensions; Disease; Dominant-Negative Mutation; E-Selectin; Endothelial Cells; Endothelium; Energy Metabolism; Environmental Tobacco Smoke; Enzymes; Epoprostenol; Equilibrium; Etiology; Eukaryotic Cell; Event; Exhibits; Exposure to; Functional disorder; Generations; Genetic; Glucose; Goals; Health; Hepatocyte; Human; Hypertension; Hypertriglyceridemia; Hypoxia; ICAM1 gene; Incubated; Inflammation; Inflammatory; Ischemic Preconditioning; Knock-out; Knockout Mice; Lesion; Link; Lipid Peroxidation; Lipids; Mediating; Medicine; Metabolism; Metformin; Modeling; Molecular; Motion; Mus; NADPH Oxidase; NG-Nitroarginine Methyl Ester; Nicotine; Nicotinic Receptors; Nitric Oxide; Obesity; Oxidants; Oxidases; Oxidation-Reduction; Peripheral; Peroxonitrite; Phenotype; Phosphotransferases; Physiological; Plasma; Play; Polyethylene Glycols; Process; Production; Prostacyclin synthase; Prostaglandins; Prostaglandins I; Protein Kinase C; Protein-Serine-Threonine Kinases; Rattus; Reaction; Reactive Nitrogen Species; Reactive Oxygen Species; Relative (related person); Relaxation; Resistance; Risk Factors; Role; Signal Transduction; Smoke; Smoker; Smoking; Smooth Muscle; Stimulus; Stress; Superoxide Dismutase; Superoxides; System; Thromboxane Receptor; Transgenic Mice; Triglycerides; Tyrosine; Vascular Cell Adhesion Molecule-1; Western Blotting; Whole Organism; Wild Type Mouse; acetovanillone; arginine methyl ester; atherogenesis; biological adaptation to stress; cardiovascular disorder risk; cardiovascular risk factor; catalase; cell growth regulation; cigarette smoking; cigarette smoking; exposed human population; genetic manipulation; human CYBA protein; human NOS3 protein; hypercholesterolemia; imidazole-4-carboxamide; improved; in vivo; inhibitor/antagonist; insight; mouse model; mutant; neutrophil cytosol factor 67K; novel; overexpression; oxidant stress; polyethylene glycol-superoxide dismutase; premature; prevent; progesterone 11-hemisuccinate-(2-iodohistamine); promoter; public health relevance; response; riboside; sensor; stressor; tetrahydrobiopterin; therapeutic target

DESCRIPTION (provided by applicant): AMPK is a serine/threonine protein kinase involved in the regulation of cellular and organismal metabolism. The AMPK system acts as a sensor of cellular energy status that is conserved in all eukaryotic cells. AMPK has been suggested to serve as an energy gauge in cells, by detecting changes in the ratios of AMP to ATP. The consequences of AMPK activation include inhibition of ATP-consuming processes and activation of an ATP-producing process. The applicant's group was the first to demonstrate that physiologically relevant concentrations of ONOO- increased AMPK activity as well as its downstream enzymes such as eNOS and acetyl Co-A carboxylase (ACC). In addition, we have also demonstrated that ONOO--dependent AMPK activation is operative in hypoxia-reoxygenation, metformin- stimulated endothelial cells, thromboxane receptor stimulation, and nicotine-treated adipocytes. Similarly, we found that nicotine, a major constituent of cigarette smoke, activates AMPK in differentiated 3T3L1 adipocytes in a ROS-dependent fashion. Interestingly, AMPK activation by pharmacological AMPK activators (AICAR and metformin), physiological stimuli (glucose-depletion and osmotic stress), or genetic manipulation (adenoviruses encoding constitutively active AMPK) protects the endothelium against the adverse effects of nicotine. Concomitantly, pharmacological or genetic inhibition of AMPK markedly increased ROS, NF?B, and inflammatory genes (ICAM-1, VCAM-1, and E-selectin) in cultured endothelial cells, differentiated 3T3L1 adipocytes, and cultured primary rat hepatocytes. In parallel, in supporting that AMPK functions as a suppressor of oxidant stress, we have obtained preliminary evidence suggesting that AMPK activation by ischemic preconditioning (IPC) effectively blocked hypoxia/reoxygenation-triggered oxidant stress. The most conclusive evidence that AMPK reduced oxidant stress is that IPC failed to alter both the markers of oxidant stress and endothelial function in the AMPK 11 knockout (KO) mice. Consistently, we have found that compared to the wild types, aortas isolated from AMPK 11 or 12 KO mice exhibited impaired endothelial relaxation together with increased detections of both O2.- and ONOO-. Similarly, there was greater levels of NADPH oxidase subunits including gp91phox (NOX2), NOX-4, p22phox, p47phox, and p67phox together with increased NAD(P)H oxidase activity in AMPK12 KO mice than those in C57BL6 wild types. Importantly, apocynin, a potent NAD(P)H oxidase inhibitor, restored acetylcholine-induced endothelium-dependent relaxation in AMPK 12 KO mice, further suggesting NAD(P)H oxidase is functionally active and is responsible for impaired endothelial function in AMPK 12KO mice. Finally, without altering plasma lipids (cholesterol and triglyceride), the aortas isolated from Apo-E/AMPK11 or AMPK/AMPK 12 dual KO mice exhibited increased detection of oxidant stress markers, increased atherosclerotic lesions, and increased expression of proinflammatory adhesion molecules when compared to those in Apo-E KO mice. The goal of this application is to establish (a) that AMPK, through its ability to respond to very small changes in AMP levels, may be the proximal "oxidant stress-sensor" of the cell; (b) that AMPK activation may trigger physiological responses to suppress processes that generate oxidants (modulator) and or increase anti-oxidant defense systems; and (c) AMPK, via a reduction of oxidant stress, maintains the non-angiogenic, non-inflammatory, and atherosclerosis-resistant phenotypes in vascular cells. PUBLIC HEALTH RELEVANCE: A basic premise of this proposal is that AMPK activation could protect cells and the whole organism against the adverse effects of cardiovascular risk factors such as nicotine by setting in motion events that decrease oxidant stress. For the sake of clarity and being focused of the current application we use nicotine as a pathological stimulus relevant to cardiovascular diseases. As there is growing evidence that oxidant stress is involved in the vascular effects of cardiovascular risk factors including hypercholesterolemia, diabetes, and hypertension, thus, the significance of this proposal is beyond nicotine and will be applicable to other cardiovascular diseases. The current application addresses a fundamental question in biology and medicine, i.e., how oxidant stressors are sensed and modulated in health and disease. Completion of this application, thus, may add an entirely new dimension to the role of AMPK in stress responses and provide an interface between oxidant stress, energy metabolism, and cardiovascular biology. Completion of the proposed studies will provide novel insights into whether AMPK is a potential target for therapy in smoking and common diseases including aging, obesity, diabetes, hypertension, and atherosclerosis.

描述（由申请人提供）：AMPK是一种丝氨酸/苏氨酸蛋白激酶，参与细胞和生物体代谢的调节。 AMPK 系统充当细胞能量状态的传感器，在所有真核细胞中都是保守的。人们建议 AMPK 通过检测 AMP 与 ATP 比率的变化来充当细胞中的能量计。 AMPK 激活的后果包括抑制 ATP 消耗过程和激活 ATP 生成过程。申请人的小组第一个证明了生理相关浓度的ONOO-增加了AMPK及其下游酶如eNOS和乙酰Co-A羧化酶(ACC)的活性。此外，我们还证明ONOO依赖性AMPK激活在缺氧-复氧、二甲双胍刺激的内皮细胞、血栓素受体刺激和尼古丁处理的脂肪细胞中起作用。同样，我们发现尼古丁（香烟烟雾的主要成分）以 ROS 依赖性方式激活分化的 3T3L1 脂肪细胞中的 AMPK。有趣的是，通过药理学 AMPK 激活剂（AICAR 和二甲双胍）、生理刺激（葡萄糖消耗和渗透压）或基因操作（编码组成型活性 AMPK 的腺病毒）来激活 AMPK，可以保护内皮细胞免受尼古丁的不利影响。同时，药理或基因抑制 AMPK 显着增加培养的内皮细胞、分化的 3T3L1 脂肪细胞和培养的原代大鼠肝细胞中的 ROS、NF?B 和炎症基因（ICAM-1、VCAM-1 和 E-选择素）。与此同时，为了支持 AMPK 作为氧化应激抑制剂发挥作用，我们获得了初步证据，表明缺血预处理 (IPC) 激活 AMPK 可以有效阻止缺氧/复氧触发的氧化应激。 AMPK 减少氧化应激的最确凿证据是，IPC 未能改变 AMPK 11 敲除 (KO) 小鼠的氧化应激标志物和内皮功能。我们一致发现，与野生型相比，从 AMPK 11 或 12 KO 小鼠中分离的主动脉表现出内皮舒张受损以及 O2.- 和 ONOO- 检测增加。同样，与 C57BL6 野生型小鼠相比，AMPK12 KO 小鼠中的 NADPH 氧化酶亚基（包括 gp91phox (NOX2)、NOX-4、p22phox、p47phox 和 p67phox）水平更高，并且 NAD(P)H 氧化酶活性增加。重要的是，夹竹桃麻素是一种有效的 NAD(P)H 氧化酶抑制剂，可恢复 AMPK 12 KO 小鼠中乙酰胆碱诱导的内皮依赖性松弛，进一步表明 NAD(P)H 氧化酶具有功能活性，并且是 AMPK 12KO 小鼠中内皮功能受损的原因。最后，在不改变血浆脂质（胆固醇和甘油三酯）的情况下，与相比，从 Apo-E/AMPK11 或 AMPK/AMPK 12 双 KO 小鼠分离的主动脉表现出氧化应激标记物检测增加、动脉粥样硬化病变增加以及促炎粘附分子表达增加与 Apo-E KO 小鼠相比。本申请的目标是确定 (a) AMPK 通过其对 AMP 水平的微小变化做出反应的能力，可能是细胞的近端“氧化应激传感器”； (b) AMPK 激活可能触发生理反应，抑制产生氧化剂（调节剂）和/或增强抗氧化防御系统的过程； (c) AMPK 通过减少氧化应激，维持血管细胞中的非血管生成、非炎症和抗动脉粥样硬化表型。公共健康相关性：该提案的基本前提是 AMPK 激活可以通过启动减少氧化应激的活动来保护细胞和整个有机体免受尼古丁等心血管危险因素的不利影响。为了清楚起见并集中当前应用，我们使用尼古丁作为与心血管疾病相关的病理刺激物。由于越来越多的证据表明氧化应激与包括高胆固醇血症、糖尿病和高血压在内的心血管危险因素的血管效应有关，因此，该提议的意义超越了尼古丁，并将适用于其他心血管疾病。当前的应用解决了生物学和医学中的一个基本问题，即如何在健康和疾病中感知和调节氧化应激源。因此，该申请的完成可能会为 AMPK 在应激反应中的作用增加一个全新的维度，并在氧化应激、能量代谢和心血管生物学之间提供一个接口。拟议研究的完成将为 AMPK 是否是吸烟和常见疾病（包括衰老、肥胖、糖尿病、高血压和动脉粥样硬化）治疗的潜在靶点提供新的见解。