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，参与细胞和机体代谢的调节。AMPK系统作为细胞能量状态的传感器，在所有真核细胞中都是保守的。AMPK通过检测AMP与ATP比率的变化，被认为是细胞中的能量测量器。AMPK激活的结果包括抑制atp消耗过程和激活atp产生过程。申请人的小组是第一个证明生理相关浓度的ONOO-增加AMPK活性及其下游酶，如eNOS和乙酰辅酶- a羧化酶（ACC）。此外，我们还证明了ONOO依赖的AMPK激活在缺氧再氧化、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是有效的。同样，我们发现尼古丁（香烟烟雾的主要成分）以ros依赖的方式激活分化3T3L1脂肪细胞中的AMPK。有趣的是，AMPK通过药理学激活剂（AICAR和二甲双胍）、生理刺激（葡萄糖消耗和渗透应激）或基因操作（腺病毒编码构成活性AMPK）激活，可以保护内皮细胞免受尼古丁的不良影响。同时，AMPK的药理学或遗传抑制显著增加ROS、NF？B和炎性基因（ICAM-1、VCAM-1和e -选择素）在培养的内皮细胞、分化的3T3L1脂肪细胞和培养的原代大鼠肝细胞中表达。同时，为了支持AMPK作为氧化应激抑制因子的功能，我们已经获得了初步证据，表明缺血预处理（IPC）激活AMPK可以有效阻断缺氧/再氧化引发的氧化应激。AMPK降低氧化应激的最确凿证据是，IPC未能改变AMPK 11敲除（KO）小鼠的氧化应激和内皮功能标记。我们一致地发现，与野生型相比，从AMPK 11或12 KO小鼠分离的主动脉内皮松弛受损，同时两种O2的检测增加。-和ONOO-。同样，与C57BL6野生型相比，AMPK12 KO小鼠的NADPH氧化酶亚基gp91phox （NOX2）、NOX-4、p22phox、p47phox和p67phox水平更高，NAD(P)H氧化酶活性也更高。重要的是，罗布宁是一种有效的NAD(P)H氧化酶抑制剂，可恢复乙酰胆碱诱导的AMPK 12KO小鼠内皮依赖性松弛，进一步表明NAD(P)H氧化酶具有功能活性，是AMPK 12KO小鼠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apoo - e KO小鼠相比，从Apo-E/AMPK11或AMPK/AMPK 12双KO小鼠分离的主动脉显示出氧化应激标志物的检测增加，动脉粥样硬化病变增加，促炎粘附分子的表达增加。本应用的目标是建立(a) AMPK，通过其对AMP水平非常小的变化作出反应的能力，可能是细胞的近端“氧化应激传感器”；(b) AMPK激活可能引发生理反应，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，维持血管细胞的非血管生成、非炎症和抗动脉粥样硬化表型。公共卫生相关性：该建议的一个基本前提是AMPK激活可以通过设置减少氧化应激的运动事件来保护细胞和整个生物体免受心血管危险因素（如尼古丁）的不利影响。为了明确和关注当前的应用，我们使用尼古丁作为与心血管疾病相关的病理刺激。由于越来越多的证据表明，氧化应激参与了包括高胆固醇血症、糖尿病和高血压在内的心血管危险因素的血管效应，因此，这一建议的意义将超越尼古丁，并将适用于其他心血管疾病。目前的应用解决了生物学和医学中的一个基本问题，即在健康和疾病中如何感知和调节氧化应激源。因此，这一应用的完成可能会为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。上述研究的完成将为AMPK是否为吸烟和常见疾病（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。