Project 5: ROS, Glutathione and Vascular Response to Diesel Exhaust

项目 5：ROS、谷胱甘肽和柴油机尾气的血管反应

• 批准号: 7328339
• 负责人: Terrance J Kavanagh
• 金额: $ 29.18万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7328339
• 关键词: Air Pollution; Angiotensin II; Antioxidants; Apoptosis; Binding; Biochemical; Blood Vessels; Candidate Disease Gene; Cardiac Myocytes; Cardiovascular Diseases; Cardiovascular Physiology; Cardiovascular system; Catalytic Domain; Cells; Chromosome Mapping; Consumption; Cultured Cells; Cysteine; Diesel Exhaust; Doctor of Philosophy; Doppler Ultrasound; Endothelial Cells; Endothelin-1; Endothelium; Enzymes; Event; Exposure to; Functional disorder; GCLC gene; GCLM gene; Gene Expression; Generations; Genes; Genetic; Genetic Determinism; Genetic Polymorphism; Glutamate-Cysteine Ligase; Glutamates; Glutathione; Glycine; Heat shock proteins; Homeostasis; Human; Hydrogen Peroxide; Hydroxyl Radical; In Vitro; Inbred Strains Mice; Inflammatory; Ischemia; Ligase; Ligase Gene; Lipid Peroxides; Maps; Metabolic Control; Metabolism; Microscopy; Mitochondria; Mus; Myocardial Infarction; NADPH Oxidase; Nitric Oxide; Nitric Oxide Synthase; Oxidation-Reduction; Oxidative Stress; Performance; Peroxonitrite; Predisposition; Preparation; Process; Production; Proteins; Quantitative Trait Loci; Rate; Reactive Oxygen Species; Recombinant Inbred Strain; Regulation; Risk; Risk Factors; Role; Serum; Signal Transduction; Single Nucleotide Polymorphism; Smooth Muscle Myocytes; Sulfhydryl Compounds; Superoxides; Thrombosis; Transgenic Organisms; Ultrasonics; Umbilical vein; Vasoconstrictor Agents; Vasodilation; Vasodilator Agents; Vasomotor; aqueous; cell injury; cell type; chemokine; cofactor; comparative; cytokine; cytotoxicity; exhaust; in vivo; interest; mouse model; nitration; oxidation; particle; prevent; programs; protein function; response; stress protein; tetrahydrobiopterin; tool; trafficking; vasoconstriction

Exposure to traffic-related air pollution such as diesel exhaust (DE) is associated with an increase in the level of reactive oxygen species (ROS) in multiple cell types. These ROS include the superoxide anion radical, hydrogen peroxide, lipid peroxides and hydroxyl radical. ROS can have deleterious effects on cells, but they are also known to be involved in normal transmembrane signaling and metabolic control in the cardiovascular system. For instance, two potent vasoconstrictors, angiotensin II and endothelin-1, can increase production of ROS by smooth muscle cell NADPH oxidase, thus causing an increase in the intracellular concentrations of superoxide and hydrogen peroxide. Superoxide can bind to nitric oxide (NO) to form peroxynitrite, which can cause cellular injury, but also limit the amount of NO available for vasorelaxation. ROS generated in this way can also oxidize tetrahydrobiopterin, a necessary cofactor for NO synthase (NOS), resulting in uncoupling of NOS and production of more superoxide and peroxynitrite. Consumption of NO by superoxide in this way can result in vasoconstriction. ROS are thus able to influence cardiovascular performance and vascular reactivity. Glutathione (GSH) is an abundant non-protein thiol which is a potent scavenger of ROS, including hydrogen peroxide, lipid peroxides, and importantly, peroxitrite (ONOO-). GSH is a tripeptide thiol present in millimolar concentrations in most cells. The first and rate-limiting step in GSH synthesis is carried out by the enzyme glutamate cysteine ligase (GCL), which is composed of two subunits, a catalytic subunit (GCLC) and a modifier or regulatory subunit (GCLM). Importantly, single nucleotide polymorphisms (SNPs) in both GCLC and GCLM have been shown to be important in myocardial infarction, as well as controlling vascular reactivity in humans. In this project, we will investigate the reasons for this effect of GCL by using a comparative approach. We will 1) use mouse models of differential GCL expression and activity to investigate its role in DE-induced changes in vascular reactivity 2), use cultured endothelial cells from these mice and from humans to investigate the underlying biochemical events responsible for GSH and DE-induced modulation of endothelial NO production; and 3) characterize genetically defined inbred strains of mice that have variability in their vascular responses to DE, and identify single nucleotide polymorphisms (SNPs) and quantitative trait loci that are associated with these changes in mice. Using these tools we will define the role of GSH synthesis in DE-induced changes in vascular reactivity, investigate the underlying pathophysiological mechanisms, and map and identify genetic determinants of DE-induced changes in vascular reactivity. These candidate genes will then be referred to Projects 1 and 2 where they will be further evaluated as potential genetic factors in DE-induced vascular abnormalities in humans.

暴露于与交通相关的空气污染（例如柴油排气（DE））与增加有关 多种细胞类型中的活性氧（ROS）水平。这些ROS包括超氧化阴离子 自由基，过氧化氢，脂质过氧化物和羟基自由基。 ROS会对细胞产生有害影响， 但已知它们参与了正常的跨膜信号传导和代谢控制 心血管系统。例如，两个有效的血管收缩剂，血管紧张素II和内皮素1，可以 通过平滑肌细胞NADPH氧化酶增加ROS的产生，从而导致增加 超氧化物和过氧化氢的细胞内浓度。超氧化物可以与一氧化氮结合（NO） 形成过氧亚硝酸盐，这可能会导致细胞损伤，但也限制了无法进行血管征的量。 以这种方式产生的ROS还可以氧化四氢无生物蛋白，这是一个必要的辅助因子 合酶（NOS），导致NOS的解偶联以及产生更多的超氧化物和过氧亚硝酸盐。 以这种方式消耗超氧化物可以导致血管收缩。 ROS因此能够影响 心血管表现和血管反应性。谷胱甘肽（GSH）是一种丰富的非蛋白硫醇 这是对ROS的有效清除，包括过氧化氢，脂质过氧化物，重要的是， 过氧石（Onoo-）。 GSH是大多数细胞中以毫米浓度存在的三肽硫醇。第一个和 GSH合成中的速率限制步骤是通过谷氨酸半胱氨酸连接酶（GCL）进行的，这是 由两个亚基组成，一个催化亚基（GCLC）和一个修饰符或调节亚基（GCLM）。 重要的是，GCLC和GCLM中的单核苷酸多态性（SNP）已证明为 在心肌梗塞以及控制人类的血管反应性中很重要。在这个项目中，我们将 通过使用比较方法研究GCL的这种效果的原因。我们将1）使用鼠标 差异GCL表达和活性的模型，以研究其在血管变化中的作用 反应性2），使用来自这些小鼠和人类的培养的内皮细胞来研究潜在的细胞 造成GSH的生化事件，并导致对内皮无生产的调节； 3） 表征具有遗传定义的小鼠的近交菌株，这些小鼠的血管反应变异性变异 并鉴定与这些相关的单核苷酸多态性（SNP）和定量性状基因座 小鼠的变化。使用这些工具，我们将定义GSH综合在DE诱导的变化中的作用 血管反应性，研究潜在的病理生理机制，并绘制并识别遗传 决定性诱导的血管反应性变化的决定因素。然后将这些候选基因转到 项目1和2将它们进一步评估为脱离诱导血管的潜在遗传因素 人类异常。