Botanical Phenolics on Oxidative/Nitrosative Signaling Pathways/Grace Sun

植物酚类物质对氧化/亚硝化信号通路的影响/Grace Sun

• 批准号: 8007165
• 负责人: DENNIS B LUBAHN
• 金额: $ 35.62万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8007165
• 关键词: Adult; Affect; Age; Agonist; Animal Nutrition; Animal Testing; Animals; Anti-Inflammatory Agents; Anti-inflammatory; Antioxidants; Area; Arts; Astrocytes; Behavioral; Bioinformatics; Biological Availability; Botanicals; Brain; Brain Injuries; Brain region; Cause of Death; Cell Survival; Cells; Cerebral Ischemia; Cerebrum; Characteristics; Core Facility; Coupling; Crude Extracts; Cysteine; Data; Diet; Dietary Supplementation; Disease; Epidemiologic Studies; Erinaceidae; Estrogen Receptors; Estrogens; Europe; Event; Figs - dietary; Filament; Fruit; Gene Expression; Genomics; Glutamate Receptor; Goals; Health; Homeostasis; Human; In Vitro; Infarction; Inflammatory; Inflammatory Response; Injury; Knockout Mice; Light; Link; Measures; Mediating; Messenger RNA; Microglia; Middle Cerebral Artery Occlusion; Modeling; Molecular; Mus; N-Methylaspartate; NADPH Oxidase; Neurodegenerative Disorders; Neuroglia; Neurons; Nitric Oxide; Nitric Oxide Signaling Pathway; North America; Outcome; Oxidation-Reduction; Pathway Analysis; Pathway interactions; Physiological; Plants; Post-Translational Protein Processing; Process; Production; Proteins; Proteomics; Protocols documentation; Public Health; Reaction; Reactive Oxygen Species; Regimen; Reperfusion Injury; Reperfusion Therapy; Research; Risk; Role; Scheme; Secondary to; Signal Pathway; Signal Transduction; Signaling Molecule; Small Interfering RNA; Source; Stroke; Stroke prevention; Sulfhydryl Compounds; Supplementation; Technology; Testing; The Sun; Transgenic Organisms; Tyrosine; adduct; base; brain cell; brain tissue; cell type; combat; cytokine; dietary supplements; disability; excitotoxicity; experience; feeding; human NOS2A protein; human disease; in vivo; inhibitor/antagonist; insight; neurochemistry; neuronal survival; nitrosative stress; novel; polyphenol; protective effect; protein structure function; response

Excessive oxidafive and nitrosative (O/N) activifies in the brain have been recognized as the basis for the neuronal excitotoxicity and glial cell inflammatory responses that underiie many neurodegenerative diseases, including stroke. Recent studies have discovered the role of NADPH oxidase as an important source of reactive oxygen species (ROS) in brain cells. Activafion of NADPH oxidase is associated with a number of downstream signaling pathways, including the NF-KB pathway that induces INOS and produces nitric oxide (NO). Among other reactions, NO can regulate protein structure and function by interacfing with protein reactive cysteine thiol residues to form S-nitrosothiol. Many proteins associated with NADPH oxidase signaling pathways appear to be modified by S-nitrosylation. While the mechanisms are unknown, there is considerable evidence to suggest that a number of botanical compounds may protect the brain against O/N insults, including those resulfing from cerebral ischemia. In keeping with the central goal ofthis Center, Project 2 will identify new botanicals and test the hypothesis that stroke-mediated neuronal excitotoxicity and glial inflammatory responses are due in part to ROS from NADPH oxidase and subsequent S-nitrosylafion of key proteins in the NADPH oxidase signaling pathways. In Aim 1. neurons and astrocytes/microglia will be used to screen botanical extracts and pure compounds isolated from the extracts for their ability to suppress NMDA-mediated neuronal excitotoxicity and cytokine-induced inflammatory responses in glial cells. The effects of botanicals on S-nitrosylation of proteins and differentially-expressed mRNA and proteins in neurons and glial cells will be invesfigated using the newly developed NitroDIGE protocol and mRNA megasequencing facilities in the Interactions Core. In Aim 2, the mouse focal cerebral ischemia model induced by occlusion of the middle cerebral artery will be used to investigate whether dietary supplementation with specific botanicals offers protective effects against cerebral ischemia/reperfusion damage. These studies will include using mice deficient in N0X2 and iNOS to test the effects of botanicals on behavioral outcome, neuronal survival, glial activation, and inflammatory responses. Aim 3 will test the hypothesis that identified botanicals offers neuroprotective effects by reducing cerebral ischemia/reperfusion damage through activation of the Nrf2/Keap1 antioxidant signaling pathway. In these studies, astrocyte- or neuron-targeted conditional Keapl-null mice will be used. In Aims 2 and 3, we will also identify changes in S-nitrosylated and inflammatory proteins and mRNA associated with oxidafive and anfi-oxidafive signaling pathways in disfinct brain regions. The in vitro and in vivo approaches of Project 2, together with support from the unique and state-of-the-art core facilities of this Center, will combine to identify specific cellular and molecular mechanisms through which novel botanicals and/or their active components may exert beneficial effects and protect against stroke damage.

大脑中过度的氧化和亚硝化（O/N）激活已被认为是导致许多神经退行性疾病（包括中风）的神经元兴奋毒性和神经胶质细胞炎症反应的基础。最近的研究发现 NADPH 氧化酶作为脑细胞中活性氧 (ROS) 的重要来源。 NADPH 氧化酶的激活与许多下游信号通路相关，包括诱导 INOS 并产生一氧化氮 (NO) 的 NF-KB 通路。在其他反应中，NO 可以通过与蛋白质反应性半胱氨酸硫醇残基相互作用形成 S-亚硝基硫醇来调节蛋白质结构和功能。许多与 NADPH 氧化酶信号通路相关的蛋白质似乎都经过 S-亚硝基化修饰。虽然其机制尚不清楚，但有大量证据表明，许多植物化合物可以保护大脑免受 O/N 损伤，包括脑缺血引起的损伤。为了与该中心的中心目标保持一致，项目 2 将鉴定新的植物药并测试以下假设：中风介导的神经元兴奋性毒性和神经胶质炎症反应部分归因于 NADPH 氧化酶的 ROS 以及随后 NADPH 氧化酶信号通路中关键蛋白的 S-亚硝基化。在目标 1 中，神经元和星形胶质细胞/小胶质细胞将用于筛选植物提取物和从提取物中分离的纯化合物，以确定它们抑制神经胶质细胞中 NMDA 介导的神经元兴奋性毒性和细胞因子诱导的炎症反应的能力。将使用 Interactions Core 中新开发的 NitroDIGE 协议和 mRNA 巨测序设施来研究植物药对蛋白质 S-亚硝基化以及神经元和神经胶质细胞中差异表达 mRNA 和蛋白质的影响。在目标2中，将利用大脑中动脉闭塞诱导的小鼠局灶性脑缺血模型来研究膳食补充剂是否对脑缺血/再灌注损伤具有保护作用。这些研究将包括使用缺乏 N0X2 和 iNOS 的小鼠来测试植物药对行为结果、神经元存活、神经胶质激活和炎症反应的影响。目标 3 将检验以下假设：已确定的植物药通过激活 Nrf2/Keap1 抗氧化信号通路，减少脑缺血/再灌注损伤，从而提供神经保护作用。在这些研究中，将使用星形胶质细胞或神经元靶向的条件性 Keapl 缺失小鼠。在目标 2 和 3 中，我们还将鉴定与不连续大脑区域中的氧化和抗氧化信号通路相关的 S-亚硝基化和炎症蛋白以及 mRNA 的变化。项目2的体外和体内方法，加上该中心独特且最先进的核心设施的支持，将结合起来确定特定的细胞和分子机制，通过这些机制，新型植物药和/或其活性成分可以发挥有益作用并预防中风损伤。