Mechanisms of Embryo Response to Oxidative Stress

胚胎对氧化应激的反应机制

基本信息

批准号：
8244524
负责人：
Mark E Hahn
金额：
$ 37.08万
依托单位：
WOODS HOLE OCEANOGRAPHIC INSTITUTION
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-06-15 至 2014-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8244524
关键词：
Affect Animals Antioxidants Basic Science Binding Biological Models Cardiovascular Diseases Chemicals Cysteine Development Diquat Disease Drug or chemical Tissue Distribution Embryo Embryonic Development Environmental Exposure Family Fishes Foundations Gene Expression Generations Genes Genetic Transcription Genomics Glutamate-Cysteine Ligase Glutathione Glutathione S-Transferase Growth Homologous Gene Human Individual Life Ligase Link Mammals Mediating Modeling Molecular NAD(P)H dehydrogenase (quinone) 1, human NF-E2-related factor 2 NQO1 gene Neurodegenerative Disorders Oligonucleotides Oxidants Oxidative Stress Phenotype Predisposition Production Protein Biosynthesis Protein Family Protein Isoforms Proteins Reactive Oxygen Species Regulation Regulatory Element Reporter Genes Research Response Elements Reverse Transcriptase Polymerase Chain Reaction Risk Assessment Role Screening procedure Staging Sulforaphane Superoxide Dismutase Testing Time Toxic effect Transgenes Transgenic Organisms Zebrafish abstracting bZIP Domain biological adaptation to stress experimental analysis human disease in vivo insight knock-down mature animal member paralogous gene promoter public health relevance research study response tool toxicant

项目摘要

Project Summary/Abstract Oxidative stress resulting from environmental exposures is associated with a variety of human diseases ranging from chemical teratogenesis to cardiovascular and neurodegenerative diseases. Developing animals appear to be especially sensitive to chemicals causing oxidative stress. The expression and inducibility of anti- oxidant defenses are critical factors affecting susceptibility to oxidants at these early life stages, but the ontogenic development of these responses in embryos is not well understood. In adult animals, oxidants initiate an anti-oxidant response by activating NF-E2-related factor 2 (NRF2) and related proteins, which bind to the anti-oxidant response element and activate transcription of genes such as glutathione S-transferases, NAD(P)H-quinone oxidoreductase, glutamyl-cysteine ligase, and superoxide dismutase. The overall objective of the research proposed here is to elucidate the mechanisms by which vertebrate embryos respond to oxidative stress during development. We will test the central hypothesis that responsiveness to oxidative stress and the set of regulated genes vary during development. Because of these developmental differences, some stages may be more sensitive to oxidative stress-induced damage. These studies will be performed in vivo using embryos of the zebrafish (Danio rerio), a valuable model in which to examine mechanisms of toxicity in developing animals and to screen chemicals for developmental toxicity. Aim 1 will use transcriptional profiling and phenotypic anchoring to identify the core set of genes that comprise the oxidative stress response in embryos, establish how the responsiveness and composition of the core set of oxidant-responsive genes vary with developmental stage of embryos, and determine how the timing and gene profile of the oxidative stress response differ in embryos exposed to structurally and mechanistically distinct activators of NRF2 (tBHQ, diquat, sulforaphane). Aim 2 will elucidate the roles of different NRF paralogs in the transcriptional response to oxidative stress during development in embryos in vivo, using targeted knock-down of NRF protein synthesis with morpholino oligonucleotides. Aim 3 will establish the mechanism of regulation of anti-oxidant response genes during embryonic development through computational and in vivo experimental analysis of oxidant-responsive gene promoters, leading to the generation of a stable line of transgenic zebrafish expressing a reporter gene (GFP) in response to oxidative stress. Finally, we will test a set of mammalian developmental toxicants for the ability to activate the transgene in embryos. The results of these studies will establish the composition and ontogeny of the transcriptional response to oxidative stress in vertebrate embryos, elucidate fundamental mechanisms underlying this response, generate tools for screening chemicals for activity as developmental toxicants or antioxidants, and provide insight into the role of oxidative stress in human disease.

项目概要/摘要环境暴露引起的氧化应激与多种人类疾病有关从化学致畸到心血管和神经退行性疾病。发育中的动物似乎对引起氧化应激的化学物质特别敏感。抗-的表达和诱导氧化防御是影响生命早期阶段对氧化剂敏感性的关键因素，但胚胎中这些反应的个体发育尚不清楚。在成年动物中，氧化剂通过激活 NF-E2 相关因子 2 (NRF2) 和相关蛋白来启动抗氧化反应，这些蛋白与抗氧化反应元件并激活谷胱甘肽 S-转移酶等基因的转录， NAD(P)H-醌氧化还原酶、谷氨酰半胱氨酸连接酶和超氧化物歧化酶。总体目标这里提出的研究的目的是阐明脊椎动物胚胎对发育过程中的氧化应激。我们将测试中心假设，即对氧化应激的反应并且受调控的基因组在发育过程中会发生变化。由于这些发育差异，一些阶段可能对氧化应激引起的损伤更敏感。这些研究将在体内进行使用斑马鱼（Danio rerio）的胚胎，这是一个有价值的模型，可用于检查斑马鱼的毒性机制发育中的动物并筛选化学品的发育毒性。目标 1 将使用转录分析和表型锚定来识别核心基因集包括胚胎中的氧化应激反应，确定氧化应激反应的反应性和组成如何氧化反应基因的核心组随胚胎发育阶段的不同而变化，并决定了其发生的时间暴露于结构和机制的胚胎中氧化应激反应的基因谱和基因谱有所不同 NRF2 的不同激活剂（tBHQ、敌草快、萝卜硫素）。目标 2 将阐明不同 NRF 的作用使用旁系同源物在体内胚胎发育过程中对氧化应激的转录反应使用吗啉代寡核苷酸靶向抑制 NRF 蛋白质合成。目标 3 将建立通过计算调节胚胎发育过程中抗氧化反应基因的机制以及氧化反应基因启动子的体内实验分析，从而产生稳定的表达报告基因（GFP）以应对氧化应激的转基因斑马鱼品系。最后，我们将测试一组哺乳动物发育毒物激活胚胎中转基因的能力。这些研究的结果将确定转录反应的组成和个体发育脊椎动物胚胎中的氧化应激，阐明这种反应的基本机制，产生用于筛选化学物质作为发育毒物或抗氧化剂的活性的工具，并提供深入了解氧化应激在人类疾病中的作用。