Mechanisms of Embryo Response to Oxidative Stress

胚胎对氧化应激的反应机制

• 批准号: 7655110
• 负责人: Mark E Hahn
• 金额: $ 37.62万
• 依托单位: WOODS HOLE OCEANOGRAPHIC INSTITUTION
• 依托单位国家: 美国
• 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/7655110
• 关键词: Affect; Animals; Antioxidants; Basic Science; Binding; Biological Models; Cardiovascular system; 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; 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

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: Oxidative stress is involved in a variety of environmentally influenced human diseases ranging from chemical teratogenesis to cardiovascular and neurodegenerative diseases. The proposed research will enhance our basic understanding of how some chemicals interfere with embryonic development by generating oxidative stress and how embryos can mitigate these effects. This research will i) determine the basal expression and inducibility of anti-oxidant defenses during embryonic development, ii) elucidate fundamental mechanisms of the response of vertebrate embryos to oxidative stress, iii) establish a model system for screening chemicals for activity as developmental toxicants or antioxidants, and iv) provide a mechanistic foundation that will facilitate the extrapolation of results obtained in zebrafish to humans, supporting risk assessment and providing insight into the role of oxidative stress in human disease.

描述（由申请人提供）：环境暴露导致的氧化应激与多种人类疾病相关，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。抗氧化防御的表达和诱导是影响这些早期生命阶段对氧化剂敏感性的关键因素，但这些反应在胚胎中的个体发育尚未得到很好的理解。在成年动物中，氧化剂通过激活NF-E2相关因子2（NRF 2）和相关蛋白质启动抗氧化反应，NRF 2和相关蛋白质与抗氧化反应元件结合并激活谷胱甘肽S-转移酶、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等基因的转录。本研究的总体目标是阐明脊椎动物胚胎在发育过程中对氧化应激的反应机制。我们将测试的核心假设，对氧化应激的反应和一套调控基因在发展过程中的变化。由于这些发育差异，某些阶段可能对氧化应激引起的损伤更敏感。这些研究将在体内使用斑马鱼（Danio rerio）胚胎进行，这是一种有价值的模型，可用于检查发育中动物的毒性机制，并筛选化学品的发育毒性。目的1将使用转录谱和表型锚定来鉴定构成胚胎中氧化应激反应的核心基因集，确定核心氧化剂反应基因集的反应性和组成如何随胚胎发育阶段而变化，并确定暴露于结构和机制不同的NRF 2激活剂的胚胎中氧化应激反应的时间和基因谱如何不同。（tBHQ、敌草快、萝卜硫素）。目的2将阐明不同的NRF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中的作用，使用靶向敲低NRF蛋白合成与吗啉代寡核苷酸。目的3通过计算和体内实验分析氧化反应基因启动子，建立抗氧化反应基因在胚胎发育过程中的调控机制，从而产生稳定的表达报告基因（GFP）的转基因斑马鱼品系，以响应氧化应激。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。这些研究的结果将建立在脊椎动物胚胎中的氧化应激的转录反应的组成和个体发生，阐明这种反应的基本机制，产生筛选化学品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。公共卫生相关性：氧化应激参与了从化学致畸到心血管和神经退行性疾病的各种环境影响的人类疾病。这项拟议中的研究将增强我们对一些化学物质如何通过产生氧化应激干扰胚胎发育以及胚胎如何减轻这些影响的基本理解。这项研究将i）确定胚胎发育过程中抗氧化防御的基础表达和诱导，ii）阐明脊椎动物胚胎对氧化应激反应的基本机制，iii）建立筛选化学品作为发育毒物或抗氧化剂活性的模型系统，iv）提供机制基础，促进将斑马鱼中获得的结果外推到人类，支持风险评估，并深入了解氧化应激在人类疾病中的作用。