Differential Sensitivity to Oxidative Stress during Embryonic Development

胚胎发育过程中对氧化应激的不同敏感性

基本信息

批准号：
8097330
负责人：
Alicia R Timme-Laragy
金额：
$ 5.3万
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：
2009-07-01 至 2012-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8097330
关键词：
Address Affect Antioxidants Area Blood flow Calcium Calcium Signaling Cardiac Cardiotoxicity Cell physiology Cells Chemicals Cocaine Detection Development Diquat Disease Embryo Embryonic Development Enzymes Ethanol Exposure to Extravasation Fertilization Fiber Optics Fluorescent Probes Foundations Gene Expression Profile Generations Genes Genetic Techniques Glutathione Glutathione Disulfide Government Agencies Heart Heart Rate Hour Human Hydrogen Peroxide Hydroquinones Image Individual Industry Learning Measurement Measures Microelectrodes Modeling Morphology Movement National Institute of Environmental Health Sciences Outer Mitochondrial Membrane Oxidants Oxidation-Reduction Oxidative Stress Oxygen Oxygen Consumption Pattern Pesticides Pharmacologic Substance Physiological Population Dynamics Reactive Oxygen Species Research Resistance Resolution Respiration Response Elements Role Screening procedure Signal Transduction Source Specificity Staging Stem cells Sulforaphane Superoxides System Techniques Teratogens Testing Time Toxic effect Translational Research Translations United States National Institutes of Health Vertebrates Xenobiotics Zebrafish biological adaptation to stress catalase developmental toxicology gain of function hydroquinone interdisciplinary approach interest multidisciplinary nanoparticle optical imaging oxidative damage research study transcription factor

项目摘要

DESCRIPTION (provided by applicant): The embryonic development of vertebrates is highly susceptible to disruption by exposure to chemicals, such as pharmaceuticals, pesticides, and nanoparticles. Xenobiotics can also cause developmental cardiotoxicity through generation of reactive oxygen species (ROS) and oxidative stress (1), defined as "a disruption of redox signaling and control" (2). The zebrafish is a powerful model in developmental toxicology and has great potential for use in screening and understanding possible human teratogens, including those that act via oxidative stress. Much remains to be learned about the oxidative stress response during development. We have found that while embryos grow increasingly resistant to pro-oxidant exposure from 18-52 hours post fertilization (hpf), this is followed by a surprising window of heightened sensitivity between 72-96 hpf. Taking a collaborative and multidisciplinary approach, the objective of this proposal is to understand the mechanisms involved in the differential sensitivity to pro-oxidants during embryogenesis. Aim 1 will identify critical windows of sensitivity, the role of the antioxidant response element transcription factor NRF2, and the impact of pro-oxidant exposure on cardiac progenitor cell fate decisions. These experiments will involve ROS detection by chemiluminescence, cardiac function and cell fate analysis using calcium imaging, as well as embryo survival studies and assessment of the role of NRF2 through loss and gain of function experiments. Aim 2 will examine the role of glutathione, the most abundant component of antioxidant defenses, using quantitative real-time PCR, measurements of total glutathione, GSH:GSSG ratios, and modulation of GSH content. Aim 3 will examine the contribution of a physiologic increase in respiration and associated ROS towards oxidant sensitivity during development by measuring oxygen consumption using self-referencing oxygen sensing microelectrodes and reducing endogenous sources of ROS by over-expressing antioxidant enzymes on the outer mitochondrial membrane. Given the growing movement of government agencies and private industry to employ zebrafish for teratogenicity and developmental toxicity, it is especially important to comprehend the critical windows of sensitivity to oxidative stress and the underlying mechanisms. This research will provide a detailed and mechanistic understanding of how oxidative stress can impact embryonic development, and identify important considerations necessary when examining chemicals (and chemical mixtures that cause oxidative stress. It will also result in a better understanding of how oxidative stress influences the development and progression of human embryotoxicity and resulting teratogenesis and diseases.

描述（由申请人提供）：脊椎动物的胚胎发育非常容易因接触化学品（例如药物、杀虫剂和纳米颗粒）而受到破坏。异生素还可能通过产生活性氧 (ROS) 和氧化应激 (1) 导致发育性心脏毒性，氧化应激被定义为“氧化还原信号传导和控制的破坏”(2)。斑马鱼是发育毒理学中的强大模型，在筛选和了解可能的人类致畸剂（包括通过氧化应激起作用的致畸剂）方面具有巨大的潜力。关于发育过程中的氧化应激反应还有很多东西有待了解。我们发现，虽然胚胎在受精后 18-52 小时 (hpf) 内对促氧化剂暴露的抵抗力越来越强，但随后的 72-96 hpf 之间出现了一个令人惊讶的敏感性增强窗口。该提案的目的是采用协作和多学科的方法，了解胚胎发生过程中对促氧化剂的不同敏感性所涉及的机制。目标 1 将确定关键的敏感性窗口、抗氧化反应元件转录因子 NRF2 的作用以及促氧化剂暴露对心脏祖细胞命运决定的影响。这些实验将涉及通过化学发光进行ROS检测、使用钙成像进行心脏功能和细胞命运分析，以及胚胎存活研究和通过功能丧失和获得实验评估NRF2的作用。目标 2 将使用定量实时 PCR、总谷胱甘肽测量、GSH:GSSG 比率以及 GSH 含量调节来研究谷胱甘肽（抗氧化防御中最丰富的成分）的作用。目标 3 将通过使用自参考氧传感微电极测量耗氧量并通过在线粒体外膜上过度表达抗氧化酶来减少 ROS 的内源来源，研究发育过程中呼吸和相关 ROS 的生理增加对氧化剂敏感性的贡献。鉴于政府机构和私营企业越来越多地利用斑马鱼来研究致畸性和发育毒性，了解氧化应激敏感性的关键窗口及其潜在机制尤为重要。这项研究将对氧化应激如何影响胚胎发育提供详细和机械的理解，并确定在检查化学物质（以及引起氧化应激的化学混合物）时必要的重要考虑因素。它还将有助于更好地理解氧化应激如何影响发育人类胚胎毒性和由此产生的致畸和疾病的进展。