Polybrominated Diphenyl Ether Effects on Human Neuronal Development

描述(由候选人提供)：我寻求资金，通过获得三个领域的额外专业知识来扩展我在环境健康科学和发育毒理学方面的培训：人类胚胎干细胞(HESC)生物学、分子生物学和生物信息学。学习在这三个领域使用的基于实验室的方法和统计方法将为我提供过渡到独立学术生涯所需的技能，调查环境暴露如何改变人类早期胚胎发育，以及它们对儿童和成人疾病的潜在贡献。在K99阶段，我的主要导师是苏珊·费舍尔博士，她的专长包括使用hESC模型来研究人类发育。我还将从我的其他导师那里获得宝贵的意见，他们是人类神经元发育、产前环境暴露和生物信息学领域的领导者。我将在hESC生物学、将这些细胞分化为神经元的方法以及研究毒物对这一过程的影响的方法方面获得广泛的培训。人类胚胎干细胞是人类胚胎发生的一个有价值的模型，因为它们发育成三个初级生殖层及其衍生细胞，如神经元。此外，我还将学习研究转录组的最先进技术，例如RNA测序(RNA-seq)。总体而言，我的理论是，怀孕期间特定的环境暴露影响hESC的自我更新和/或其向神经元分化的一个或多个步骤，导致有害的发育和神经行为结果。在这里，我的目标是检验一个特定的假设，即在与环境相关的水平上，广泛使用的阻燃剂和潜在的神经发育毒剂多溴二苯醚(PBDE)会扰乱人类的神经发生。首先，使用神经元分化的hESC模型，我将建立相应的同系物PBDE-47和/或PBDE-99的有效剂量(目标1)。其次，在相关范围的磨练中，我将在转录组水平上表征多溴二苯醚的剂量依赖效应(目标2)。最后，我将确定PBDE基因靶点在hESC自我更新和/或神经元发育方面的功能相关性(目标3)。拟议的实验是第一次研究PBDE对人胚胎干细胞及其神经元衍生物的影响。预期的结果是识别转录组水平上的变化，并对其表达水平发生变化的基因进行功能分析。随后，这项工作可能会导致几个新的研究方向。例如，根据不利的神经发育结果，可以使用体内模型来验证在体外确定的基因-环境相互作用。此外，了解多溴二苯醚靶标是否针对这些化学物质或更广泛地指示神经发育毒性将是有趣的。我的主要导师和共同导师，在互补的领域，将指导我学习开展拟议研究所需的技术。通过完成这个项目，我将掌握强大的方法，使我能够达到我的最终目标，作为一名独立研究员，研究潜在毒物对人类早期发育的种子方面的影响。

DESCRIPTION (provided by candidate): I seek funding to extend my training in environmental health science and developmental toxicology by acquiring additional expertise in three areas: human embryonic stem cell (hESC) biology, molecular biology, and bioinformatics. Learning laboratory-based methods and statistical approaches used in these three areas will provide me with the skills that are required to transition into an independent academic career, investigating how environmental exposures alter early human embryogenesis, and their potential contributions to childhood and adult diseases. For the K99 phase, my primary mentor is Dr. Susan Fisher, whose expertise includes using hESC models for studying human development. I will also receive valuable input from my other mentors who are leaders in the fields of human neuronal development, prenatal environmental exposures, and bioinformatics. I will obtain extensive training in hESC biology, methods for differentiating these cells into neurons, and approaches for studying the effects of toxicants on this process. HESCs are a valuable model of human embryogenesis because they develop into the three primary germ layers and their derivatives such as neurons. Additionally, I will learn state-of-the-art technique for investigating the transcriptome, e.g., RNA- sequencing (RNA-seq). In general, I theorize that specific environmental exposures during pregnancy impact hESC self-renewal and/or one or more steps in their differentiation to neurons, resulting in detrimental developmental and neurobehavioral outcomes. Here, I aim to test the specific hypothesis that, at environmentally relevant levels, polybrominated diphenyl ethers (PBDE), widely-used flame retardants and potential neurodevelopmental toxicants, disrupt human neurogenesis. First, using an hESC model of neuronal differentiation, I will establish the relevant effective doses of congeners PBDE-47 and/or PBDE-99 (Aim 1). Second, honing in on the relevant ranges, I will characterize the dose-dependent effects of PBDEs at the level of the transcriptome (Aim 2). Finally, I will determine the functional relevance of PBDE gene targets in terms of hESC self-renewal and/or neuronal development (Aim 3). The proposed experiments are the first investigation of PBDE effects on hESCs and their neuronal derivatives. The expected result is identification of alterations at the level of the transcriptome and a functional analysis of genes whose expression levels change. Subsequently, this work could lead to several new research directions. For example, gene-environment interactions identified in vitro could be validated, in terms of adverse neurodevelopmental outcomes, using in vivo models. Additionally, it will be interesting to learn if the PBDE targets are specific to these chemicals or more broadly indicative of neurodevelopmental toxicity. My primary mentor and co-mentors, in complementary fields, will guide me in learning the techniques required to carry out the proposed research. By completing this project, I will master powerful approaches that will enable me to reach my ultimate goal of studying, as an independent investigator, the effects of potential toxicants on seminal aspects of early human development.