Project 3: Mitochondrial and cellular mechanisms of neurotoxicity of Superfund chemical co-exposures

摘要 在这个项目正在进行的工作中，我们发现发育过程中暴露于线粒体毒物会导致 秀丽隐杆线虫的神经毒性结果，包括神经元的形态学改变， 行为，并且，从长远来看，增加了对神经退行性疾病的易感性。根据EPA，我们描述 所有这些都被称为“发育神经毒性（DNT）”，因为它们是由暴露于 发展DNT的两个重要和支配性机制是：1）神经发生的变化，导致 改变的细胞命运、形态和连接性（“硬接线”），和2） 神经元在形态上似乎是正常的（表观遗传“编程”）。区分这些是 我们提出了一种新颖而强大的方法来评估每种可能性。我们将开始与体内尚未 相对高通量和经济的模式，C.优雅C. elegans提供了一个额外的关键好处： 发育性神经发生通常是不变的，这也使得我们能够清楚地识别硬连线的变异 作为行为和应激反应的变化，而没有形态改变（编程）。工作在C。 随后将在人类神经干细胞（hNSCs）中进行测试， 测试，加上机会，以确定性别特异性差异和表观遗传修饰。 相对较少的化学品已被严格评估为DNT。信息的匮乏更是 化学品共同接触是明显的，尽管事实上，综合接触是现实。这种缺乏 混合物的测试部分来自监管政策，部分来自实验室测试中的技术挑战 共同曝光。我们结合体内-体外方法将使我们能够严格测试由以下原因引起的DNT： 既有复杂的环境混合物，也有我们确定的个别超级基金化学品的组合， 将评估非累加效应。我们将测试典型的发育神经毒剂的作用 铅、镉和多环芳烃，单独和组合由已知的环境 浓度的我们将在C中比较结果。elegans和hNSCs，与其他项目在 鱼、老鼠和人。证明C.线虫可以可靠地用于研究混合物DNT将添加一个 用于测试和调节环境混合物的强大新模型。 最后，我们将测试线粒体功能障碍（神经发育的关键）驱动DNT的程度， 这些典型的化学物质。这些化学物质具有多个分子靶点，包括但不限于 不同的线粒体大分子。事实上，这些化学物质都单独影响线粒体， 神经元，但通过不同的机制，这就是为什么我们预测协同作用。然而，虽然线粒体 是这些化学物质的已知目标，线粒体毒性驱动其DNT的程度尚不清楚。 我们的工作将确定线粒体功能障碍在单一和联合化学DNT中的作用， 为不良后果途径和干预努力的发展提供信息。

Abstract In ongoing work by this Project, we have found that developmental exposures to mitochondrial toxicants cause neurotoxic outcomes in Caenorhabditis elegans, including morphological alterations in neurons, altered behavior, and, in the long term, increased susceptibility to neurodegeneration. In line with the EPA, we describe all of these as “developmental neurotoxicity (DNT),” because they result from exposures that occur during development. Two important and overarching mechanisms of DNT are 1) changes to neurogenesis resulting in altered cell fate, morphology, and connectivity (“hardwiring”), and 2) persistent changes to the function of neurons that appear to be morphologically normal (epigenetic “programming”). Distinguishing these is challenging; we propose a novel and powerful way to assess each possibility. We will begin with an in vivo yet relatively high-throughput and economic model, C. elegans. C. elegans offers an additional, key benefit: developmental neurogenesis is normally invariant, permitting clear identification of variation in hardwiring as well as behavioral and stress-responsive changes without morphological alteration (programming). Work in C. elegans will be followed by testing in human neuronal stem cells (hNSCs) that permit human-relevant DNT testing, plus the opportunity to identify sex-specific differences and epigenetic modifications. Relatively few chemicals have been rigorously evaluated for DNT. The paucity of information is even more pronounced for chemical co-exposures, despite the fact that combined exposures are the reality. This lack of testing of mixtures results partly from regulatory policy, and partly from technical challenges in laboratory testing of co-exposures. Our combined in vivo-in vitro approach will permit us to rigorously test for DNT resulting from both complex environmental mixtures, and from defined combinations of individual Superfund chemicals that we will evaluate for non-additive effects. We will test the effects of the prototypical developmental neurotoxicants Pb, Cd, and polycyclic aromatic hydrocarbons, singly and in combinations dictated by known environmental concentrations. We will compare our outcomes in C. elegans and hNSCs, to those obtained by other Projects in fish, rats, and people. Demonstration that C. elegans can be reliably used to investigate mixture DNT will add a powerful new model for testing and regulation of environmental mixtures. Finally, we will test the degree to which mitochondrial dysfunction, key to neurodevelopment, drives DNT by these prototypical chemicals. These chemicals have multiple molecular targets, including but not limited to different mitochondrial macromolecules. The fact that these chemicals individually all affect mitochondria and neurons, but by different mechanisms, is why we predict synergistic interactions. However, while mitochondria are known targets of these chemicals, the extent to which mitochondrial toxicity drives their DNT is not known. Our work will establish the contribution of mitochondrial dysfunction in single and combined chemical DNT, informing development of adverse outcome pathways and intervention efforts.