Functional and epigenetic effects of preconceptional EDCs on the female HPG axis

摘要 暴露于环境内分泌干扰化学品（EDCs），特别是在生命早期， 与不良生殖结果有关。多氯联苯（PCBs）是一种常见的 EDCs，在工业应用中使用了几十年。虽然几十年前就被禁止了，但人类仍然暴露在多氯联苯中。 现在主要是通过食物链。在人类和动物中， 内分泌和生殖功能失调然而，在这方面存在着几个重要的知识空白。 我们建议填写的字段。首先，虽然多氯联苯扰乱下丘脑-垂体-性腺（HPG）轴的每个水平， 在单独评估时，尚未探讨这些水平之间的相互影响。第二，虽然早 发育（胎儿，婴儿）被认为是易受内分泌干扰物影响的时期， 尚未研究多氯联苯对HPG系统的作用的另一个关键窗口，特别是在女性中， 我们建议纠正这一问题。第三，EDC暴露后的绝大多数表型分析是在 在一个生命阶段，通常是在成年期。然而，生殖系统经历了巨大的动态变化， 在整个生命的结构和功能，强调需要一个生命周期的方法来表型。的 第四个也可能是最大的差距是先入为主的暴露是否以及如何重新编程种系 表观基因组，如果这导致躯体效应。在这项提案中，我们将开发和利用一种新的 多氯联苯先入为主的代际接触模型。我们将喂养怀孕的SD大鼠母鼠 (F0祖母）人类相关水平的PCB混合物，或车辆，从E8-18，当生殖细胞表观遗传 F1胎仔性腺中的标记被擦除;和/或在哺乳期间，当F1幼仔为P1-21时， 当生殖细胞表观遗传标记重新建立时。F1代雌性大鼠与未处理的雄性大鼠交配， 生成F2代。F2雌性后代（孙子）将在3个生殖生命期进行表型分析 阶段：新生儿、青春期和成人。生理结果，关键HPG基因和蛋白质的表达 参与生殖能力的控制，以及潜在的分子表观遗传标记（DNA和组蛋白 甲基化）进行评估。此外，我们将使用一种新的方法分离原始生殖细胞（PGCs）， PCB和溶剂谱系的新生F1和F2雌性大鼠，以定量基因表达和甲基化 标记.这使我们能够将HPG表型与先前的生殖细胞编程联系起来。因此，这一提议是 预计提供新的见解的影响，preconceptibility多氯联苯暴露对女性HPG 生理学、功能和潜在的表观遗传调控机制，其结果与 女性的环境健康。

ABSTRACT Exposures to environmental endocrine-disrupting chemicals (EDCs), especially during early life, are strongly linked to adverse reproductive outcomes. Polychlorinated biphenyls (PCBs) are a well-established family of EDCs, used for decades in industrial applications. While banned decades ago, humans are exposed to PCBs today primarily through the food chain. In both humans and animals, higher PCB body burdens are associated with endocrine and reproductive dysfunctions. However, there are several important gaps in knowledge in this field that we propose to fill. First, while PCBs perturb each level of the hypothalamic-pituitary-gonadal (HPG) axis when assessed individually, the cross-talk among these levels has not been explored. Second, while early development (fetus, infant) is recognized as a period of vulnerability to EDCs, the preconceptional period is another critical window that has not been studied for PCB actions on HPG systems, especially in females, something we propose to redress. Third, the vast majority of phenotyping following EDC exposures is conducted at a single life stage, typically in the adult. Yet, the reproductive system undergoes enormous dynamic change in structure and function throughout life, underscoring the need for a lifecycle approach to phenotyping. The fourth and perhaps largest gap is whether and how preconceptional exposures reprogram the germline epigenome, and if that leads to somatic effects. In this proposal, we will develop and utilize a novel intergenerational model of preconceptional exposure to PCBs. We will feed pregnant Sprague-Dawley rat dams (F0 grandmothers) human-relevant levels of a PCB mixture, or vehicle, from E8-18, when germ cell epigenetic marks are being erased in the gonads of the F1 fetuses; and/or during lactation when F1 pups are P1-21 and when germ cell epigenetic marks are re-established. The F1 females are bred with untreated male rats to generate the F2 generation. The F2 female offspring (grandchildren) will be phenotyped at 3 reproductive life stages: neonatal, pubertal, and adult. Physiological outcomes, expression of key HPG genes and proteins involved in the control of reproductive competence, and underlying molecular epigenetic marks (DNA and histone methylation) will be assessed. In addition, we will use a novel method to isolate primordial germ cells (PGCs) of newborn F1 and F2 females rats of the PCB and vehicle lineage, to quantify gene expression and methylation marks. This allows us to relate HPG phenotypes to prior germ cell programming. Thus, this proposal is anticipated to provide novel insights into the effects of preconceptional PCB exposures on female HPG physiology, function, and underlying epigenetic mechanisms of regulation, results of which are directly relevant to women’s environmental health.