Mechanism of developmental toxicity of Bisphenol-A

描述（由申请人提供）：生殖道畸形和激素相关癌症的增加是工业化世界的重要问题。人们一直怀疑环境雌激素是其中一个致病因素。其中，双酚A （BPA）由于广泛的人体暴露而引起特别关注。啮齿动物围产期暴露于低剂量、与环境相关的双酚a会在雌激素靶组织中引起多营养效应，这种效应在暴露结束后很长时间才会显现出来。特别是，对发情周期至关重要的细胞核性别分化的改变和促性腺激素释放激素（GnRH）神经元激活的改变可能对生育能力和生殖力产生影响，而乳腺形态发生的改变可能会损害泌乳。我们预计BPA暴露的雌性雌激素靶组织中所观察到的影响将损害其生育健康存活后代的能力。拟议的研究将建立双酚a作用的因果机制链，包括细胞、组织和组织水平；因此，他们将兼具综合性和分析性。在综合方面，我们将评估围产期暴露的雌性小鼠的生殖成功率。这一信息对于阐明前一个资助周期中描述的分子事件的生理后果至关重要。在分析方面，我们提出了一种双重方法来研究BPA如何改变两个重要目标组织，发育中的下丘脑（HYP）和乳腺（MG）的组织组织。HYP对整体繁殖成功至关重要，而MG对新生儿的生存至关重要。此外，HYP可以通过调节垂体促性腺激素和卵巢激素的合成和催乳素的释放来影响MG的发展。目标1:BPA如何影响围产期暴露的雌性的生殖结果？为了确定发育性低剂量双酚a暴露对生殖的影响，将评估生育能力、生殖力和MG功能。目的2:BPA暴露如何改变发育中的HYP的组织结构？我们假设BPA改变了细胞核的结构和连通性，这对调节促性腺激素的释放很重要。我们将检查这些细胞核：i)细胞存活、凋亡和连接模式的变化；Ii)类固醇受体、睾酮代谢酶及雌激素作用下游因子如谷氨酸脱羧酶、星形胶质细胞分化的表达。这些研究的完成将确定GnRH神经元激活改变的机制。目标3:BPA暴露如何影响MG的基因表达和组织？我们假设：i)双酚a作为形态因子直接作用于MG源性胶原（在MG器官培养中通过QRT-PCR检测）；ii)双酚a效应是由内质网α和/或β介导的（将用无内质网小鼠进行QRT-PCR测试），iii)这些初始事件转化为基质-上皮相互作用的改变。为了从BPA对内分泌系统的影响分析BPA暴露对MG角原的影响，将BPA暴露和未暴露动物的MG分别移植到暴露和未暴露的宿主体内。为了评估间质、上皮或两者是否因BPA暴露而发生永久性改变，将进行组织重组研究。这一目的将开始揭示BPA扰乱雌激素靶器官的组织和结构的机制。该项目的实现将提供BPA在目标组织中的作用及其机体后果的机制信息。它还将揭示在替代模型中，当前的环境暴露水平是否会产生重大的健康影响。这一信息对于制定关于内分泌失调的公共政策至关重要。

DESCRIPTION (provided by applicant): Increased malformations of the genital tract and hormone-related cancers are significant problems in the industrialized world. Suspicions have focused on environmental estrogens as one causal agent. Among them, bisphenol A (BPA) is of particular interest due to widespread human exposure. Perinatal exposure of rodents to low, environmentally-relevant doses of BPA induces pleiotrophic effects in estrogen target tissues that manifest long after exposure has ended. In particular, altered sexual differentiation of a nucleus important for estrous cyclicity, and altered gonadotropin-releasing hormone (GnRH) neuronal activation may have repercussions on fertility and fecundity, while altered morphogenesis of the mammary gland may impair lactation. We expect that effects observed in estrogen target tissues of BPA exposed females will impair their ability to produce viable' and healthy offspring. The proposed studies will establish a causal mechanistic chain for BPA action encompassing cellular, tissue and organismal levels of organization; hence, they will be both integrative and analytical. On the integrative side, we will evaluate the reproductive success of perinatally exposed female mice. This information is essential to elucidate the physiological consequences of the molecular events described in the previous funding cycle. On the analytical side, we propose a dual approach to study how BPA alters the tissue organization of two important target tissues, the developing hypothalamus (HYP) and the mammary gland (MG). The HYP is critical to overall reproductive success, and the MG is critical to the survival of the neonates. In addition, the HYP can influence MG development by modulating pituitary gonadotropins and ovarian hormone synthesis and prolactin release. Aim 1: How does BPA affect the reproductive outcome of perinatally exposed females? Fertility, fecundity, and MG function will be assessed in order to define the reproductive impact of developmental low dose BPA exposure. Aim 2: How does BPA exposure alter tissue organization in the developing HYP? We hypothesize that BPA alters the architecture and connectivity of nuclei important for the regulation of gonadotropin release. We will examine these nuclei for: i) changes in patterns of cell survival, apoptosis, and connectivity; ii) expression of steroid receptors, enzymes of testosterone metabolism, and factors downstream of estrogen action such as glutamic acid decarboxylase and astrocyte differentiation. Completion of these studies will identify mechanisms underlying altered GnRH neuronal activation. Aim 3: How does BPA exposure affect gene expression and tissue organization in the MG? We hypothesize that: i) BPA acts as a morphogen directly on the MG anlagen (to be tested by QRT-PCR in MG organ culture); ii) BPA effects are mediated by ER alpha and/or beta (to be tested by QRT-PCR using null ER mice), and iii) these initial events translate into altered stroma-epithelium interactions. To dissect the effects resulting from BPA exposure of the MG anlagen from systemic effects due to the action of BPA on the endocrine system, the MG of BPA exposed and unexposed animals will be transplanted into exposed and unexposed hosts. To assess whether the stroma, the epithelium or both compartments are permanently altered by BPA exposure, tissue recombination studies will be performed. This Aim will begin to reveal the mechanisms by which BPA disturbs the organization and architecture of an estrogen target organ. The realization of this project will provide mechanistic information linking BPA action in target tissues and its organismal consequences. It will also reveal whether current levels of environmental exposure produce significant health effects in a surrogate model. This information is critically needed to develop public policy on endocrine disruption.