Environmental Estrogen Induced Epigenetic Alteration of Uterine Stem Cells

描述(申请人提供)：在人类中，接触到异种雌激素，如己烯雌酚(DES)和双酚A(BPA)，与女性生殖道发育障碍有关。研究人员此前已经证明，胎儿接触这些药物会改变子宫内膜中雌激素反应元件(ERE)的甲基化。ERE甲基化的改变导致体内雌激素受体(ER)对ERE的占据改变，并在整个生命过程中改变对雌激素的反应的基因表达。在人类或啮齿动物的每个月经周期或动情周期中，子宫内膜分别被替换。研究人员首先确定了再生这种组织的干细胞。由于雌激素反应的表观遗传变化在成人中持续存在，它们很可能在每个生殖周期中再生子宫内膜的干细胞中编码。研究人员假设，异种雌激素暴露会影响子宫干细胞中多个ERE的甲基化，导致成年后雌激素敏感性的改变。为了检验这一假设， 研究人员将确定DES或BPA暴露后整个基因组中子宫肌层甲基化的程度，以及在体外和体内暴露于这些药物对子宫内膜干细胞生长和雌激素反应的影响。具体目标包括1)使用染色质沉淀、大规模平行测序和亚硫酸氢盐测序来确定暴露是否导致多个ERE的优先甲基化以及这种情况是否发生在子宫干细胞中；2)确定暴露是否导致体外子宫内膜干细胞雌激素反应改变；以及3)使用体内模型来确定来自暴露于暴露的动物的子宫内膜干细胞是否更容易发生子宫内膜异位症或子宫内膜癌。这些研究将检验这样一种假设，即子宫干细胞中ERE的甲基化和由此引起的雌激素反应的改变将导致雌激素介导的疾病发生率增加，从而为与异种雌激素暴露相关的生殖道疾病提供表观遗传学机制。该模型解释了弱雌激素如何导致与其内在雌激素活性不成比例的雌激素反应，以及这种表观遗传信号如何持续存在，尽管由于子宫内膜干细胞甲基化的改变导致每个生殖周期的子宫内膜丢失。

DESCRIPTION (provided by applicant): In humans, exposure to xenoestrogens such as diethylstilbestrol (DES) and bisphenol A (BPA) has been associated with developmental disorders of the female reproductive tract. The investigators have previously shown that fetal exposure to these agents alters the methylation of estrogen response elements (EREs) in the endometrium. The change in ERE methylation results in altered ERE occupancy by estrogen receptor (ER) in vivo and altered gene expression in response to estrogens throughout life. The uterine endometrium is replaced in each menstrual cycle or estrus cycle in humans or rodents, respectively. The investigators were the first to identify the stem cells that regenerate this tisse. As the epigenetic changes in estrogen response persist in the adult, they are likely to be encoded in the stem cells that regenerate the endometrium in each reproductive cycle. The investigators hypothesize that xenoestrogen exposure affects methylation of multiple EREs in uterine stem cells leading to altered estrogen sensitivity as an adult. To test this hypothesis the investigators will determine the extent of uterine ERE methylation in the entire genome after DES or BPA exposure as well as the effect of exposure to these agents on endometrial stem cell growth and estrogen response in vitro and in vivo. The specific aims include 1) the use of chromatin precipitation, massively parallel sequencing and bisulfite sequencing to determine if exposure leads to preferential methylation of multiple EREs and if this occurs in uterine stem cells; 2) determine if exposure leads to altered endometrial stem cell estrogen response in vitro; and 3) use of an in vivo model to determine if endometrial stem cells from exposed animals are more prone to endometriosis or endometrial cancer. These studies will test the hypothesis that methylation of EREs in uterine stem cells and resultant altered estrogen responsiveness will lead to an increased incidence of estrogen mediated disorders, thus providing an epigenetic mechanism for reproductive tract disease associated with xenoestrogen exposure. This model explains how a weak estrogen results in an estrogenic response disproportionate to its intrinsic estrogenic activity and how this epigenetic signal persists despite loss of endometrium in each reproductive cycle due to altered methylation in endometrial stem cells.