Crosstalk between estrogen and bile acid signaling pathway

描述(由申请人提供)：除了众所周知的在发育和维持正常的性和生殖功能方面的作用外，雌激素在非经典的内分泌器官系统中发挥着广泛的生物学作用，如心血管系统和肝脏。胆汁酸是胆固醇的代谢产物，是核受体法尼醇X受体(FXR)的一类信号分子。胆汁酸激活FXR调控着一系列FXR靶基因，参与维持胆汁酸、胆固醇、血脂和葡萄糖的稳态。胆汁酸盐导出泵(BSEP)通过激活FXR受到胆汁酸的正向调节，作为胆汁酸流出器，在维持胆汁酸在肝脏和胆汁中的动态平衡中起着重要作用。在胆汁酸水平失衡引起的疾病中，妊娠期肝内胆汁淤积症和胆石症都与性激素雌激素有关。尽管这些疾病与多种风险因素有关，但研究表明，雌激素在这些疾病的诱发中起着关键作用。雌激素诱导的胆汁淤积症和胆石症都已在动物模型上复制。然而，我们对潜在机制的理解还远远不完整。在我们的初步研究中，我们发现雌激素在体外和体内都抑制人BSEP的表达，这种抑制是由雌激素受体1(ER1)通过与FXR物理相互作用而介导的，表明雌激素/ER1和胆汁酸/FXR信号通路之间存在串扰。有待检验的中心假设是，雌激素下调BSEP的表达是通过一种新的非经典反式抑制途径介导的，即ER1和FXR之间的直接相互作用，因此是肝内胆汁淤积症和胆石病的共同危险因素。为了检验这一假说，本文提出了两个具体目标。具体目的1是通过以下三种方法确定雌激素介导的BSEP表达抑制的机制：(1)在体外和体内将牢固地建立ER1和FXR之间的物理相互作用；(2)将确定雌激素对BSEP启动子的FXR和ER1共调节因子的变化；以及(3)将绘制ER1中与FXR直接相互作用的区域。具体目的2是通过以下两种方法在体内研究雌激素介导的BSEP抑制、肝脏和胆道病理后果：(1)将使用Living Image技术确定整个妊娠期间在小鼠体内人BSEP反式激活的动态；(2)将在wt和ER1(-/-)小鼠中研究雌激素介导的BSEP抑制在雌激素诱导的胆汁淤积和胆结石疾病中的病因学作用。研究完成后，我们将通过ER1和FXR之间的直接相互作用，建立一个新的非经典雌激素转录抑制通路，揭示雌激素/ER1和胆汁酸/FXR信号通路之间的相互作用，并建立BSEP抑制在雌激素诱导的胆汁淤积和胆石病中的共同病因学作用。这些发现将通过研究雌激素对多种FXR靶基因的影响，为我们深入了解与雌激素相关的各种生物学和病理活动开辟新的领域。同样重要的是，这些发现将促进我们对雌激素诱导的妊娠肝内胆汁淤积症和胆石病的机制和发病机制的理解。这一结果将首次在这两种不同但相关的疾病之间建立起共同的病因学联系，最后，该发现将为开发治疗药物提供分子基础，通过调节BSEP表达和ER1/FXR信号通路来治疗或预防这些疾病。

DESCRIPTION (provided by applicant): In addition to the well-known roles in development and maintenance of normal sexual and reproductive function, estrogen exerts a vast range of biological effects in nonclassical endocrine organ systems, such as cardiovascular system and liver. Bile acids are metabolites of cholesterol and have been identified as a class of signaling molecules for nuclear receptor farnesoid X receptor (FXR). Activation of FXR by bile acids regulates a myriad of FXR target genes involving in maintenance of bile acids, cholesterol, lipids and glucose homeostasis. Bile acid salt export pump (BSEP) positively regulated by bile acids through activating FXR plays a critical role in maintaining bile acid homeostasis in the liver and bile as a canalicular bile acid effluxer. Among diseases resulted from imbalance of bile acid levels, intrahepatic cholestasis of pregnancy (ICP) and gallstone disease are both associated with sex hormone estrogen. Although multiple risk factors are linked to those disorders, studies have demonstrated that estrogen plays a key role in the induction of the diseases. Both estrogen-induced cholestasis and gallstone disease have been reproduced in animal model. However, our understanding on the underlying mechanisms is far from complete. In our preliminary study, we found that estrogen repressed human BSEP expression in vitro and in vivo, and the repression was mediated by estrogen receptor 1 (ER1) through physically interacting with FXR, indicating a crosstalk between estrogen/ER1 and bile acids/FXR signaling pathway. The central hypothesis to be tested is that down-regulation of BSEP expression by estrogen is mediated through a novel nonclassical transrepression pathway, a direct interaction between ER1 and FXR and as a consequence is a common risk factor for ICP and gallstone disease. Two specific aims are proposed to test the hypothesis. Specific Aim 1 is to determine the mechanisms for estrogen-mediated repression of BSEP expression by the following three approaches: (1) the physical interaction between ER1 and FXR will be firmly established in vitro and in vivo; (2) altered recruitment of FXR and ER1 coregulators to the BSEP promoter by estrogen will be determined; and (3) the region in ER1 directly interacting with FXR will be mapped. Specific Aim 2 is to investigate estrogen-mediated BSEP repression, hepatic and biliary pathological consequences in vivo with the following two approaches: (1) the dynamics of human BSEP transactivation in vivo in mice during the entire pregnancy will be determined with the Living Imaging technology; and (2) the etiological role of estrogen-mediated BSEP repression in estrogen-induced cholestasis and gallstone disease will be investigated in wt and ER1(-/-) mice. Upon completion of the proposed study, we will have firmly established a novel nonclassical estrogen transrepression pathway through a direct interaction between ER1 and FXR, revealed mechanistic insights into such crosstalk between the estrogen/ER1 and bile acids/FXR signaling pathway, and established a common etiological role of BSEP repression in estrogen-induced cholestasis and gallstone disease. The findings will open a new field to advance our understanding on the diverse biological and pathological activities associated with estrogens through investigating estrogen's effects on various FXR target genes. Equally significant, the findings will advance our understanding on the mechanisms and pathogenesis of estrogen-induced ICP and gallstone disease. For the first time, the results will establish a common etiological link between the two distinct but related diseases, and finally, the findings will provide a molecular basis for developing therapeutic agents to treat or prevent those disorders by modulating BSEP expression and ER1/FXR signaling pathways.