Regulation of Intestinal Bile Acid Transport

描述（由申请方提供）：已证明抑制肠道胆汁酸吸收可降低血浆胆固醇。虽然临床上证明胆汁酸螯合剂可降低胆固醇，但患者依从性差需要开发更好的治疗方式来直接抑制胆汁酸吸收。顶端钠依赖性胆汁酸转运蛋白（ASBT）在回肠中吸收大部分胆汁酸，并且对于维持肝肠循环中的胆汁酸池是必需的。因此，ASBT抑制代表了降低血浆胆固醇的有吸引力的治疗手段。在这方面，我们实验室最近的研究表明，ASBT被信号中间体抑制，包括蛋白酪氨酸磷酸酶（PTPases）通过膜再循环事件。此外，有益的膳食组分如绿色茶儿茶素、（-）-表没食子儿茶素-3-没食子酸酯、EGCG以脂筏依赖性方式抑制ASBT功能。我们假设磷酸化/去磷酸化过程，脂筏依赖性机制和膜运输事件在抑制ASBT功能中起关键作用。因此，全面了解这些抑制途径是至关重要的，利用它们作为一种有效的治疗高胆固醇血症与糖尿病。我们的初步研究表明，在链脲佐菌素（STZ）诱导的糖尿病大鼠模型中，ASBT的功能和表达上调。这种糖尿病体内模型将提供一种特殊的工具来研究糖尿病中ASBT上调的潜在机制以及确定ASBT抑制对相关高胆固醇血症的影响。本研究旨在利用体外模型系统地阐明ASBT抑制的细胞和分子机制，并利用糖尿病大鼠模型研究其体内失调。在特定目标1中，我们将研究细胞培养模型中蛋白磷酸酶（PPase）对ASBT功能和磷酸化的调节。在具体目标2中，我们的研究将集中在阐明ASBT功能的抑制机制，通过膜再循环事件和脂筏，以及描绘EGCG介导的抑制的分子基础。针对特定目标3设计的研究将侧重于研究STZ诱导的糖尿病大鼠模型中ASBT上调的潜在机制，并确定有益的饮食化合物EGCG和特定ASBT抑制剂（由生物技术公司Albireo开发）在降低血浆胆固醇水平方面的功效。我们提出的研究对于在正常和病理生理条件下对ASBT的调节提供新的见解至关重要，并可能为与多种疾病相关的高胆固醇血症的管理提供更好的策略。 公共卫生相关性：肠胆汁酸吸收的增加与糖尿病患者血浆胆固醇水平升高和心血管疾病风险增加有关。负责胆汁酸吸收的主要蛋白质是回肠ASBT。因此，研究ASBT抑制的机制对于改善目前对高胆固醇血症的治疗是至关重要的，特别是在糖尿病患者中，这些患者对现有的治疗方式没有有效的反应。拟开展的研究将侧重于阐明ASBT功能抑制的机制，并评估其对糖尿病大鼠模型高胆固醇血症的影响。由于糖尿病的高发病率，这些研究与公共卫生直接相关，并且对于设计未来管理胆固醇相关疾病的更好治疗方式具有重要意义。

DESCRIPTION (provided by applicant): Inhibition of intestinal bile acid absorption has been shown to reduce plasma cholesterol. Although, bile acid sequestrants are clinically proven to lower cholesterol, poor patient compliance necessitates the development of better therapeutic modalities to directly inhibit bile acid absorption. Apical Sodium Dependent Bile Acid Transporter (ASBT) absorbs majority of the bile acids in the ileum and is essential for maintaining bile acid pool in the enterohepatic circulation. Therefore, ASBT inhibition represents an attractive therapeutic means for lowering plasma cholesterol. In this regard, recent studies from our laboratory demonstrated that ASBT is inhibited by signaling intermediates including protein tyrosine phosphatases (PTPases) via membrane recycling events. Also beneficial dietary components such as green tea catechin, (-)-epigallocatechin-3-gallate, EGCG, inhibits ASBT function in a lipid-raft dependent manner. We hypothesized that phosphorylation/dephosphorylation processes, lipid raft-dependent mechanisms and membrane trafficking events play critical roles in the inhibition of ASBT function. Hence, a comprehensive understanding of these inhibitory pathways is crucial to exploit their utilization as an effective therapy for hypercholesterolemia associated with diabetes mellitus. Our preliminary studies showed that ASBT function and expression are upregulated in rat model of streptozotocin (STZ)-induced diabetes mellitus. This in vivo model of diabetes mellitus will provide an exceptional tool to investigate the underlying mechanisms of ASBT upregulation in diabetes mellitus as well as determining the impact of ASBT inhibition on associated hypercholesterolemia. Our studies are designed to systematically delineate the cellular and molecular mechanisms inhibiting ASBT utilizing in vitro models and to examine their dysregulation in vivo utilizing diabetes mellitus rat model. In Specific Aim 1, we will investigate the regulation of ASBT function and phosphorylation by protein phosphatases (PPase) in cell culture models. In Specific Aim 2, our studies will focus on elucidating the inhibitory mechanisms of ASBT function by membrane recycling events and lipid rafts as well as delineating the molecular basis for EGCG-mediated inhibition. Studies designed for Specific Aim 3 will focus on investigating mechanisms underlying ASBT upregulation in rat model of STZ-induced diabetes mellitus and determine the efficacy of the beneficial dietary compound EGCG and specific ASBT inhibitors (developed by Biotechnology Company Albireo) in lowering the levels of plasma cholesterol. Our proposed studies are critical for providing novel insights into the regulation of ASBT under normal and pathophysiological conditions and may provide better strategies for the management of hypercholesterolemia associated with several disorders. PUBLIC HEALTH RELEVANCE: An increase in intestinal bile acid absorption has been implicated in the elevated plasma cholesterol levels and the increased risk of cardiovascular disorders in patients with diabetes mellitus. The major protein responsible for bile acid absorption is ileal ASBT. Therefore, investigating the mechanisms of ASBT inhibition is crucial to improve current therapy of hypercholesterolemia especially in diabetic patients, who do not efficiently respond to available therapeutic modalities. The proposed studies will focus on elucidating the mechanisms involved in the inhibition of ASBT function and evaluating their impact on hypercholesterolemia in rat model of diabetes mellitus. These studies have direct relevance to public health because of the high incidence of diabetes mellitus, and are of a great importance for designing better therapeutic modalities for the management of cholesterol-related disorders in the future.