The Nuclear Receptor-Aster Pathway in Enterohepatic Metabolism

摘要 脂质是肠肝轴中重要的内分泌信号，通过激活基因表达来控制基因的表达。 核激素受体。肝脏和肠道的脂质代谢是建立全身性 哺乳动物体内胆固醇和甘油三酯的动态平衡。这些组织中过多的脂肪堆积与 糖尿病、NASH、癌症和其他疾病。我们的长期目标是通过以下方式揭示基本机制 哪些脂质激活的核受体协调细胞和全身的脂质动态平衡。在当前 应用方面，我们重点介绍了LXR和FXR对一种新的类固醇转运途径的调控。我们发现 哺乳动物蛋白质家族(Aster-A、-B和-C)，在胆固醇运动中起重要作用。这个 3个ASTs以组织特异性的方式表达，并受核受体的差异调控。艾斯特- B是类固醇激活的LXR调节的靶点，而Aster-C是胆汁酸激活的FXR调节的靶标。 我们假设酯在肠肝轴的脂平衡中起关键作用，包括饮食中的脂类。 吸收、乳胶粒的产生和胆固醇的反向运输。我们进一步假设，星座是 对LXR和FXR激动剂的药理作用有重要贡献。我们将解决这些问题 有以下具体目的的假设。特定目标1将定义FXR调节的Aster-C在 肝脏胆固醇的运输。我们将结合细胞、生化、成像和活体研究 明确Aster-C依赖的胆固醇在肝细胞中的转运途径。我们产生了肝脏- 特异性Aster-C基因敲除小鼠，初步分析表明它们已经改变了肝脏和血脂 级别。我们将进行代谢分析和体内胆固醇示踪研究，以询问全身 胆固醇流动。我们将分析Aster蛋白的丢失或过表达如何影响高密度脂蛋白的运动- 基础水平和FXR反应时胆固醇进入胆汁和全身反向胆固醇转运 激动剂。具体目标2将阐明ASTs在肠道胆固醇运输中的作用。我们将分析 缺乏Aster-B和/或Aster-C的小鼠，以测试Aster在饮食中胆固醇吸收和 乳胶管生产。初步分析显示，饮食中胆固醇的摄入量减少， 肠细胞胆固醇酯含量在没有这两种紫杉醇的情况下。我们将进一步测试Asters是否 有助于跨肠道胆固醇的排泄。最后，我们发现，批准的药物 Ezetimibe抑制肠道胆固醇摄取，是Aster-C的选择性配体，我们已经解决了 Aster-C-ezetimibe络合物的晶体结构。根据这些发现，我们将确定 Aster途径参与依折麦布的药理作用。我们的目标有望实现 提供对脂类转运途径的基本洞察，并可能确定新的机会 对代谢性疾病的干预。

ABSTRACT Lipids are important endocrine signals in the enterohepatic axis that control gene expression by activating nuclear hormone receptors. Hepatic and intestinal lipid metabolism are key factors in establishing systemic cholesterol and triglyceride homeostasis in mammals. Excess lipid accumulation in these tissues is linked to diabetes, NASH, cancer and other diseases. Our long-term objective is to reveal fundamental mechanisms by which lipid-activated nuclear receptors orchestrate cellular and systemic lipid homeostasis. In the current application, we focus on regulation of a novel sterol-transport pathway by LXR and FXR. We have discovered a family of mammalian proteins (Aster-A, -B and -C) that play an important role in cholesterol movement. The 3 Asters are expressed in a tissue-specific manner and are differentially regulated by nuclear receptors. Aster- B is a target for regulation by sterol-activated LXRs, while Aster-C is regulated by the bile acid-activated FXR. We hypothesize the Asters play key roles in lipid homeostasis in the enterohepatic axis, including dietary lipid absorption, chylomicron production and reverse cholesterol transport. We further hypothesize that Asters are important contributors to the pharmacological effects of LXR and FXR agonists. We will address these hypotheses with the following specific aims. Specific Aim 1 will define the role of FXR-regulated Aster-C in hepatic cholesterol transport. We will use a combination of cellular, biochemical, imaging, and in vivo studies to define the pathway for Aster-C-dependent cholesterol transport in hepatocytes. We have generated liver- specific Aster-C knockout mice, and preliminary analysis reveals them to have altered hepatic and plasma lipid levels. We will perform metabolic analyses and in vivo cholesterol tracer studies to interrogate systemic cholesterol flux. We will analyze how loss or overexpression of Aster proteins affects the movement of HDL- cholesterol into bile and for systemic reverse cholesterol transport at baseline and in response to FXR agonists. Specific Aim 2 will elucidate the role of Asters in intestinal cholesterol transport. We will analyze mice lacking Aster-B, Aster-C, or both to test the importance of Asters in dietary cholesterol absorption and chylomicron production. Preliminary analysis has revealed reduced uptake of dietary cholesterol and reduced enterocyte cholesterol ester content in the absence of both Asters. We will further test whether Asters contribute to trans-intestinal cholesterol excretion. Finally, we have discovered that the approved drug ezetimibe, which inhibits intestinal cholesterol uptake, is a selective ligand for Aster-C, and we have solved the crystal structure of the Aster-C–ezetimibe complex. Based on these findings we will determine whether the Aster pathway contributes to the pharmacological effects of ezetimibe. Completion of our aims is expected to provide fundamental insight into pathways governing lipid transport, and may identify new opportunities for intervention in metabolic disease.