Role of cholesteryl ester transfer protein in cellular lipid homeostasis

项目概要/摘要 胆固醇酯转移蛋白（CETP）在胆固醇酯（CE）和甘油三酯（TG）之间交换 脂蛋白，并被公认为血浆脂蛋白代谢的调节剂。 CETP也驻留在里面 存在两种亚型的细胞：全长 (FL) 和外显子 9 (E9) 删除的 CETP。 CETP内部的功能 细胞中的作用尚不清楚，但它显然是必要的，因为抑制脂肪细胞中的 CETP 生物合成会损害脂质 存储。我们假设发生这种情况是因为细胞内 CETP 是 CE 和 TG 从其细胞中转运所必需的。 合成位点到储存位点。为了检验这个一般假设，我们提出了三个具体目标。目标1 - 确定促进细胞脂质储存的CETP亚型，并定义CETP的结构特征 这对于其细胞内功能至关重要。该目标测试了 E9 删除的 CETP 促进的假设 直接或通过与 FL CETP 相互作用来检测细胞内脂质转运，并检查这种细胞活性如何 取决于CETP结构。目标 2 - 定义 CETP 在细胞脂质代谢中的作用。为了解开 CETP 缺乏破坏细胞脂质代谢的机制，该目标量化代谢 改变并定义了当 CETP 同种型表达改变时发生的脂滴形成的变化。 目标 3 - 定义 CETP 在脂蛋白中脂质储存液滴的形成和利用中的作用 合成细胞。与初步研究一致，我们提出在脂蛋白合成细胞中CETP 有助于脂滴形成和将储存的脂质逆向转运至微粒体以获得脂蛋白 装配。遗传和腺病毒方法将改变 SW872 中 FL 和/或 E9 删除的 CETP 的表达 脂肪细胞、Caco-2 肠细胞和仓鼠。这些分子的后果 脂质合成、细胞器间脂质转运、脂滴形成和脂蛋白组装的操作 将通过生化和显微镜技术进行研究。这些研究将描述一个新功能 CETP 并增进我们对细胞脂质运输和储存机制的理解。脂质储存于 脂肪细胞与其分泌调节葡萄糖和脂质代谢的激素有关，这些激素直接 影响炎症和动脉粥样硬化等过程。

PROJECT SUMMARY/ABSTRACT Cholesteryl ester transfer protein (CETP) exchanges cholesteryl ester (CE) and triglyceride (TG) between lipoproteins and is well recognized as a regulator of plasma lipoprotein metabolism. CETP also resides inside cells where two isoforms exist: full-length (FL) and exon 9 (E9)-deleted CETP. The function of CETP inside cells is not understood, but it is clearly essential since inhibiting CETP biosynthesis in adipocytes impairs lipid storage. We postulate this occurs because intracellular CETP is required for CE and TG transport from their site of synthesis to their site of storage. To test this general hypothesis, we propose three specific aims. Aim 1 - Determine the CETP isoform(s) that promote cellular lipid storage, and define the structural features of CETP that are essential to its intracellular function. This aim tests the hypothesis that E9-deleted CETP facilitates intracellular lipid transport directly or by interacting with FL CETP, and also examines how this cellular activity depends on CETP structure. Aim 2 - Define the role of CETP in cellular lipid metabolism. To unravel the mechanisms by which CETP deficiency disrupts cellular lipid metabolism, this aim quantifies metabolic alterations and defines changes in lipid droplet formation that occur when CETP isoform expression is altered. Aim 3 - Define the role of CETP in the formation and utilization of lipid storage droplets in lipoprotein- synthesizing cells. Consistent with preliminary studies, we propose that in lipoprotein-synthesizing cells CETP assists in both lipid droplet formation and the reverse transport of stored lipids to microsomes for lipoprotein assembly. Genetic and adenoviral approaches will alter expression of FL and/or E9-deleted CETPs in SW872 adipocytes, Caco-2 intestinal enterocytes, and in hamster. The consequences of these molecular manipulations on lipid synthesis, interorganelle lipid transport, lipid droplet formation and lipoprotein assembly will be studied through biochemical and microscopy techniques. These studies will describe a novel function for CETP and advance our understanding of cellular lipid transport and storage mechanisms. Lipid storage in adipocytes is linked to their secretion of hormones that regulate glucose and lipid metabolism, which directly affect processes such as inflammation and atherogenesis.