Role of NPC1L1 in regulating energy metabolism, nutrient utilization & weight gai

描述（由申请人提供）：本研究的目的是了解膳食脂质吸收和总体代谢之间的机制关系。依折麦布药物通过阻断其在肠道中的吸收来降低血浆胆固醇，显然是通过破坏与NPC 1 L1（尼曼-匹克病，C1型蛋白样蛋白1）相关的途径。对依折麦布其他生理学效应的初步研究表明，依折麦布给药小鼠的餐后肠道脂蛋白在大小和组成上与对照小鼠不同。我们的研究结果还表明，通过依折麦布治疗或基因缺失，NPC 1 L1功能的丧失可保护小鼠免受饮食诱导的肥胖和胰岛素抵抗。脂肪吸收减少可能是体重增加差异的部分原因，但其他机制，包括增加能量消耗，也可能有助于预防糖尿病和肥胖。该提案的重点是确定负责这些不同表型的机制。在特定目标1中，将对依折麦布给药小鼠、Npc 1 l1-/-小鼠和对照小鼠饲喂高脂肪饲料，这些饲料要么被各组充分吸收（基于红花油），要么被Npc 1 l1-/-和依折麦布给药小鼠吸收较差（基于可可脂）。将监测体重增加、血浆葡萄糖和能量消耗，以检验以下假设：除减少脂肪吸收外，药物治疗和基因敲除小鼠的机制有助于抵抗肥胖和糖尿病。其他实验将检查膳食脂肪在这三组小鼠的高能量组织如脂肪、肌肉、心脏和肝脏中的分配，以检验NPC 1 L1失活改变餐后脂质递送至这些组织并被这些组织利用的假设。目标1还将确定代谢变化是否源于慢性胆固醇丢失引起的肝细胞特异性受体活化增加。具体目标2将使用体内方法来确定肌肉、肝脏和脂肪对膳食甘油三酯利用的差异是否是由这些组织中的NPC 1 L1依赖性途径破坏引起的，或者是否是由胆固醇和脂肪酸吸收减少引起的乳糜微粒组成改变引起的。还将专门检测NPC 1 L1和依折麦布对这些组织葡萄糖代谢的影响。具体目标3将确定抑制胆固醇吸收减少饱和脂肪酸吸收而不影响不饱和脂肪酸吸收的机制。体外、细胞培养和体内实验将检验未吸收的胆固醇抑制胆盐胶束溶解饱和脂肪酸导致其沉淀和排泄的假设。总之，这些研究将为肥胖和糖尿病的根本原因提供新的见解，特别是脂肪吸收和代谢与葡萄糖代谢和整体能量平衡的关系。这些研究还将提供有关现有药物作用机制的有价值的信息，具有设计和开发用于治疗高脂血症、肥胖症和糖尿病等密切相关疾病的新治疗实体的潜力。 公共卫生相关性：该研究项目将提供有关肥胖和糖尿病的根本原因的重要新信息，特别是脂肪吸收和代谢与葡萄糖代谢和整体能量平衡的关系。这些研究还将为现有药物的作用机制提供有价值的信息，并将为设计和开发用于治疗高脂血症、肥胖症和糖尿病等密切相关疾病的新治疗实体确定潜在的新靶点。

DESCRIPTION (provided by applicant): The goal of this research is to understand the mechanistic relationship between dietary lipid absorption and overall metabolism. The drug ezetimibe lowers plasma cholesterol by blocking its absorption in the intestine, apparently by disrupting the pathway associated with NPC1L1 (Nieman-Pick disease, type C1-protein -like protein 1). Preliminary studies into other physiological effects of ezetimibe have revealed that postprandial intestinal lipoproteins from ezetimibe-treated mice are different in size and composition than those from control mice. Our results also show that loss of NPC1L1 function, through ezetimibe treatment or gene deletion, protects mice from diet-induced obesity and insulin resistance. Reduced fat absorption may account for part of the difference in weight gain, but additional mechanisms, including increased energy expenditure, may also confer protection against diabetes and obesity. The focus of this proposal is to determine the mechanism(s) responsible for these different phenotypes. In Specific Aim 1, ezetimibe-treated, Npc1l1-/-, and control mice will be fed high fat diets that are either well absorbed by each group (safflower oil based) or poorly absorbed by Npc1l1-/- and ezetimibe-treated mice (cocoa butter based). Weight gain, plasma glucose, and energy expenditure will be monitored to test the hypothesis that mechanisms in addition to reduced fat absorption contribute resistance to obesity and diabetes of drug-treated and knockout mice. Other experiments will examine the partitioning of dietary fat among high energy tissues such as adipose, muscle, heart and liver in these three groups of mice to test the hypothesis that NPC1L1 inactivation alters postprandial lipid delivery to and utilization by these tissues. Aim 1 will also determine if metabolic changes derive from increased activation of specific receptors in hepatocytes due to chronic cholesterol loss. Specific Aim 2 will use in vivo methods to determine if the difference in dietary triglyceride utilization by muscle, liver and adipose is caused by disruption of NPC1L1-dependent pathways in those tissues or if it is caused by altered chylomicron composition from decreased cholesterol and fatty acid absorption. The effect of NPC1L1 and ezetimibe on glucose metabolism by these tissues will also be specifically tested. Specific Aim 3 will identify the mechanism by which inhibiting cholesterol absorption reduces saturated fatty acid absorption without affecting absorption of unsaturated fatty acids. In vitro, cell culture, and in vivo experiments will test the hypothesis that unabsorbed cholesterol inhibits solubilization of saturated fatty acids by bile salt micelles causing their precipitation and excretion. Together, these studies will provide novel insights about underlying causes of obesity and diabetes, especially the relationship of fat absorption and metabolism with glucose metabolism and overall energy balance. These studies will also provide valuable information about mechanisms of action for existing drugs, with potential for the design and development of new therapeutic entities for treatment of the closely related diseases of hyperlipidemia, obesity and diabetes. PUBLIC HEALTH RELEVANCE: This research project will provide important new information about underlying causes of obesity and diabetes, especially the relationship of fat absorption and metabolism with glucose metabolism and overall energy balance. These studies will also provide valuable information about mechanisms of action for existing drugs, and will identify potential new targets for the design and development of new therapeutic entities for treatment of the closely related diseases of hyperlipidemia, obesity and diabetes.