MECHANISMS AND REGULATION OF INTESTINAL CHOLESTEROL TRANSPORT

肠道胆固醇转运的机制和调节

• 批准号: 8543330
• 负责人: Waddah A. Alrefai
• 金额: --
• 依托单位: JESSE BROWN VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8543330
• 关键词: Address; Anticholesteremic Agents; Azacitidine; Binding; Binding Proteins; Blood; Cell Line; Cell physiology; Cells; Cholesterol; Cholesterol Homeostasis; Chylomicrons; Coronary heart disease; DNA; DNA Methylation; Data; Development; Diabetes Mellitus; Disease; Enterocytes; Epigenetic Process; Epithelial Cells; Esterification; Fatty Acids; Gastrointestinal tract structure; Gene Expression; Gene Silencing; Genes; Hepatic; In Vitro; Intestines; LDL Cholesterol Lipoproteins; Lead; Lipids; Liver; Low-Density Lipoproteins; Lysosomes; MAP Kinase Gene; MAPK Signaling Pathway Pathway; Maintenance; Mediating; Mediator of activation protein; Microarray Analysis; Mitogen-Activated Protein Kinases; Modality; Molecular; Mus; Pathway interactions; Patients; Pharmaceutical Preparations; Plasma; Play; Production; Proteins; Regulation; Regulatory Pathway; Research Design; Response Elements; Risk; Role; Serum; Site; Sterols; Therapeutic; Transgenic Mice; Ubiquitination; Very low density lipoprotein; absorption; cholesterol absorption; cholesterol biosynthesis; design; diabetic patient; effective therapy; ezetimibe; high risk; hypercholesterolemia; in vivo; in vivo Model; inhibitor/antagonist; intestinal epithelium; jejunum; mouse model; novel; overexpression; patient population; promoter; protein degradation; public health relevance; response; transcription factor

DESCRIPTION (provided by applicant): Inhibition of intestinal Niemann-Pick type C1 Like 1 (NPC1L1) transporter, an essential mediator of intestinal cholesterol absorption, has emerged as an important target for management of hypercholesterolemia. Despite such advances in the therapeutic modalities, reducing plasma cholesterol to low levels in patients with high risk for coronary heart disease remains poor. Therefore, identifying novel mechanisms targeting NPC1L1 is warranted for more effective treatment of hypercholesterolemia. The inhibition of NPC1L1, however, triggers the activation of adaptive cellular pathways such as the Sterol Response Element Binding Protein-2 (SREBP2) that could hinder the efficiency of cholesterol lowering drugs. Suppressing these compensatory mechanisms should also be taken into consideration when designing more effective treatment of hypercholesterolemia. Our efforts are focused at investigating the novel mechanisms involved in the inhibition of NPC1L1 as well as delineating roles of intestinal SREBP2 in body cholesterol homeostasis. Our preliminary data showed that proteasomal and lysosomal-dependent pathways are involved in the normal turnover of NPC1L1 protein. Also, inhibition of MAPK pathway caused a decrease in the cellular level of NPC1L1 by decreasing its protein stability. Our findings showed that DNA methylation is a major epigenetic determinant involved in controlling NPC1L1 expression in intestinal epithelia cells. To understand the role of compensatory pathways, we have recently generated novel transgenic mice with intestine- specific overexpression of constitutively active SREBP2 (designated as ISR2 mice). Our data showed an increase in levels of plasma cholesterol in the LDL and VLDL fractions in ISR2 mice indicating that the activation of intestinal SREBP2 alone is sufficient to increase plasma cholesterol. We hypothesized that inhibition of MAPK pathway decreases NPC1L1 protein stability via proteasomal and/or lysosomal degradation, and that DNA methylation is involved in NPC1L1 gene silencing. We further hypothesize that the increase in intestinal lipid synthesis/absorption and chylomicron production by active intestinal SREBP2 lead to an increase in plasma cholesterol. Our studies are designed to address these hypotheses utilizing both in vitro and in vivo models. Studies designed for Specific Aim 1 will investigate the mechanisms involved in the regulation of NPC1L1 protein stability by the MAPK dependent pathway and determine the structural domains of NPC1L1 involved. Specific Aim 2 will systematically investigate mechanisms of NPC1L1 gene silencing by DNA methylation. Studies designed for Specific Aim 3 will focus on investigating the compensatory mechanisms underlying hypercholesterolemia in mice with intestine specific-overexpression of active SREBP2 (ISR2 mice). Our proposed studies are important to elucidate the critical role of the intestine in the maintenance of cholesterol homeostasis and to unravel potential targets for more effective treatment of hypercholesterolemia.

描述（由申请人提供）： 抑制肠Niemann-pick型C1，例如1（NPC1L1）转运蛋白是肠道胆固醇吸收的必不可少的介体，已成为治疗高胆固醇血症的重要靶标。尽管治疗方式取得了这样的进步，但患有冠心病高风险的患者的血浆胆固醇降低到较低的水平仍然很差。因此，有必要确定针对NPC1L1的新型机制，以更有效地治疗高胆固醇血症。然而，NPC1L1的抑制作用会触发适应性细胞途径的激活，例如固醇反应元件结合蛋白-2（SREBP2），这可能会阻碍降低胆固醇降低药物的效率。在设计更有效的高胆固醇血症治疗时，还应考虑抑制这些补偿机制。我们的努力集中在研究抑制NPC1L1的新机制，以及描述肠道SREBP2在人体胆固醇稳态中的作用。我们的初步数据表明，蛋白酶体和溶酶体依赖性途径与NPC1L1蛋白的正常周转有关。同样，抑制MAPK途径通过降低其蛋白质稳定性，导致NPC1L1的细胞水平降低。我们的发现表明，DNA甲基化是控制肠上皮细胞中NPC1L1表达的主要表观遗传决定因素。为了了解补偿途径的作用，我们最近产生了新型的转基因小鼠，具有组成型活性SREBP2（指定为ISR2小鼠）的肠道特异性过表达。我们的数据显示，ISR2小鼠中LDL和VLDL级分中血浆胆固醇水平升高，表明仅肠道SREBP2的激活就足以增加血浆胆固醇。我们假设抑制MAPK途径可通过蛋白酶体和/或溶酶体降解降低NPC1L1蛋白的稳定性，并且DNA甲基化参与NPC1L1基因沉默。我们进一步假设，活跃的肠道SREBP2肠道脂质合成/吸收和乳糜微粒的增加导致血浆胆固醇的增加。我们的研究旨在通过体外和体内模型来解决这些假设。专为特定目标1设计的研究将研究通过MAPK依赖途径调节NPC1L1蛋白稳定性的机制，并确定所涉及的NPC1L1的结构结构域。具体目标2将系统地研究通过DNA甲基化的NPC1L1基因沉默的机制。专为特定目标3设计的研究将重点研究在活性SREBP2（ISR2小鼠）肠道特异性过表达的小鼠中的高胆固醇血症基础的补偿机制（ISR2小鼠）。我们提出的研究对于阐明肠道在维持胆固醇稳态中的关键作用和揭开潜在的靶标以更有效的高胆固醇血症的靶标很重要。