Membrane-Based Mechanisms of Hepatic Insulin Resistance

肝胰岛素抵抗的膜机制

• 批准号: 9350309
• 负责人: Baran Ersoy
• 金额: $ 15.19万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2019-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9350309
• 关键词: Ablation; Activation Analysis; Advisory Committees; Affect; Applications Grants; Attenuated; Automobile Driving; Basic Science; Binding; Binding Proteins; Biological; Bone Marrow Transplantation; Calcium Channel; Cell Culture System; Cell Culture Techniques; Cells; Cellular biology; Chemicals; Committee Members; Data; Defect; Development; Diabetes Mellitus; Diet; Disease; Education; Endocrinologist; Endoplasmic Reticulum; Environment; Enzyme-Linked Immunosorbent Assay; Exhibits; Extrahepatic; Fasting; Genetic; Glucose Metabolism Disorders; Goals; Harvest; Hepatic; Hepatocyte; Hepatology; High Fat Diet; Hospitals; IRS2 gene; Immune response; Immunoblot Analysis; Impairment; In Vitro; Inflammation; Inflammatory; Insulin; Insulin Receptor; Insulin Resistance; Interleukin-6; Intervention; Laboratories; Lead; Lecithin; Link; Lipid Binding; Lipids; Lipopolysaccharides; Liver; MAPK8 gene; Macrophage Activation; Mass Spectrum Analysis; Measures; Mediating; Medication Management; Medicine; Membrane; Mentored Research Scientist Development Award; Mentors; Mentorship; Metabolic; Mission; Molecular; Molecular Biology; Molecular Immunology; Mus; National Institute of Diabetes and Digestive and Kidney Diseases; Non-Insulin-Dependent Diabetes Mellitus; Obesity; Outcome Study; Pathway interactions; Peritoneal Macrophages; Pharmacology; Phospholipids; Phosphotransferases; Physician Executives; Physiological; Play; Positioning Attribute; Prevention; Proteins; Proteomics; Public Health; RNA Splicing; Reagent; Regulation; Research; Research Personnel; Research Proposals; Resistance; Role; Scientist; Signal Transduction; Small Interfering RNA; Starvation; TNF gene; Techniques; Testing; Time; Tissues; Training; Transgenic Mice; Tunicamycin; Woman; base; biological adaptation to stress; blood glucose regulation; career; common treatment; cytokine; endoplasmic reticulum stress; experience; experimental study; feeding; glucose metabolism; glucose production; improved; instructor; insulin sensitivity; insulin signaling; knock-down; lipid metabolism; loss of function; medical schools; mouse model; new therapeutic target; non-alcoholic fatty liver; novel; novel therapeutic intervention; phosphatidylcholine transfer protein; professor; protein complex; public health relevance; response; skills; small molecule; small molecule inhibitor; success

DESCRIPTION (provided by applicant): This is an application for a K01 Award for Dr. Baran A. Ersoy, an Instructor in Medicine at the Brigham and Women's Hospital (BWH) and Harvard Medical School (HMS). Dr. Ersoy is a young investigator in basic research on disorders of glucose and lipid metabolism. His application outlines a short-term plan to further his education by gaining experience in advanced molecular biology and immunology techniques, including mass spectrometry of proteomics and bone marrow transplantation. This plan will build upon his previous experience and provide him the time and training to establish new research skills, strategies and projects that will contribute to his success upon becoming an independent scientist. It is his long-term goal to become an establish independent investigator in an academic setting and study the mechanisms driving insulin resistance and promoting type 2 diabetes and non-alcoholic fatty liver disease (NAFLD). To achieve these goals, Dr. Ersoy will be mentored by Dr. David E. Cohen, Professor of Medicine at HMS and Director of Hepatology at BWH. Dr. Ersoy has also assembled an advisory committee comprised of Dr. Steven P. Gygi, Professor of Cell Biology at HMS, who focuses on mass spectrometry of proteomics; Dr. Umut Ozcan, an endocrinologist with expertise in endoplasmic reticulum stress, inflammation and insulin signaling; and Dr. Morris F. White, an expert in insulin signaling who played a major role in the discovery insulin receptor substrate proteins. Dr. Ersoy's mentor and advisory committee members have established mentorship and training record, whose previous trainees hold independent investigator positions throughout the world. Dr. Ersoy will conduct his training at BWH and HMS, which will provide an intellectually rich research environment and has a long- standing commitment to training young scientists for independent academic research careers. Obesity-induced resistance to insulin action is the primary pathophysiological defect that predisposes to NAFLD and type 2 diabetes. Because current management options remain limited, identification of new regulatory mechanisms that govern the metabolic response to insulin should serve to identify novel opportunities for pharmacologic intervention. The objective of this research proposal is to elucidate a fundamental new mechanism for the regulation of glucose homeostasis. The rationale is that the identification of a novel mechanism that regulates hepatic insulin sensitivity could lead to the identification of new therapeutic targets for the treatment of NAFLD and type 2 diabetes. Guided by extensive preliminary data, the central hypothesis of this research plan is that phosphatidylcholine transfer protein (PC-TP) regulates glucose homeostasis by controlling the activity of insulin receptor substrate 2 (IRS2) both directly and also indirectly by modulating endoplasmic reticulum (ER) stress and inflammation. This will be tested in three specific aims: 1) Define the mechanisms by which PC-TP suppresses IRS2 activity; 2) Determine the role of ER stress in PC- TP-mediated regulation of insulin signaling; 3) Elucidate the role of immune response in PC-TP-mediated inhibition of insulin sensitivity. In Aim 1, a small molecule inhibitor of PC-TP (compound A1) and IRS2 deficient mice will be used to examine the role of IRS2 in the regulation of insulin signaling by PC-TP. The biological pathways in the control of IRS2 inhibition by PC-TP will be unveiled using mass spectroscopy of proteomics in Pctp+/+ and Pctp-/- mice as well as compound A1-treated hepatocytes harvested from these mice. In Aim 2, Pctp+/+ and Pctp-/- mice and hepatocytes harvested from these mice will be used to determine whether PC-TP modulates key ER stress pathways that regulate insulin signaling and IRS2 activity by immunoblot analyses. ER stress signaling will be activated by refeeding following 24 h starvation in mice, and by the chemical reagent tunicamycin in cell culture systems. Aim 3 will utilize peritoneal macrophages harvested from Pctp+/+ and Pctp-/- mice and bone marrow transplantation of Pctp-/- donor cells into Pctp+/+ recipient mice. Inflammation will be induced by high fat diet feeding in mice, and by lipopolysaccharide treatment in cell culture. Inflammatory signals will be measured by ELISA, immunoblot and qPCR analyses. It is anticipated that PC-TP will regulate insulin signaling by regulating IRS2 activity both directly and by modulating ER stress and inflammation. This is important because loss of IRS2 expression and increased ER stress and inflammation are associated with development of insulin resistance. These studies are expected to identify new therapeutic targets for the management of insulin resistance and related disorders. The data generated from this research proposal will form the basis for an R01 grant application at the end of the K01 award.

描述（由申请人提供）：这是Baran a . Ersoy博士的K01奖申请，Baran a . Ersoy博士是布莱根妇女医院（BWH）和哈佛医学院（HMS）的医学讲师。Ersoy博士是葡萄糖和脂质代谢紊乱基础研究的年轻研究者。他的申请概述了他的短期计划，通过获得先进分子生物学和免疫学技术的经验，包括蛋白质组学的质谱分析和骨髓移植。该计划将以他以前的经验为基础，为他提供时间和培训，以建立新的研究技能、策略和项目，这将有助于他成为一名独立的科学家。他的长期目标是在学术环境中成为一名独立的研究者，研究驱动胰岛素抵抗和促进2型糖尿病和非酒精性脂肪性肝病（NAFLD）的机制。为了实现这些目标，Dr. Ersoy将得到Dr. David E. Cohen的指导，David E. Cohen是HMS医学教授和BWH肝病学主任。Ersoy博士还组建了一个咨询委员会，由HMS细胞生物学教授Steven P. Gygi博士组成，他专注于蛋白质组学的质谱分析；Umut Ozcan博士是内分泌学家，擅长内质网应激、炎症和胰岛素信号传导；以及胰岛素信号专家莫里斯·f·怀特（Morris F. White）博士，他在发现胰岛素受体底物蛋白方面发挥了重要作用。Ersoy博士的导师和咨询委员会成员建立了指导和培训记录，其以前的学员在世界各地担任独立调查员职位。Ersoy博士将在BWH和HMS进行培训，这将提供一个智力丰富的研究环境，并长期致力于培养年轻科学家从事独立的学术研究事业。肥胖诱导的胰岛素抵抗是诱发NAFLD和2型糖尿病的主要病理生理缺陷。由于目前的管理选择仍然有限，确定控制胰岛素代谢反应的新调节机制应该有助于确定药物干预的新机会。本研究计划的目的是阐明葡萄糖稳态调节的基本新机制。其基本原理是，确定一种调节肝脏胰岛素敏感性的新机制可能导致确定治疗NAFLD和2型糖尿病的新治疗靶点。在大量初步数据的指导下，本研究计划的中心假设是磷脂酰胆碱转移蛋白（PC-TP）通过调节内质网（ER）应激和炎症，直接或间接地控制胰岛素受体底物2 （IRS2）的活性，从而调节葡萄糖稳态。这将在三个具体目标中进行测试：1)定义PC-TP抑制IRS2活性的机制；2)确定内质网应激在PC- tp介导的胰岛素信号调节中的作用；3)阐明免疫应答在pc - tp介导的胰岛素敏感性抑制中的作用。在Aim 1中，将使用PC-TP的小分子抑制剂（化合物A1）和IRS2缺陷小鼠来检测IRS2在PC-TP调节胰岛素信号传导中的作用。利用Pctp+/+和Pctp-/-小鼠的蛋白质组学质谱，以及从这些小鼠中收获的经化合物a1处理的肝细胞，将揭示PC-TP控制IRS2抑制的生物学途径。在Aim 2中，将使用Pctp+/+和Pctp-/-小鼠以及从这些小鼠中获取的肝细胞，通过免疫印迹分析确定PC-TP是否调节调节胰岛素信号和IRS2活性的关键内质网应激途径。内质网应激信号将被小鼠饥饿24小时后的重新喂食和细胞培养系统中的化学试剂脲霉素激活。目的3将利用从Pctp+/+和Pctp-/-小鼠中获取的腹膜巨噬细胞，并将Pctp-/-供体细胞骨髓移植到Pctp+/+受体小鼠中。小鼠高脂饮食喂养和细胞培养中脂多糖处理均可引起炎症。通过ELISA、免疫印迹和qPCR分析检测炎症信号。预计PC-TP将通过直接调节IRS2活性和调节内质网应激和炎症来调节胰岛素信号。这一点很重要，因为IRS2表达的缺失和内质网应激和炎症的增加与胰岛素抵抗的发展有关。这些研究有望为胰岛素抵抗和相关疾病的治疗找到新的治疗靶点。该研究计划产生的数据将在K01奖励结束时成为R01资助申请的基础。