Membrane-Based Mechanisms of Hepatic Insulin Resistance

肝胰岛素抵抗的膜机制

• 批准号: 8758031
• 负责人: Baran Ersoy
• 金额: $ 15.79万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 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 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8758031
• 关键词: Ablation; Activation Analysis; Advisory Committees; Affect; Applications Grants; Attenuated; Automobile Driving; Basic Science; Binding; Binding Proteins; Biological; Bone Marrow Transplantation; Boxing; 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; Fatty acid glycerol esters; Genetic; Glucose Metabolism Disorders; Goals; Harvest; Hepatic; Hepatocyte; Hepatology; Hospitals; Immune response; Immunoblotting; In Vitro; Inflammation; Inflammatory; Insulin; Insulin Receptor; Insulin Resistance; Interleukin-6; Intervention; JNK-activating protein kinase; JUN gene; Laboratories; Lead; Lecithin; Link; Lipid Binding; Lipids; Lipopolysaccharides; Liver; Liver diseases; 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; Phospholipids; 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; Tumor Necrosis Factor-alpha; Tunicamycin; Woman; base; biological adaptation to stress; blood glucose regulation; career; common treatment; cytokine; endoplasmic reticulum stress; experience; feeding; glucose metabolism; glucose production; human IRS2 protein; improved; inhibitor/antagonist; instructor; insulin sensitivity; insulin signaling; 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; protein expression; public health relevance; research study; response; skills; small molecule; 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博士的K 01奖申请。她是布里格姆妇女医院（BWH）和哈佛医学院（HMS）的医学讲师。Ersoy博士是一位年轻的研究者，从事葡萄糖和脂质代谢紊乱的基础研究。他的申请概述了一个短期计划，通过获得先进的分子生物学和免疫学技术，包括蛋白质组学和骨髓移植的质谱分析的经验，进一步他的教育。该计划将建立在他以前的经验，并为他提供时间和培训，以建立新的研究技能，战略和项目，这将有助于他成为一名独立的科学家后取得成功。他的长期目标是成为学术环境中的独立研究者，并研究驱动胰岛素抵抗和促进2型糖尿病和非酒精性脂肪肝（NAFLD）的机制。为了实现这些目标，Ersoy博士将由大卫E博士指导。科恩说，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，一位在内质网应激，炎症和胰岛素信号传导方面具有专长的内分泌学家;以及Morris F.白色，胰岛素信号专家谁发挥了主要作用，在发现胰岛素受体底物蛋白。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研究员职位。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导致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，确定新的调节机制，管理胰岛素的代谢反应，应有助于确定新的机会，药物干预。这项研究的目的是阐明一个基本的新机制，调节葡萄糖稳态。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）通过直接和间接调节内质网（ER）应激和炎症来控制胰岛素受体底物2（IRS 2）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IRS 2活性的机制; 2）确定ER应激在PC-TP介导的胰岛素信号传导调节中的作用; 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。在目标1中，将使用PC-TP的小分子抑制剂（化合物A1）和IRS 2缺陷小鼠来检查IRS 2在PC-TP调节胰岛素信号传导中的作用。将使用Pctp+/+和Pctp-/-小鼠以及从这些小鼠收获的化合物A1处理的肝细胞中的蛋白质组学质谱来揭示PC-TP控制IRS 2抑制的生物学途径。在目标2中，Pctp+/+和Pctp-/-小鼠和从这些小鼠收获的肝细胞将用于通过免疫印迹分析确定PC-TP是否调节调节胰岛素信号传导和IRS 2活性的关键ER应激途径。ER应激信号传导将通过在小鼠中饥饿24小时后再喂食以及通过细胞培养系统中的化学试剂衣霉素来激活。目的3利用从Pctp+/+和Pctp-/-小鼠收获的腹腔巨噬细胞，将Pctp-/-供体细胞骨髓移植到Pctp+/+受体小鼠中。在小鼠中通过高脂肪饮食喂养和在细胞培养物中通过脂多糖处理诱导炎症。将通过ELISA、免疫印迹和qPCR分析测量炎症信号。预期PC-TP将通过直接调节IRS 2活性以及通过调节ER应激和炎症来调节胰岛素信号传导。这是重要的，因为IRS 2表达的丧失和ER应激和炎症的增加与胰岛素抵抗的发展相关。这些研究有望为胰岛素抵抗和相关疾病的治疗确定新的治疗靶点。从这个研究计划产生的数据将形成在K 01奖结束时的R 01补助金申请的基础。