Genetic and Proteomic Studies of Lipid Metabolism in Zebrafish

斑马鱼脂质代谢的遗传和蛋白质组学研究

• 批准号: 7442221
• 负责人: AMNON SCHLEGEL
• 金额: $ 11.88万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7442221
• 关键词: Abetalipoproteinemia; Adverse effects; Animal Model; Biochemical; Biological Assay; Blood Glucose; Cardiovascular Diseases; Caring; Chemicals; Chronic Kidney Failure; Complement; Consumption; Cultured Cells; Daily; Defect; Degenerative polyarthritis; Deposition; Development; Diabetes Mellitus; Dietary Fats; Disease; Embryo; Endocrinologist; Enzymes; Fatty Liver; Fatty acid glycerol esters; Foundations; Gene Expression; Gene Proteins; Genes; Genetic; Genetic Screening; Global Change; Healthcare Systems; Hereditary Disease; Human; Hyperlipidemia; Hypertension; Hypotension; Immunoblotting; Individual; Intervention Trial; Investigation; Kinetics; Lead; Life; Life Expectancy; Lipids; Lipoproteins; Liver diseases; Malabsorption Syndromes; Malignant Neoplasms; Mass Spectrum Analysis; Measurement; Metabolic; Metabolism; Methods; Microscopic; Modeling; Modern 1601-history; Monitor; Morbidity - disease rate; Mutagenesis; Mutate; Mutation; Natural History; Obesity; Obesity associated disease; Obstructive Sleep Apnea; Organ; Overweight; Pathologic; Pathway interactions; Patients; Phenotype; Physicians; Physiological; Polycystic Ovary Syndrome; Prevalence; Principal Investigator; Proteins; Proteomics; Public Health; Quality of life; Rate; Relative (related person); Risk; Scientist; Serum; Staining method; Standards of Weights and Measures; Therapeutic Effect; Time; Tobacco; Tobacco use; Transport Process; Triglycerides; United States; Upper arm; Vertebrates; Whole Organism; Yolk Sac; Zebrafish; absorption; career; cost; day; energy balance; feeding; gel electrophoresis; hypercholesterolemia; improved; knock-down; lipid I; lipid metabolism; lipid transport; microsomal triglyceride transfer protein; mortality; mutant; non-alcoholic fatty liver; novel; novel strategies; novel therapeutics; pandemic disease; particle; positional cloning; prevent; programs; protein expression; research study; two-dimensional; uptake

DESCRIPTION (provided by applicant): As an Endocrinologist, I see daily the devastating impacts obesity, hyperlipidemia, and diabetes mellitus have on patients. These three diseases are reaching pandemic status in the United States, and novel therapeutic candidates are needed to comabt [sic] these grave risks to the public health. In order to identify pharmacologically amenable targets in these three related diseases, I propose to use zebrafish in forward genetic and unbiased proteomic studies of lipid absorption, transport, and metabolism. In the first part of this project, I shall develop a simple and reliable method for staining and tracking yolk-derived and dietary lipids in the transparent zebrafish embryo. With this method, I shall confirm the suitability of the model organism for studying lipid metabolism by examining the effect of targeted knock-down of the evolutionary central and physiologically critical lipoprotein particle-packaging enzyme microsomal triglcyeride [sic] transfer protein (Mtp). Biochemical, microscopic, and physiologic assays shall be developed in order to compare the effects of decreasing zebrafish Mtp on development and dietary lipid absorption with those of abetalipoproteinemia, the human hereditary disease in which Mtp is mutated. In the second part of the project, an unbiased, forward genetic screen shall be performed to identify mutants that have alterations in lipid uptake, transport, and storage. The genetic screen shall rely on the methods developed in the first part of the project to identify and fully characterize the defects in lipid metabolism. Mutants that model human diseas [sic] states characterized by malabsorption of dietary lipid, altered kinetics of lipid consumption, and inappropriate deposition of lipids in organs (i.e., hepatic steatosis and intramyocellular liposis) shall be identified, and the genes giving rise to these mutant phenotypes shall be identified using standard positional cloning methods. The final arm of the study shall examine global changes in expression of proteins following high fat feeding. I shall use proteomic methods to find proteins whose expression is altered by high fat feeding, and employ mass spectrometric methods to identify the differentially regulated proteins. As with the genes identified in the forward genetic screen, the proteins identified in these proteomic studies shall be studied in order to develop novel strategies to treat obesity, hyperlipidemia and diabetes mellitus. These studies are well suited to the development of my career as an academic physician-scientist and they shall help combat the looming public health crisis.

描述（由申请人提供）： 作为一名内分泌科医生，我每天都会看到肥胖、高脂血症和糖尿病对患者造成的破坏性影响。这三种疾病在美国已达到大流行状态，需要新的候选治疗药物来应对这些对公共卫生的严重风险。为了确定这三种相关疾病的药理学靶标，我建议使用斑马鱼进行脂质吸收、转运和代谢的正向遗传和无偏蛋白质组学研究。在该项目的第一部分中，我将开发一种简单可靠的方法来染色和跟踪透明斑马鱼胚胎中的卵黄来源和膳食脂质。通过这种方法，我将通过检查靶向敲低进化中心和生理关键脂蛋白颗粒包装酶微粒体甘油三酯[原文如此]转移蛋白（Mtp）的效果来确认模型生物体用于研究脂质代谢的适用性。应开发生化、显微镜和生理测定，以比较斑马鱼 Mtp 减少对发育和膳食脂质吸收的影响与无β脂蛋白血症（Mtp 突变的人类遗传性疾病）的影响。在该项目的第二部分中，将进行公正的正向遗传筛选，以鉴定在脂质摄取、运输和储存方面发生改变的突变体。遗传筛选应依靠项目第一部分开发的方法来识别和充分表征脂质代谢缺陷。应鉴定以膳食脂质吸收不良、脂质消耗动力学改变和脂质在器官中不适当沉积（即肝脂肪变性和肌细胞内脂肪变性）为特征的人类疾病模型突变体，并应使用标准定位克隆方法鉴定产生这些突变表型的基因。该研究的最后一部分将检查高脂肪喂养后蛋白质表达的整体变化。我将使用蛋白质组学方法来寻找表达因高脂肪喂养而改变的蛋白质，并使用质谱方法来鉴定差异调节的蛋白质。与正向遗传筛选中鉴定的基因一样，应研究这些蛋白质组学研究中鉴定的蛋白质，以便开发治疗肥胖、高脂血症和糖尿病的新策略。这些研究非常适合我作为学术医师科学家的职业发展，它们将有助于应对迫在眉睫的公共卫生危机。