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: 7295460
• 负责人: AMNON SCHLEGEL
• 金额: $ 11.78万
• 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/7295460
• 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)： 作为一名内分泌学家，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。这三种疾病在美国正达到大流行状态，需要新的治疗方案来消除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作用靶点，我建议使用斑马鱼对脂肪的吸收、运输和代谢进行正向遗传和无偏倚的蛋白质组学研究。在这个项目的第一部分，我将开发一种简单而可靠的方法来染色和跟踪透明斑马鱼胚胎中来自蛋黄和饲料的脂肪。用这种方法，我将通过检测进化中心和生理关键的脂蛋白颗粒包装酶微体三环素[SIC]转移蛋白(MTP)的靶向性下调的效果来确认模式生物是否适合研究脂代谢。应开展生化、显微镜和生理分析，以比较降低斑马鱼MTP对发育和膳食脂肪吸收的影响，以及降低MTP突变的人类遗传性疾病--脂蛋白血症的影响。在项目的第二部分，应该进行无偏见的正向遗传筛查，以识别在脂质吸收、运输和储存方面发生变化的突变株。基因筛查应依靠项目第一部分开发的方法来识别和充分描述脂类代谢缺陷。应识别模拟人类疾病状态的突变，其特征是膳食脂肪吸收不良，脂肪消耗动力学改变，以及器官中脂质的不适当沉积(即，肝脏脂肪变性和肌内脂肪沉积)，并应使用标准的位置克隆方法来鉴定导致这些突变表型的基因。这项研究的最后一环将检查高脂肪喂养后蛋白质表达的全球变化。我将使用蛋白质组学方法来寻找其表达被高脂肪喂养改变的蛋白质，并使用质谱学方法来鉴定差异调节的蛋白质。与正向遗传筛查中确定的基因一样，应研究这些蛋白质组研究中确定的蛋白质，以开发治疗肥胖症、高脂血症和糖尿病的新策略。这些研究非常适合我作为一名学术内科科学家的职业发展，它们将有助于对抗迫在眉睫的公共卫生危机。