Role of Ceramide in Hepatic Stellate Cell Activation and Liver Fibrosis

神经酰胺在肝星状细胞活化和肝纤维化中的作用

• 批准号: 10222658
• 负责人: Jennifer Y. Chen
• 金额: $ 18.36万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-01 至 2023-08-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10222658
• 关键词: Address; Advisory Committees; Amino Acids; Award; Basic Science; Carbon Tetrachloride; Cell Differentiation process; Cells; Ceramides; Chemicals; Choline; Cicatrix; Complement; Cytoplasm; Data; Development; Disease Progression; Ensure; Environment; Enzyme Inhibition; Enzymes; Extracellular Matrix; Farber' s lipogranulomatosis; Fibroblasts; Fibrosis; Funding; Gastroenterologist; Gene Expression; General Hospitals; Genes; Genetic; Genetic Transcription; Goals; Hepatic Fibrogenesis; Hepatic Stellate Cell; High Fat Diet; Human; Immunoblotting; Knowledge; Life; Lipids; Liver Failure; Liver Fibrosis; Liver diseases; Massachusetts; Mechanics; Mediating; Mentorship; Metabolism; Methodology; Mission; Modeling; Molecular; Molecular Biology; Mus; Myofibroblast; Nortriptyline; Pathway interactions; Patients; Pharmacology; Phenotype; Phosphorylation; Physicians; Production; Proteins; Public Health; Public Health Schools; Publications; RNA Interference; Research; Research Personnel; Rest; Role; Scientist; Signal Pathway; Signal Transduction; Sphingolipids; Supervision; Techniques; Therapeutic; Time; Training; Transgenic Mice; Tricyclic Antidepressive Agents; United States National Institutes of Health; Work; career; career development; cell type; chronic liver disease; chronic liver injury; collaborative environment; college; experience; fibrogenesis; galactosylgalactosylglucosylceramidase; genome-wide; genome-wide analysis; improved; in vivo; in vivo Model; liver injury; medical schools; mouse model; multidisciplinary; mutant; non-alcoholic fatty liver disease; novel; novel therapeutics; prevent; programs; response; small molecule; small molecule inhibitor; success; supportive environment; targeted treatment

PROJECT SUMMARY/ABSTRACT This is an application for a K08 award for Dr. Jennifer Y. Chen, a gastroenterologist and hepatologist at the Massachusetts General Hospital (MGH). Dr. Chen’s long-term career goal is to become an independently funded physician scientist, devoting more than 75% of her time to establish and maintain a basic science research program in hepatic fibrosis. This K08 award will provide Dr. Chen with the support necessary to achieve her short-term goals: 1) develop additional training in molecular biology, including investigating protein interactions and induction of stable gene expression; 2) gain expertise in genome-wide transcriptional analysis; 3) become proficient in transgenic mouse work and mouse models of hepatic fibrosis; and 4) produce the data and publication record necessary for a successful R01 application. In prior work, the candidate has developed a small molecule screen to identify compounds that inactivate hepatic stellate cells (HSCs), the primary cell type responsible for hepatic fibrosis. Through analysis of a hit, she identified that the sphingolipid ceramide can profoundly inhibit the activated effector phenotype of HSCs. Her work has identified a potential antifibrotic role for inhibition of acid ceramidase (aCDase), the enzyme responsible for ceramide metabolism. In this proposal, the candidate seeks to elucidate the mechanism by which ceramide accumulation inactivates HSCs and define the role of aCDase in fibrosis progression. The specific goals of the study are to: 1) determine how ceramide mediates HSC inactivation; and 2) define the impact of aCDase depletion and inhibition on fibrosis development in vivo. Under the first aim, the applicant will utilize a combination of techniques including immunoblotting, small molecule inhibition and activation, RNA interference, and expression of constitutively activated mutant proteins to understand how ceramide inactivates HSCs. Genome-wide expression analysis will be performed to elucidate transcriptional targets of ceramide. Under the second aim, the applicant will generate mice with a conditional fibroblast-specific deletion of aCDase, and will determine the extent to which they experience reduced fibrosis in two independent models of hepatic fibrosis. She will also determine the antifibrotic effects of a small molecule inhibitor of aCDase in vivo. As an integral part of this proposal, the candidate’s career development will be complemented by participation in advanced coursework and research seminars to develop expertise in molecular biology, genome-wide transcriptional analysis, and in vivo models of hepatic fibrosis. A formal mentorship committee and advisory team will provide supervision, guidance, and assistance for the candidate to achieve her goals. The research environment, which includes the MGH GI Unit, Harvard Fibrosis Network, Harvard College, Harvard Medical School, and the Harvard School of Public Health, will provide a rich, collaborative, and supportive atmosphere to ensure the candidate’s success. Through this award, the candidate will become an independent basic science investigator by contributing to the understanding of the role of the sphingolipid ceramide and its inactivation of HSCs ex vivo and in vivo.

项目摘要/摘要 这是为Jennifer Y.Chen博士申请K08奖项的申请表，她是一名胃肠病专家和肝病专家 麻省总医院(MGH)。陈博士的长期职业目标是成为一名独立的 资助内科科学家，75%以上的时间致力于建立和维护一门基础科学 肝纤维化研究计划。这一K08奖项将为陈博士提供必要的支持， 实现她的短期目标：1)发展额外的分子生物学培训，包括研究蛋白质 相互作用和诱导稳定的基因表达；2)获得全基因组转录分析的专业知识； 3)精通转基因小鼠的工作和肝纤维化的小鼠模型；4)制作数据 以及成功申请R01所需的发布记录。在之前的工作中，应聘者已经开发出 一种小分子筛选以鉴定使肝星状细胞(HSCs)失活的化合物，HSCs是主要细胞 导致肝纤维化的类型。通过对一次命中的分析，她发现神经鞘磷脂神经酰胺可以 深刻抑制HSCs激活的效应表型。她的研究已经确定了一种潜在的抗纤维化作用。 用于抑制酸性神经酰胺酶(ACDase)，负责神经酰胺代谢的酶。在这份提案中， 候选人试图阐明神经酰胺积累使HSCs失活的机制，并确定 ACDase在纤维化进展中的作用。这项研究的具体目标是：1)确定神经酰胺如何 介导HSC失活；以及2)确定aCDase耗竭和抑制对纤维化的影响 体内发育。在第一个目标下，申请者将利用一系列技术，包括 免疫印迹、小分子抑制和激活、RNA干扰和组成型表达 激活突变蛋白以了解神经酰胺如何使肝星状细胞失活。全基因组表达分析 将被用来阐明神经酰胺的转录靶标。在第二个目标下，申请人将 用条件成纤维细胞特异性缺失aCDase的小鼠，并将确定在多大程度上 他们在两个独立的肝纤维化模型中经历了纤维化减轻。她还将决定 一种aCDase小分子抑制剂的体内抗纤维化作用。作为这项提案的一个组成部分， 候选人的职业发展将通过参与高级课程和研究来补充 发展分子生物学、全基因组转录分析和活体模型方面的专业知识的研讨会 肝纤维化的症状。一个正式的指导委员会和咨询团队将提供监督、指导和 帮助候选人实现她的目标。研究环境，其中包括MGH GI股， 哈佛大学纤维化网络、哈佛学院、哈佛医学院和哈佛公共卫生学院， 将提供丰富、协作和支持的氛围，以确保候选人的成功。通过这件事 获奖后，候选人将成为一名独立的基础科学研究员，为 了解鞘磷脂神经酰胺的作用及其在体内和体外对HSCs的灭活作用。