HEPATIC PROGENITORS IN TRANSPLANTATION

描述(由申请人提供)： 应聘者：这份职业发展提案旨在为我的学术生涯做好正式准备，成为一名免疫和细胞生物学领域的移植外科医生和独立研究员。培训计划由三个部分组成：正式的课程作业，其他教学内容，以及完成拟议研究计划目标的密集指导。在1-2年级：将选修两门免疫学高级课程和两门细胞生物学课程。作为补充，在第一年将有一个密集的研讨会，它将扩展我在分子生物学方面的基本科学技术。每周的实验室会议和期刊俱乐部的活动将持续进行。在过去的两年里，我一直专注于学习分离干细胞的原理，这项前期工作是研究计划的一部分。我将与两位导师密切合作，以更广泛地了解分离肝脏干细胞所需的关键实验。这个结构化的项目将使我准备好批判性思考、提出问题、形成假设，并随后在移植和细胞生物学领域进行创新实验。最后，作为一名移植外科医生，我的临床培训是对我的科学追求的理想补充，因为我对开发解决相关临床问题的翻译项目有洞察力。 环境：北卡罗来纳大学在校园内有多个核心实验室设施，可以完成这些和未来的实验。它们包括荧光激活的细胞分选设施、胃肠道和胆道疾病生物学中心，其中包括先进细胞技术和组织工程核心、免疫分析核心和共焦设施。这项提议得到了外科部门的支持，并允许PI将75%的时间投入到这个项目中。大部分工作将在PI的实验室进行。 研究：随着挑战器官移植的器官持续短缺，以及新鲜分离的成熟肝细胞在肝细胞治疗中缺乏成功，我提出了一系列基于肝脏祖细胞(具有增殖能力)的实验，作为一种有希望的替代方案。为了验证这一假设，我将确定肝祖细胞在移植后的扩张和功能。研究将集中在它们的免疫原性上，并将为未来的实验提供基础。 其具体目的是1)鉴定肝脏富含转录因子和肝细胞谱系基因在肝祖细胞扩增过程中的表达序列；2)利用绿色荧光蛋白(Green)转基因小鼠确定肝祖细胞的植入、迁移模式和随后的细胞分化；3)利用精选的转基因小鼠模型(即神经鞘磷脂酶缺陷、高胆固醇血症)，我将分析肝祖细胞的功能及其在损伤后重建肝脏的能力；4)分析CD4+、CD8+和非T细胞启动同种异体肝祖细胞急性排斥反应的能力。

DESCRIPTION (provided by applicant): Candidate: This career development proposal is designed to complete formal preparation for my academic career as a transplant surgeon and independent investigator in immuno- and cellular biology. The training program has three components: formal coursework, other didactics, and an intensive mentorship to complete the proposed research plan's aims. During years 1-2: two advanced courses in immunology and two in cellular biology will be taken. These will be complemented by an intensive workshop during year 1 that will expand my basic science techniques in molecular biology. Weekly laboratory meetings and journal club activities will be ongoing. During the past two years I have focused on learning the principles of isolating stem cells and this preliminary work is part of the research plan. I will be working closely with both mentors to develop a broader understanding of the critical experiments required to isolate hepatic stem cells. This structured program will prepare me to think critically, ask questions, form hypotheses and subsequently perform innovative experiments in the transplant and cell biology field. Lastly, my clinical training as a transplant surgeon is an ideal complement to my scientific pursuits, as I have insight into developing translational projects that address pertinent clinical questions. Environment: The University of North Carolina has multiple core laboratory facilities on campus that will allow the completion of these and future experiments. They include the Fluorescence Activated Cell Sorting facility, the Center for Gastrointestinal and Biliary Disease Biology that includes the Advanced Cell Technologies and Tissue Engineering Core, the Immunoassay Core, and the Confocal Facility. This proposal is supported by the Department of Surgery and allows the PI to dedicate 75% of his time on this project. The majority of the work will be carried out in the PI's lab. Research: With the ongoing shortage of organs that challenges organ transplantation and the lack of success with freshly isolated mature hepatocytes in liver cell therapy I have proposed a series of experiments based on hepatic progenitors (with their ability to proliferate), as a promising alternative. To test this hypothesis I will determine the hepatic progenitor's ability to expand and function after transplantation. Research will focus on their immunogenicity and will provide a foundation for future experiments. The specific aims are to 1) To identify the sequence of expression of liver-enriched transcription factors and hepatocyte lineage genes in hepatic progenitors during cellular expansion; 2) To determine the engraftment and migration pattern of hepatic progenitors and subsequent cellular differentiation using a green fluorescent protein (transgenic) "green" mouse; 3) Using selected transgenic mouse models (i.e. sphingomyelinase deficient, hypercholesterolemia) I will analyze the function of hepatic progenitor cells along with their ability to reconstitute a liver after injury; 4) Analyze the capacity of CD4+, CD8+ and non-T cells to initiate acute rejection of allogeneic hepatic progenitor cells.