Deconstructing the Hematopoietic Stem Cell Niche

解构造血干细胞生态位

• 批准号: 9766280
• 负责人: Elliott Jennings Hagedorn
• 金额: $ 15.12万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 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 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9766280
• 关键词: Anatomy; Animal Model; Area; Award; Bioinformatics; Blood Cells; Bone Marrow; Boston; Cell Adhesion; Cell Adhesion Molecules; Cell Communication; Cell Lineage; Cells; Clustered Regularly Interspaced Short Palindromic Repeats; Data; Data Set; Disease; Dyes; Embryo; Engraftment; Environment; Erythrocytes; Fetal Liver; Future; Gene Expression; Gene Expression Profiling; Genes; Genetic; Goals; Grant; Hematological Disease; Hematologist; Hematopoietic; Hematopoietic Stem Cell Transplantation; Hematopoietic stem cells; Human; Imagery; Individual; Investigation; Label; Laboratories; Life; Location; Maintenance; Maps; Marrow; Mediating; Mentors; Molecular; Organism; Oxidative Stress; Pediatric Hospitals; Phase; Population; Postdoctoral Fellow; Production; RNA; Reactive Oxygen Species; Reporter; Research; Research Training; Resolution; Role; Scientist; Signal Transduction; Source; Stem cells; Stress; Stromal Cells; System; Techniques; Testing; Tissue-Specific Gene Expression; Tissues; Transgenes; Transplantation; Vascular Endothelial Cell; Work; Zebrafish; base; blood treatment; bone; cell type; curative treatments; experimental study; gene discovery; hematopoietic engraftment; hematopoietic stem cell niche; hematopoietic stem cell self-renewal; hematopoietic tissue; improved; live cell imaging; loss of function; macrophage; medical schools; neutrophil; new technology; new therapeutic target; novel; overexpression; promoter; selenoprotein; therapeutic target; tomography; transcriptome sequencing

Project Summary/Abstract The hematopoietic stem cell (HSC) niche is a specialized microenvironment that supports the life-long self- renewal of HSCs and their ability to produce all blood cell lineages. A number of different cell types and molecular factors have been associated with HSC niches in the mammalian bone marrow. To date, however, the challenges of directly visualizing the bone marrow have precluded a rigorous, systematic investigation of the cell-cell interactions that promote the niche engraftment of HSCs. Zebrafish offer an unparalleled setting in which the dynamic interactions between HSCs and their supporting niche cells can be experimentally manipulated and directly visualized at a resolution and throughput not possible in any other organism. The overarching goal of this proposal is to use live cell imaging in the zebrafish embryo, in combination with new technologies in gene expression analysis and tissue-specific gene disruption, to elucidate novel cellular and molecular mechanisms that are required for the engraftment of HSCs within their niche. The goal of the first aim is to integrate multiple spatial and tissue-specific gene expression datasets to generate a comprehensive map of gene expression within the zebrafish caudal hematopoietic tissue (CHT), a transient HSC niche akin to the mammalian fetal liver. The resulting map of cell-specific gene expression will be used to guide functional investigations of the cell-cell interactions that are required for HSC engraftment within the CHT. This gene expression data will be used to generate a reporter transgene that specifically labels stromal cells in the CHT. In addition, this gene expression data will be used, in combination with a tissue-specific CRISPR system, to identify the cellular adhesion molecules that mediate macrophage-HSC interactions and are required for HSC engraftment in the CHT. The second aim will investigate the effects of oxidative stress on the HSC niche within the CHT. This will be done by treating embryos with compounds that induce oxidative stress in combination with a live cell dye that permits visualization of reactive oxygen species (ROS). Additionally, the function of two genes (sepp1a and nrros), with known roles in maintaining low ROS levels, will be investigated using loss-of- function and overexpression experiments. These studies could identify novel therapeutic targets to enhance the engraftment and maintenance of HSCs during transplantation-based treatment of blood disorders. As a postdoctoral fellow, Dr. Hagedorn will conduct his research in the laboratory of Dr. Leonard Zon, a renowned hematologist and stem cell biologist. Building on a strong background in genetic and live cell imaging, Dr. Hagedorn will expand his expertise to include new technologies in spatial gene expression and tissue-specific gene disruption. Under the guidance of Dr. Zon and an exceptional mentoring committee, Dr. Hagedorn has constructed a rigorous research and training plan that will allow him to succeed in the mentored and independent phases of the award. The environment at Boston Children's Hospital and Harvard Medical School will provide the ideal surroundings for Dr. Hagedorn to become a successful independent scientist.

项目总结/摘要 造血干细胞（HSC）生态位是一个专门的微环境，支持终身自我造血。 HSC的更新及其产生所有血细胞谱系的能力。许多不同的细胞类型， 分子因子与哺乳动物骨髓中的HSC小生境有关。然而，迄今为止， 直接观察骨髓的挑战已经排除了对骨髓的严格的、系统的研究， 细胞间相互作用促进造血干细胞的小生境植入。斑马鱼提供了一个无与伦比的环境， HSC与其支持的小生境细胞之间的动态相互作用可以通过实验来确定。 操作和直接可视化的分辨率和吞吐量在任何其他生物体中是不可能的。的 这项提议的首要目标是在斑马鱼胚胎中使用活细胞成像，结合新的 基因表达分析和组织特异性基因破坏技术，以阐明新的细胞和 HSC在其小生境内植入所需的分子机制。第一个目标 目的是整合多个空间和组织特异性基因表达数据集， 斑马鱼尾部造血组织（CHT）内的基因表达图，CHT是一种类似于 哺乳动物的胎儿肝脏由此产生的细胞特异性基因表达图谱将用于指导功能性基因表达。 研究CHT内HSC植入所需的细胞-细胞相互作用。该基因 表达数据将用于产生特异性标记CHT中基质细胞的报告转基因。 此外，该基因表达数据将与组织特异性CRISPR系统结合使用，以 鉴定介导巨噬细胞-HSC相互作用和HSC所需的细胞粘附分子 在CHT中移植。第二个目的是研究氧化应激对HSC生态位的影响， CHT。这将通过用诱导氧化应激的化合物联合处理胚胎来实现 与活细胞染料，允许可视化的活性氧（ROS）。此外，两个功能 基因（sepp 1a和nrros），与已知的作用，在维持低活性氧水平，将研究使用损失- 功能和过表达实验。这些研究可以确定新的治疗靶点， 在基于移植的血液疾病治疗期间HSC的植入和维持。 作为一名博士后研究员，哈格多恩博士将在伦纳德宗博士的实验室进行他的研究， 著名的血液学家和干细胞生物学家建立在遗传和活细胞的强大背景之上 哈格多恩博士将扩大他的专业知识，包括空间基因表达的新技术， 组织特异性基因破坏。在Zon博士和一个特殊的指导委员会的指导下， 哈格多恩已经建立了一个严格的研究和培训计划，这将使他能够成功地在指导 独立的奖项。波士顿儿童医院和哈佛医学院的环境 学校将为哈格多恩博士成为一名成功的独立科学家提供理想的环境。