The impact of inflammation on hematopoietic stem cell specification

炎症对造血干细胞规范的影响

• 批准号: 9432324
• 负责人: Raquel Espín Palazón
• 金额: $ 14.65万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-08 至 2022-09-07
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9432324
• 关键词: Address; Anemia; Animal Model; Aorta; Award; Blood; Blood Cells; Bone Marrow; Bone Marrow Transplantation; Cell Lineage; Cell Therapy; Cells; Cellular biology; Clinical; Communities; Confocal Microscopy; Cytokine Receptors; Data; Development; Discipline; Dorsal; Embryonic Development; Endothelium; Environment; Event; Focus Groups; Foundations; Future; Generations; Genes; Genetic; Goals; Hematological Disease; Hematology; Hematopoietic; Hematopoietic Stem Cell Research; Hematopoietic Stem Cell Transplantation; Hematopoietic stem cells; Hemoglobinopathies; Human; Imagery; Immunologics; Immunology; In Vitro; Individual; Inflammation; Inflammation Mediators; Inflammatory; Institution; Joints; Knowledge; Laboratories; Lead; Life; Mentors; Mentorship; Modeling; Molecular Biology; Multipotent Stem Cells; NF-kappa B; Natural regeneration; Organism; Outcome; PGRN gene; Patients; Phase; Play; Positioning Attribute; Postdoctoral Fellow; Process; Property; Protocols documentation; Recruitment Activity; Regulator Genes; Reporting; Research; Research Personnel; Role; Signal Transduction; Sorting - Cell Movement; Source; Specific qualifier value; Stem cells; System; TNF gene; Techniques; Testing; Therapeutic; To specify; Training; Transgenic Organisms; Work; Zebrafish; base; career; career development; cytokine; experimental study; genetic manipulation; genome editing; hematopoietic stem cell fate; human disease; improved; in vivo; induced pluripotent stem cell; leukemia; mutant; notch protein; novel; pgRNA; reconstitution; scientific atmosphere; self renewing cell; skills; spatiotemporal; success; tool; transcriptome sequencing; vertebrate embryos

Hematopoietic stem cells (HSCs) are rare cells within human bone marrow that are responsible both for the life-long replenishment of all blood cell lineages and for the curative effects of bone marrow transplantation. The creation of human induced pluripotent stem cells holds great promise for cellular regeneration therapies, but it is not currently possible to instruct these cells to generate HSCs in vitro. The goal of this application is to determine how pro-inflammatory signaling via NF-kB instructs HSC fate in the vertebrate embryo, and how this process is regulated by Progranulin a (Pgrna), with the ultimate goal of replicating HSC generation in vitro for clinical utility. My working hypothesis is that early pro-inflammatory inputs converge to activate NF-kB, which in turn activates key signaling events in the specification of HSC fate. These pro-inflammatory signals need to be downregulated soon after HSC specification; my preliminary results suggest that Pgrna functions in this manner. To test these hypotheses, this proposal consists of 2 aims: (1) Characterize the role of NF-kB in HSC specification; and (2) identify the role of Pgrna in HSC emergence. This study will be conducted in zebrafish, which are an ideal system for direct visualization of HSCs and have served as a model organism to study human disease. To achieve this application’s goals, novel transgenic and mutant lines will be generated, and qPCR, FACS-sorting, RNA-seq and confocal microscopy techniques will be utilized. Dr. Espin Palazon is a postdoctoral fellow in David Traver’s laboratory at UCSD whose ultimate career goal is to lead an independent research group focused on stem cells at a major research institution. Her short- term goals are (1) to determine the spatiotemporal requirements of NF-kB within hemogenic endothelium, and the downstream genes regulated to specify HSCs; and (2) to elucidate how Pgrna governs HSC emergence. She was recruited to join Dr. Traver’s group because of her strong background in immunology and the zebrafish animal model. The project outlined in this proposal will allow Dr. Espin Palazon to transition from a mentored scientific position to an independent research career, helping her gain expertise in FACS sorting, RNA-seq, genome editing techniques, and HSC biology, all of which are fundamental to establish her independent lab. Dr. Espin Palazon will meet and present her work to experts in development, immunology and hematology, in addition to presenting her data to a formal mentorship committee comprised of senior experts that will aid her transition to an independent researcher. UCSD offers numerous courses that Dr. Espin Palazon will attend, as well as seminars on career development and laboratory management. The vibrant, collaborative scientific atmosphere at UCSD is an ideal environment for Dr. Espin Palazon to develop during the mentored phase of her award, and will be instrumental in forming the foundation for the future success of these studies. She is well poised to execute the proposed work, achieve her career development and training goals and to contribute high impact research to the scientific community.

造血干细胞（HSC）是人类骨髓中稀有细胞，这两者都是 所有血细胞谱系的终身复制以及骨髓移植的治愈作用。 人类引起的多能干细胞的创建对细胞再生疗法具有巨大的希望，但 目前不可能指示这些细胞在体外产生HSC。该应用的目的是 通过NF-KB确定促炎性信号传导如何指示脊椎动物胚胎中的HSC命运，以及如何 过程受progranulin A（PGRNA）调节，其最终目标是在体外复制HSC生成 临床实用程序。我的工作假设是，早期的促炎性输入会融合激活NF-KB，这在 在HSC命运的规范中激活关键信号事件。这些促炎性信号必须是 HSC规格后不久下调；我的初步结果表明，PGRNA以这种方式起作用。 为了检验这些假设，该提案由2个目的组成：（1）表征NF-KB在HSC中的作用 规格; （2）确定PGRNA在HSC出现中的作用。这项研究将在斑马鱼进行 这是直接可视化HSC的理想系统，并已成为研究人类的模型生物 疾病。为了实现该应用程序的目标，将产生新颖的转基因和突变线，qpcr， 将利用FACS分级，RNA-SEQ和共聚焦显微镜技术。 Espin Palazon博士是UCSD的David Traver实验室的博士后研究员，他的最终职业生涯 目标是领导一个专注于主要研究机构干细胞的独立研究小组。她的短暂 术语目标是（1）确定血液内皮内NF-KB的时空要求，并且 调节的下游基因指定HSC； （2）阐明PGRNA如何控制HSC出现。 由于她的免疫学背景和斑马鱼，她被招募加入Traver博士的小组 动物模型。本提案中概述的项目将使Espin Palazon博士从事过渡 独立研究职业的科学地位，帮助她获得FACS分类的专业知识，RNA-Seq， 基因组编辑技术和HSC生物学，所有这些都是建立其独立实验室的基础。博士 Espin Palazon将与发展，免疫学和血液学专家见面并介绍她的工作， 除了将她的数据介绍给正式的高级委员会委员会完成，以帮助她 过渡到独立研究人员。 UCSD提供了许多Espin Palazon博士将参加的课程， 以及关于职业发展和实验室管理的半小伙子。充满活力的协作科学 UCSD的气氛是Espin Palazon博士在指导阶段发展的理想环境 她的奖项将有助于为这些研究的未来成功奠定基础。她很好 中毒以执行拟议的工作，实现她的职业发展和培训目标，并贡献很高 影响科学界的研究。