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: 10242117
• 负责人: Raquel Espín Palazón
• 金额: $ 15.09万
• 依托单位: IOWA STATE UNIVERSITY
• 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 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10242117
• 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 Specification; Hematopoietic Stem Cell Transplantation; Hematopoietic stem cells; Hemoglobinopathies; Human; 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; Organism; Outcome; PGRN gene; Patients; Phase; Play; Positioning Attribute; Postdoctoral Fellow; Process; Property; Protocols documentation; Regulator Genes; Reporting; Research; Research Personnel; Role; Signal Transduction; Sorting - Cell Movement; Source; Specific qualifier value; Stem Cell Development; System; TNF gene; Techniques; Testing; Therapeutic; To specify; Training; Transgenic Organisms; Visualization; Work; Zebrafish; base; career; career development; cell regeneration; cytokine; experimental study; genetic manipulation; genome editing; hematopoietic stem cell emergence; hematopoietic stem cell fate; hemogenic endothelium; human disease; improved; in vivo; induced pluripotent stem cell; leukemia; mutant; notch protein; novel; pgRNA; reconstitution; recruit; regenerative therapy; scientific atmosphere; self renewing cell; skills; spatiotemporal; stem cell biology; stem cells; 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（HSCs）是人类骨髓中的罕见细胞，负责 所有血细胞谱系的终身补充和骨髓移植的疗效。 人类诱导多能干细胞的创造为细胞再生疗法带来了巨大希望，但 目前不可能指导这些细胞在体外产生HSC。此应用程序的目标是 确定通过NF-κ B的促炎信号如何指导脊椎动物胚胎中的HSC命运，以及这是如何 这一过程由颗粒蛋白前体a（Pgrna）调节，最终目标是在体外复制HSC生成， 临床应用我的工作假设是，早期促炎性输入汇聚激活NF-κ B， turn激活HSC命运规范中的关键信号事件。这些促炎信号需要 在HSC特化后不久下调;我的初步结果表明Pgrna以这种方式起作用。 为了验证这些假设，本研究的目的有两个：（1）研究NF-κ B在HSC中的作用 （2）鉴定Pgrna在HSC出现中的作用。这项研究将在斑马鱼中进行， 这是一个理想的系统，用于直接可视化的HSC，并已作为一种模式生物，研究人类 疾病为了实现本申请的目标，将产生新的转基因和突变株系，并且qPCR， 将使用FACS分选、RNA-seq和共聚焦显微镜技术。 埃斯平·帕拉松博士是加州大学圣地亚哥分校大卫·特拉弗实验室的博士后研究员， 我的目标是在一家大型研究机构领导一个专注于干细胞的独立研究小组。她的短- 长期目标是（1）确定造血内皮细胞内NF-kB的时空需求，和 阐明Pgrna如何调控HSC的出现。 由于她在免疫学和斑马鱼方面有很强的背景， 动物模型本提案中概述的项目将允许Espin Palazon博士从一个受指导的 科学职位，以独立的研究生涯，帮助她获得专业知识，在流式细胞仪分选，RNA-seq， 基因组编辑技术和HSC生物学，所有这些都是建立她独立实验室的基础。博士 Espin Palazon将在2015年与发育、免疫学和血液学专家会面并展示她的工作。 除了将她的数据提交给由资深专家组成的正式指导委员会， 转型为独立研究员。UCSD提供了许多课程，埃斯平·帕拉松博士将参加，因为 以及有关职业发展和实验室管理的研讨会。充满活力的，协作的科学 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，并将有助于形成这些研究的未来成功的基础。她很 准备好执行拟议的工作，实现她的职业发展和培训目标，并作出贡献高 对科学界的影响。