Dissection of Hematopoietic Stem and Progenitor Cell Differentiation during Embryogenesis

胚胎发生过程中造血干细胞和祖细胞分化的剖析

• 批准号: 10607210
• 负责人: Anastasia Nizhnik
• 金额: $ 4.77万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-16 至 2026-04-15
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10607210
• 关键词: Address; Adult; Affect; Agreement; Biological Assay; Blood; Blood Cells; CRISPR/Cas technology; Candidate Disease Gene; Cell Differentiation process; Cells; Clinical; Development; Dissection; Embryo; Embryonic Development; Endothelial Cells; Endothelium; Environment; Erythroid; Excision; Experimental Genetics; Gene Mutation; Generations; Genes; Genetic; Genetic Screening; Genomic approach; Goals; Growth; Hematologic Neoplasms; Hematological Disease; Hematopoiesis; Hematopoietic; Hematopoietic System; Hematopoietic stem cells; Impairment; In Vitro; Inherited; Kinetics; Knowledge; Label; Lymphoid; Lymphoid Cell; Malignant Neoplasms; Measures; Methods; Molecular; Mus; Mutagenesis; Myelogenous; Myeloid Progenitor Cells; Natural Killer Cells; Organism; Output; Patients; Pluripotent Stem Cells; Population; Process; Property; RUNX1 gene; Regenerative capacity; Replacement Therapy; Source; System; T-Lymphocyte; Therapeutic; Tissues; Transplantation; Umbilical Cord Blood; Zebrafish; cell behavior; cell type; genetic approach; granulocyte; hematopoietic differentiation; hematopoietic stem cell differentiation; hematopoietic stem cell formation; hemogenic endothelium; improved; in vivo; induced pluripotent stem cell; loss of function; molecular marker; mutant; novel; progenitor; regenerative; regenerative cell; reverse genetics; self-renewal; single-cell RNA sequencing; spatiotemporal; stem cell therapy; stem cells; tool; transcription factor; vertebrate embryos; zebrafish development

PROJECT SUMMARY Hematopoietic Stem Cells (HSCs) are clinically valuable because they display regenerative properties of self- renewal and the ability to produce an entire hematopoietic system through multilineage differentiation. In other words, HSC can make all mature blood cell types such as myeloid, erythroid, and lymphoid and they sustain those blood types long-term in the organism. These regenerative properties make them an idea therapeutic tool for treating hematological malignancies by transplantation from a healthy donor, however a shortage of donors has prompted the effort to generate HSCs from alternative sources such as patient derived iPSCs. Knowledge about molecular mechanisms of HSC formation is needed to aid those efforts. HSCs are formed in early embryonic development alongside other HSC-independent Progenitor Cells (HPCs). One major difference between them is that HPCs are more limited in their differentiation repertoire compared to HSCs. We aim to study the differences between HSCs and HPCs to further advance our understanding of the specific molecular drivers of HSC regenerative properties. Understanding how HSCs acquire the ability to differentiate and self- renew in development can aid the efforts for generating them in vitro from alternative sources such as patient derived tissues or cord blood sources. Current molecular markers are insufficient for distinguishing HSCs from all other HPC subtypes. Moreover, most experimental systems require the removal of these cells from their native environment to study their functionality using in vitro culture assays or transplantation, which will measure the potential of a cell but may not reflect its actual endogenous function. Genetic experiments in mice and zebrafish showed that HSC and HPC fate can be molecularly uncoupled, implying that unique drivers of HSC formation exist. What genes are responsible for the formation of the rare population of HSCs independently from HPCs? To address this question, we established a genetic lineage tracing approach that measures HSC and HPC differentiation without removing them from their native environment. Using this assay, we found that HSCs show a significant delay in their ability to differentiate to myeloid and lymphoid lineages relative to HPCs, and this finding agrees with others in the field. The assay allows us to assess HSC differentiation in vivo, uncoupled from HPCs. We propose to use this functional tool in combination with other molecular and genetic assays to address the following aims: (1) Determine the differentiation potential of HSCs and HPCs in vivo and (2) Perform a reverse genetic screen for regulators of HSC differentiation. Our findings can aid the efforts to generate HSCs from alternative sources for clinical use.

项目摘要 造血干细胞（HSC）在临床上是有价值的，因为它们表现出自我的再生特性 更新和通过多琳分化产生整个造血系统的能力。在其他 单词，HSC可以使所有成熟的血细胞类型（例如髓样，红细胞和淋巴样）持续 这些血型长期在生物体中。这些再生特性使它们成为一种想法的治疗工具 通过从健康供体移植中治疗血液学恶性肿瘤，但是捐助者短缺 已经促使努力从替代来源（例如患者衍生的IPSC）产生HSC。知识 需要关于HSC形成的分子机制来帮助这些努力。 HSC在早期形成 胚胎发育与其他非HSC独立的祖细胞（HPC）一起发育。一个主要区别 与HSC相比，它们之间的HPC在分化库中受到更大的限制。我们的目标 研究HSC和HPC之间的差异，以进一步提高我们对特定分子的理解 HSC再生特性的驱动因素。了解HSC如何获得区分和自我的能力 开发的续订可以帮助从替代来源（例如患者）中产生它们的努力 衍生的组织或脐带血源。当前的分子标记不足以区分HSC和 所有其他HPC亚型。此外，大多数实验系统都需要从其中去除这些单元 使用体外培养试验或移植研究其功能的天然环境，这将测量 细胞的潜力，但不能反映其实际的内源性功能。小鼠和 斑马鱼表明，HSC和HPC命运可以分子取消耦合，这意味着HSC的独特驱动因素 形成存在。哪些基因是造成HSC罕见人群形成的基因，而不是独立于 HPCS？为了解决这个问题，我们建立了一种遗传谱系追踪方法，以测量HSC和 HPC分化而不将其从本地环境中删除。使用此测定法，我们发现HSCS 显示出与HPC相对于HPC的髓样和淋巴样谱系区分开的能力明显延迟，并且 这一发现与该领域的其他人一致。该测定使我们能够评估体内HSC差异化，未偶联 来自HPC。我们建议将此功能工具与其他分子和遗传测定结合使用 解决以下目的：（1）确定体内HSC和HPC的分化潜力，（2）执行 HSC分化调节因子的反向遗传筛选。我们的发现可以帮助产生HSC的努力 从临床用途的替代来源。