Hematopoietic Development From ES Cells

描述(申请人提供)：考虑到早期胚胎的不可获得性，调节人类造血承诺和发育的早期阶段的事件还知之甚少。胚胎干细胞(ES)在培养中分化并概括负责早期血细胞发育的关键程序的能力为更好地了解这些过程提供了一个强大的模型。对小鼠ES细胞的研究表明，ES细胞衍生的类胚体(EBS)的造血发育调控与早期胚胎相似。这项计划中的实验将把小鼠系统的发现转化为人类ES细胞分化，最终目标是建立一个模型，研究人类造血发育的最早阶段。在这项建议的第一个目标中，将在发育中的人类EBS中描述不同的造血发展阶段。这些分析是必不可少的，因为它们将定义产生人类造血系统的发育程序。第二个目标的重点是定义调节人类EBS中血细胞发育的分子。揭示这一过程中的关键因素是重要的，因为这些发现将为早期人类造血发育的调控提供第一个洞察力。此外，这些信息将使我们能够为EBS内的造血发展建立最佳方案。该提案的第三个目标是识别和表征人类胚胎干细胞(EBS)中的长期再生干细胞(LTRSC)。虽然这些研究具有挑战性，但这些研究是重要的，因为在这些文化中鉴定LTRSC将为研究调节其生成、自我更新和成熟的事件提供一个模式系统。此外，从EBS发展LTRSC将为移植提供一种新的来源。前三个目标的研究将在两个不同的细胞系WA01和ES02上进行。在最终目标中，我们将比较12个不同色调细胞系的造血潜能，以确定每种细胞系产生前三个目标中确定的群体的效率。以下线路将用于此目的：WA01、WA07、WA09、ES02、ES03、ES04、TE-03、TE-06、MI01、UC01、BG02、BG03。这些研究的发现将为人类造血系统的起源和发育调控提供新的重要信息，并由此确定血管母细胞、原始红系血统和最终造血的最早阶段。这些信息将为人类胚胎发育过程中中胚层诱导和造血规范的调节提供第一个洞察力。从人类ES细胞中产生LTRSC最终可以扩大造血干细胞移植治疗的患者数量和疾病谱。

DESCRIPTION (provided by applicant): The events regulating the earliest stages of hematopoietic commitment and development in humans are poorly understood, given the inaccessibility of the early embryo. The capacity of embryonic stem (ES) cells to differentiate in culture and recapitulate the critical programs responsible for early blood cell development provides a powerful model for gaining a better understanding of these processes. Studies with mouse ES cells have demonstrated that regulation of hematopoietic development in ES cell-derived embryoid bodies (EBs) is similar to that of the early embryo. The experiments in this proposal will translate findings from the mouse system to human ES cell differentiation with the ultimate goal of establishing a model for investigating the earliest stages of human hematopoietic development. In the first aim of this proposal, different stages of hematopoietic development will be characterized within the developing human EBs. These analyses are essential, as they will define the developmental program that gives rise to the human hematopoietic system. The focus of the second aim is to define the molecules that regulate blood cell development in human EBs. Uncovering the key factors in this process is important, as these findings will provide the first insights into the regulation of early human hematopoietic development. In addition, this information will enable us to establish optimal protocols for hematopoietic development within the EBs. The goal of the third aim of the proposal is to identify and characterize the long-term repopulating stem cell (LTRSC) in the human EBs. While challenging, these studies are important, as the identification of LTRSC in these cultures would provide a model system for investigating events that regulate their generation, self-renewal and maturation. In addition, the development of LTRSC from EBs would provide a novel source of these cells for transplantation. The studies in the first three aims will be carried out with the two different cell lines, WA01 and ES02. In the final aim, we will compare the hematopoietic potential of 12 different huES cell lines, to determine the efficiency of each to generate the populations identified in the previous three aims. The following lines will be used for this aim; WA01, WA07, WA09, ES02, ES03, ES04, TE-03, TE-06, MI01, UC01, BG02, BG03. Findings from these studies will provide new and important information on the origins and developmental regulation of the human hematopoietic system and in doing so will identify the hemangioblast, the primitive erythroid lineage and the earliest stages of definitive hematopoiesis. This information will provide the first insights into the regulation of mesoderm induction and hematopoietic specification during human embryonic development. Generation of LTRSC from human ES cells could ultimately expand the number of patients and the spectrum of diseases treated by hematopoietic stem cell transplantation.