Functional Analysis of Hematopoietic Stem Cell Origin

描述（由申请人提供）：此R21应用程序结合了 两个独立实验室的专业知识来解决一个关键问题 关于我们对造血个体发生的理解--发育起源 造血干细胞（HSC） 在小鼠胚胎中，原始造血祖细胞产生于卵黄囊 在原肠胚形成期间（E7.0-E8.5）。虽然定型血细胞在 从E.9.5开始的胎肝，HSC具有移植成人的能力 胚胎发育到E10时才开始出现异常。最近它又有了 “胚胎”HSC，其提供新生儿的长期植入，但不提供成人， 在E9.0时，在卵黄囊和胚胎中均鉴定出受体。它 尚不清楚这些HSC是否起源于胚内（AGM）或胚外 (yolk SAC）位点。在初步实验中，J. Palis发现 具有高增殖潜能的造血细胞仅在 循环前小鼠胚胎的卵黄囊（E8.25）。我们假设 胚胎HSC也在原肠胚形成期间从胚外位点产生。在 本提案的第一个目的是，我们将使用子宫内移植模型， 确定哺乳动物造血干细胞的发育起源。 囊细胞和胚胎固有细胞，来自E7.5 - E9.5将直接移植到E14.5 W 42胎仔的肝脏中。这 独特的移植系统将最大限度地提高我们识别HSC的能力， 在早期胚胎植入后的活动。在第二个目标中， 建议，我们将研究植入的时间和空间动力学， 造血干细胞是胚胎和胎儿造血器官的细胞。这些 实验将利用表型确定的E9.5卵黄囊， 表达增强的绿色荧光的E14.5肝供体干细胞群 蛋白 更好地理解哺乳动物造血的启动将 提供对HSC的个体发育，调节和扩展的见解，以及 白血病和骨髓衰竭综合征的起源。这些知识将 最终改善骨髓移植治疗 治疗先天性贫血，遗传性疾病和许多儿童和成人 癌的

DESCRIPTION (provided by applicant): This R21 application combines the expertise of two independent laboratories to address a pivotal question regarding our understanding of hematopoietic ontogeny- the developmental origin of hematopoietic stem cells (HSC). In the mouse embryo, primitive hematopoietic progenitors arise in the yolk sac during gastrulation (E7.0-E8.5). While definitive blood cells differentiate in the fetal liver beginning at E.9.5, HSC with the ability to engraft adult recipientsfirst arise within the embryo proper only at E10. Recently an "embryonic" HSC that provides long-term engraftment of newborn, but not adult, recipients has been identified at E9.0 in both yolk sac and embryo proper. It is not known if these HSC arise from intraembryonic (AGM) or extraembryonic (yolk sac) sites. In preliminary experiments, J. Palis has identified hematopoietic cells with high proliferative potential exclusively within the yolk sac of the pre-circulation mouse embryo (E8.25). We hypothesize that embryonic HSC also arise from extraembryonic sites during gastrulation. In the First Aim of this proposal, we will use an in utero transplantation model to determine the developmental origin of mammalian hematopoietic stem cells.Yolk sac and embryo proper cells from accurately staged mouse embryos between E7.5 - E9.5 will be transplanted directly into the livers of E14.5 W42 fetuses. This unique transplantation system will maximize our ability to identify HSC activity within the early post-implantation embryo. In the Second Aim of this proposal, we will study the temporal and spatial kinetics of engraftment of hematopoietic organs by embryonic and fetal hematopoietic stem cells. These experiments will take advantage of phenotypically defined E9.5 yolk sac and E14.5 liver donor stem cell populations expressing enhanced green fluorescent protein A better understanding of the initiation of mammalian hematopoiesis will provide insights into the ontogeny, regulation and expansion of HSC, as well as the origin of leukemias and bone marrow failure syndromes. This knowledge will ultimately lead to improvements in bone marrow transplantation for the curative treatment of congenital anemias, genetic diseases and many childhood and adult cancers.