Hematopoietic Precursor Development from Human Embryonic Stem (ES) Cells

描述(申请人提供)：造血已在许多动物模型中被广泛研究，包括小鼠胚胎干细胞和小鼠胚胎。然而，对人类造血发育的分析通常使用从骨髓或脐带血中分离出的血细胞。虽然这些出生后的人类造血干细胞(HSCs)已经被很好地描述了，但关于这些HSCs是如何在人类发育的初始阶段从早期的前体细胞产生的，我们知道的很少。人类胚胎干细胞为研究人类发育生物学的基本问题以及确定稀有前体细胞的表型和遗传调控提供了一个最佳的起点。这项建议的总体目标是使用人类ES细胞来系统地表征人类造血祖细胞的发育。我们之前已经展示了从人类ES细胞中获得CD34+、CD45+和成熟的髓系细胞的方法。最近，我们还从CD34+人ES细胞来源的前体细胞中获得了淋巴细胞。虽然我们证明了人类ES细胞来源的造血细胞的有序发育，但对人类ES细胞造血所需的条件仍缺乏更完整的了解。为了更好地表征早期人类造血的基本细胞和分子机制，我们将做以下工作。1)确定从人胚胎干细胞发育为血细胞所必需的可溶性蛋白和细胞结合蛋白，尤其是Wnt蛋白。2)通过体外替代试验和移植到免疫缺陷小鼠体内，鉴定人ES细胞来源的HSCs的表型。3)分离作为血液和血管内皮细胞共同前体的血管母细胞或血源性内皮细胞的克隆性群体。这些目标的成功实现将极大地提高我们利用人类胚胎干细胞了解人类基本发育的能力，并将其重要应用于更好的基于干细胞的治疗。(这些研究将使用NIH注册的WA01和WA09人类ES细胞)。与公共健康相关：这些研究将对迅速增长的再生医学领域产生重要影响。更好地了解人类ES细胞的造血细胞发育将转化为用于治疗各种恶性和非恶性血细胞疾病的造血细胞移植的新细胞来源。这些细胞也可能被用作输血药物的新来源。

DESCRIPTION (provided by applicant): Hematopoiesis has been extensively studied in many animal models, including mouse embryonic stem (ES) cells and murine embryos. However, analyses of human hematopoietic development typically utilize blood cells isolated from bone marrow or umbilical cord blood. While these post-natal human hematopoietic stem cells (HSCs) have been well characterized, considerably less is known about how these HSCs arise from earlier precursor cells during the initial stages of human development. Human ES cells provide an optimal starting point to study fundamental questions of human developmental biology, and to define phenotypic and genetic regulation of rare precursor cells. The overall goal of this proposal is to use human ES cells to systematically characterize human hematopoietic precursor cell development. We have previously demonstrated methods to derive CD34+, CD45+ and mature myeloid-lineage cells from human ES cells. More recently, we have also derived lymphocytes from CD34+ human ES cell-derived precursor cells. While we demonstrate an ordered development of hematopoietic cells derived from human ES cells, more complete understanding of the conditions required for hematopoiesis from human ES cells is still lacking. To better characterize fundamental cellular and molecular mechanisms of early human hematopoiesis, we will do the following. 1) Define soluble and cell bound proteins essential for development of blood cells from human ES cells, with particular emphasis on Wnt proteins. 2) Characterize the phenotype of human ES cell derived HSCs by in vitro surrogate assays and transplantation into immunodeficient mice. 3) Isolate clonal populations of hemangioblast or hemogenic endothelial cells that serve as a common precursor to both blood and endothelial cell lineages. Successful completion of these aims will significantly advance our ability to use human ES cells to understand basic human development, with important applications to better stem-cell based therapies. (These studies will use NIH registry WA01 and WA09 human ES cells). Relevance to public health: These studies will have important impact in the rapidly growing area of regenerative medicine. Better understanding of hematopoietic development from human ES cells will translate to novel source of cells for hematopoietic cell transplants used to treat a variety of malignant and non-malignant blood cell diseases. These cells may also be used as a new source for transfusion medicine.