Mesodermal Commitment of the Definitive Hematopoietic Program

项目摘要/摘要 我们研究的总体目标是了解胚胎的起源、发育和调控。 造血干细胞(HSC)、血源性内皮细胞(HE)的祖细胞。这项提议的重点是 以人多能干细胞和小鼠胚胎发育为互补模型 用于识别HE规范的基本信号和基因的系统。目前，信号要求(S) 胚胎发育过程中新生中胚层的规范仍然是一个鲜为人知的过程。 我们建议的研究建立在我们在定向分化明确的造血细胞方面的最新进展的基础上 来自hPSCs的祖细胞，已经鉴定出依赖WNT的中胚层群体 明确的造血潜能，通过CDX4的表达。我们现在已经确定了早期的一个新子集 中胚层完全依赖于维甲酸(RA)信号来产生HOXA-HE，具有多个 血统具有明确的造血潜能。由于RA在胚胎发育过程中是HE功能所必需的，我们 假设hPSC派生的RA依赖的HE来自一个非常独特的中胚层亚集，这是 在个体发育上不同于具有HSC非依赖性HE潜能的中胚层。我们将检验这一假设 跨越3个具体目标。在目标1中，我们将评估ALDH1A2来源的HE的生理学相关性，在 体外和体内。这些研究的目的是了解ALDH1A2中胚层是否会导致 有HSC能力的他。在目标2中，我们将定义RA依赖的HE规范的机制。目标是 这些研究的目的是确定RA信号如何影响HE的发育。在目标3中，我们将确定 RA依赖的明确造血规范中的选择性剪接。这些研究的目的是 明确早期中胚层中与HE发生发展相关的重要转录调控因子。这个 成功完成这些研究将使我们更全面地了解 造血发育。这对基础生物学和由以下方面产生的见解具有根本重要性 这些研究将具有临床意义，例如体外培养一系列的造血干细胞。 再生医学的应用。我们独特的细胞和分子工具，结合我们在 造血发育、干细胞生物学和生物信息学使我们处于一个理想的位置，可以做出显著的 在这一领域的影响。

PROJECT SUMMARY / ABSTRACT The overall goal of our research is to understand the origin, development and regulation of the embryonic progenitor to the hematopoietic stem cell (HSC), hemogenic endothelium (HE). The focus of this proposal is to use human pluripotent stem cells (hPSCs) and mouse embryonic development as complementary model systems to identify the essential signals and genes for HE specification. Currently, the signal requirement(s) for HE specification from nascent mesoderm during embryonic development remains a poorly understood process. Our proposed studies build on our recent advances in the directed differentiation of definitive hematopoietic progenitors from hPSCs, having identified a WNT-dependent mesodermal population that harbors exclusively definitive hematopoietic potential, via CDX4 expression. We have now identified a novel subset of early mesoderm that is entirely dependent on retinoic acid (RA) signaling for the generation of HOXA+ HE with multi- lineage definitive hematopoietic potential. As RA is required for HE function during embryogenesis, we hypothesize that hPSC-derived RA-dependent HE is derived from a very unique mesodermal subset, which is ontogenically distinct from mesoderm harboring HSC-independent HE potential. We will test this hypothesis across 3 Specific Aims. In Aim 1, we will assess the physiological relevance of ALDH1A2+ derived HE, both in vitro and in vivo. The objective of these studies is to understand whether ALDH1A2+ mesoderm gives rise to HSC-competent HE. In Aim 2, we will define the mechanism the specification of RA-dependent HE. The objective of these studies is to determine how RA signaling affects HE development. In Aim 3, we will identify the role of alternative splicing during RA-dependent definitive hematopoietic specification. The objective of these studies is to define the essential transcriptional regulators within early mesoderm for the development of HE. The successful completion of these studies will provide us with a more comprehensive understanding of hematopoietic development. This is of fundamental importance to basic biology, and the insights generated from these studies will have clinical implications, such as the in vitro generation of HSCs for a wide array of regenerative medicine applications. Our unique cellular and molecular tools, combined with our expertise in hematopoietic development, stem cell biology and bioinformatics puts us in an ideal position to make a significant impact in this field.