Developmental Paradigms to Direct Human Endoderm into Foregut Lineage

描述（由申请人提供）：本提案的目的是了解如何有效地将人胚胎干细胞（HESC）分化为不同的内胚层谱系，如肺、肝、胰腺和肠，用于治疗用途，以替代患病或受损的组织。我假设，目前的ES细胞分化方案可以大大改善，通过纳入一个关键的前后（A-P）图案的事件，通常发生在早期内胚层器官发育。此外，我还假设，操纵信号 负责A-P模式化的内胚层细胞将使我们能够系统地产生各种不同的内胚层谱系，包括肺、甲状腺和肠。我们和其他实验室发表的研究表明，在肠管形成之前控制FGF和Wnt信号传导水平对于沿着前后（A-P）轴形成内胚层和建立器官结构域至关重要。FGF和Wnt信号通路的内胚层图案化活性在HESC中是保守的。根据我们的初步数据，我假设FGF和Wnt通路协同作用以两种方式促进人内胚层细胞命运，第一种是通过调节关键A-P决定簇的表达，第二种是通过引导细胞沿沿着A-P轴迁移。此外，我推测，通过操纵FGF和Wnt信号传导水平，我可以有效地将定形内胚层（DE）导入特定的前肠谱系，包括胰腺和肝脏。拟议的实验将首次允许询问FGF和Wnt途径如何指导人类前肠特化的早期阶段，并更有效地将HESC引导到治疗上重要的组织中。此外，细胞培养系统的优势将使我能够测试FGF和Wnt信号之间的机制相互作用，这在动物模型中很难解决。具体目标1：确定FGF和Wnt信号传导如何协同作用以形成人内胚层和指导前肠谱系特化。具体目的2：研究HESC衍生的前肠内胚层在体内的分化潜力。具体目标3：确定FGF 4如何协调人内胚层迁移。公共卫生相关性：该提案的目标是了解如何有效地指导人胚胎干细胞（HESC）分化为不同的内胚层谱系，如肺、肝、胰腺和肠，用于治疗用途，以替代患病或受损的组织。

DESCRIPTION (provided by applicant): The goal of this proposal is to understand how to efficiently direct differentiation of human embryonic stem cells (HESCs) into different endoderm lineages, such as lung, liver, pancreas and intestine for therapeutic use to replace diseased or damaged tissue. I hypothesize that current ES cell differentiation protocols can be greatly improved by incorporating a critical anterior-posterior (A-P) patterning event that normally occurs during early endoderm organ development. In addition, I also hypothesize that manipulating signals responsible for A-P patterning will allow us to systematically generate the full range of different endoderm lineages including lung, thyroid and intestine. Published studies from our and other labs have demonstrated that controlling the levels of FGF and Wnt signaling prior to formation of a gut tube is critical for patterning the endoderm along the anterior-posterior (A-P) axis and for establishing the organ domains. The endoderm patterning activity of FGF and Wnt signaling pathways is conserved in HESCs. From our preliminary data I hypothesize that the FGF and Wnt pathways synergize to promote human endoderm cell fate in two ways, first by regulating expression of key A-P determinants and second by directing cell migration along the A-P axis. Furthermore I hypothesize that by manipulating the levels of FGF and Wnt signaling I can efficiently direct definitive endoderm (DE) into specific foregut lineages, including pancreas and liver. The proposed experiments will allow, for the first time, interrogation of how FGF and Wnt pathways direct the early stages of foregut specification in humans and to more efficiently direct HESCs into therapeutically important tissues. Moreover, the advantages of a cell culture system will allow me to test mechanistic interactions between FGF and Wnt signaling which are difficult to address in animal models. Specific Aim 1: Determine how FGF and Wnt signaling synergize to pattern human endoderm and direct foregut lineage specification. Specific Aim 2: Investigate the differentiation potential of HESC-derived foregut endoderm in vivo. Specific Aim 3: Determine how FGF4 coordinates human endoderm migration. PUBLIC HEALTH RELEVANCE: The goal of this proposal is to understand how to efficiently direct differentiation of human embryonic stem cells (HESCs) into different endoderm lineages, such as lung, liver, pancreas and intestine for therapeutic use to replace diseased or damaged tissue.