Differentiating Embryonic Stem Cells Toward Arterial and Venous Endothelial Cells

描述(申请人提供)：胚胎干细胞(ESCs)来源的内皮细胞(ECs)具有巨大的潜力用于各种治疗领域，如血管移植物的组织工程和缺血组织的再血管化。为了特定的应用，还迫切需要获得具有功能的动脉或静脉内皮细胞的均质培养。到目前为止，已经开发了各种方案来区分胚胎干细胞和血管内皮细胞。然而，使用目前的方法从ESCs获得的ECs主要表现为静脉表型。因此，发展精细的动静脉分化方法是解决这一差距的关键。根据血管发育的结果，我们推测胚胎干细胞来源的Flk1 Nrp1细胞可能是动脉内皮细胞的前体细胞。我们认为这一亚群细胞群易于动脉分化，可以结合环境线索选择来指导动脉分化。我们的初步数据支持这一假设。这项研究的目标是用人类胚胎干细胞进一步检验这一假说。然后，我们将设计最佳的体外环境，引导胚胎干细胞进入动脉和静脉细胞的命运，并比较它们在组织工程应用中的功能结果。具体地说，我们将：(1)验证Nrp1可用于从干细胞中识别动脉内皮细胞前体细胞，并确定引导ESCs走向动脉和静脉细胞命运的最佳体外环境。(2)分析干细胞来源的动、静脉内皮细胞在体外和体内构建组织工程血管网络的能力。(3)确定干细胞来源的动、静脉内皮细胞在体内和体外组织工程血管移植物重建中的作用。

DESCRIPTION (provided by applicant): Embryonic stem cells (ESCs) derived endothelial cells (ECs) have enormous potential to be used in a variety of therapeutic areas such as tissue engineering of vascular grafts and re-vascularization of ischemic tissues. It is also much desired to obtain homogeneous culture of functional arterial or venous ECs for specific applications. To date, various protocols have been developed to differentiate ESCs toward vascular ECs. However, ECs derived from ESCs using current methods display predominantly venous phenotype. Therefore, developing refined method of arterial-venous differentiation is critically needed to address this gap. Based on the findings of vascular development, we hypothesize that embryonic stem cell derived Flk1+Nrp1+ cells serve as arterial EC progenitors. We think that this subset cell population is predisposed to arterial differentiation and can be selected to guide arterial differentiation in combination with environmental cues. Our preliminary data support this hypothesis. The goal of this study is to further test this hypothesis using human ESCs. We will then engineer optimal in vitro environments that guide ESCs into arterial and venous cell fate and compare their functional consequences in tissue engineering applications. Specifically, we will: (1) Validate that Nrp1 can be used to identify arterial EC progenitor from stem cells and define optimal in vitro environments that guide ESCs into arterial and venous cell fate. (2) Analyze the ability of ESC-derived arterial and venous ECs to form interconnected functional vascular network in tissue-engineered construct both in vitro and in vivo. (3) Determine the functional consequences of ESC-derived arterial and venous ECs in the remodeling of tissue engineered vascular graft both in vitro and in vivo.