Control Arterial-Venous Differentiation Using 3-Dimensional Cell Printing Technol

描述（申请人提供）：干细胞在组织工程和再生医学中具有巨大的潜力。然而，为了利用干细胞产生用于植入的组织，我们必须克服一个主要挑战，即在组织生长时提供功能性血管网络和足够的灌注。为了实现从干细胞中产生功能性血管系统的长期目标，我们的第一步是研究如何控制干细胞分化为适当的血管谱系。在正常血管发育过程中，胚胎干细胞进入血管谱系并进一步进入动脉和静脉细胞的命运对于确定血管网络的模式和促进心脏开始跳动时的灌注至关重要。血流是维持血管身份和刺激血管成熟进入功能状态的另一重要驱动力。因此，为了从干细胞中设计出功能性的血管系统，总结血管发育过程中的这些关键步骤是很重要的。在这一提议中，我们计划研究干细胞同时分化为动脉和静脉内皮细胞并进一步维持其身份对于建立功能性血管网络至关重要的假设。本研究的目的是建立一个模型系统，使我们能够检验这一假设，并确定控制动脉和静脉细胞命运空间格局的关键因素。在具体目标1中，我们将开发一种三维细胞打印技术，能够按需控制干细胞壁龛。在Specific Aim 2中，我们将使用该技术设计一个体外环境，该环境概括了许多对正常血管发育重要的信号线索，并研究它们对胚胎干细胞分化为动脉和静脉规范的影响。基于这些研究，我们期望确定控制动脉和静脉细胞命运差异的关键因素。在Specific Aim 3中，我们将整合这些信号来构建一个三维动静脉环，以支持胚胎干细胞衍生的灌注微血管网络的生长。我们的研究通过关注如何使用一种新的细胞打印技术将胚胎干细胞分化成合适的血管谱系，向工程功能血管系统的目标迈出了第一步。此外，我们的研究将建立人类血管发育的体外模型，该模型可以揭示早期发育事件的一些详细信息，并可能为各种形式的血管异常的机制提供新的见解。

DESCRIPTION (provided by applicant): Stem cells have enormous potential in tissue engineering and regenerative medicine. However, in order to use stem cells to generate tissue for implantation, we must overcome a major challenge, which is to provide a functional vascular network and adequate perfusion while the tissue is growing. To achieve this long term goal of generating functional vasculature from stem cells, our first step is studying how to control stem cell differentiation into the proper vascular lineages. During normal vascular development, embryonic stem cell commitment into vascular lineage and further into the arterial and venous cell fate is crucial to define the patterns of vascular network and facilitate the perfusion when the heart starts to beat. Blood flow is another important driving force to maintain vascular identity and stimulate vascular maturation into a functional state. Therefore, to engineer a functional vasculature from stem cells, it is important to recapitulate these critical steps in vascular development process. In this proposal, we plan to investigate the hypothesis that simultaneously differentiating stem cells into arterial and venous endothelial cells and further maintaining their identity are critical for building a functional vascular network. The goal of this study is to build a model system that allows us to test this hypothesis and to define critical factors for controlling the spatial patterns of arterial and venous cell fate. In Specific Aim 1, we will develop a 3-Dimensional cell printing technology capable of providing on-demand control of stem cell niches. In Specific Aim 2, we will use this technology to engineer an in vitro environment that recapitulates many signal cues important for normal vascular development, and study their effects on embryonic stem cell differentiation into the arterial and venous specification. Based on these studies, we expect to define critical factors that differentially control arterial and venous cell fate. In Specific Aim 3, we will incorporate these signals to build a 3-Dimensional Arterial-Venous loop to support the growth of perfused microvascular network derived from embryonic stem cells. Our research undertakes a first step toward the goal of engineering a functional vasculature by focusing on how to differentiate embryonic stem cells into the proper vascular lineage using a novel cell printing technology. In addition, our study will create an in vitro model of human vascular development which can reveal some detailed information of early developmental events and may provide novel insights into the mechanisms of various forms of vascular anomalies. PUBLIC HEALTH RELEVANCE: This project will use a novel cell printing technology to differentiate embryonic stem cells into arterial and venous cells, and further assemble them into 3-D vascular network with perfusion, which is a critical step in engineering human tissues for therapeutic purpose. In addition, this research will also reveal information related to early vascular development process and may provide insight into the mechanisms of arterial-venous malformations.