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，为了利用干细胞生成移植组织，我们必须克服一个重大挑战，即在组织生长的同时提供一个功能良好的血管网络和足够的灌流。为了实现从干细胞生成功能性血管的长期目标，我们的第一步是研究如何控制干细胞分化为合适的血管谱系。在正常的血管发育过程中，胚胎干细胞承诺进入血管谱系，进而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流至关重要。血流是维持血管特性和刺激血管成熟进入功能状态的另一个重要驱动力。因此，要从干细胞设计出具有功能的血管系统，重要的是要概括这些血管发育过程中的关键步骤。在这项提议中，我们计划研究一种假设，即同时将干细胞分化为动脉和静脉内皮细胞，并进一步保持它们的特性对于建立一个有功能的血管网络至关重要。这项研究的目标是建立一个模型系统，使我们能够检验这一假设，并定义控制动脉和静脉细胞命运的空间模式的关键因素。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的位置。在具体目标2中，我们将使用这项技术来设计一个体外环境，该环境概括了许多对正常血管发育至关重要的信号信号，并研究它们对胚胎干细胞分化为动脉和静脉规格的影响。在这些研究的基础上，我们期望确定不同地控制动脉和静脉细胞命运的关键因素。在具体目标3中，我们将结合这些信号来构建一个三维的动-静脉环，以支持来自胚胎干细胞的灌流微血管网络的生长。我们的研究向设计功能性血管系统的目标迈出了第一步，重点是如何使用一种新的细胞打印技术将胚胎干细胞分化为合适的血管谱系。此外，我们的研究将建立一个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。 与公共卫生相关：该项目将使用一种新的细胞打印技术将胚胎干细胞分化为动脉和静脉细胞，并通过灌流将它们进一步组装成三维血管网络，这是将人体组织工程用于治疗的关键步骤。此外，这项研究还将揭示与早期血管发育过程相关的信息，并可能为动静脉畸形的机制提供洞察力。

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.