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，为了使用干细胞来产生用于植入的组织，我们必须克服一个主要挑战，即在组织生长的同时提供功能性血管网络和足够的灌注。为了实现从干细胞生成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。在正常的血管发育过程中，胚胎干细胞定向进入血管谱系并进一步进入动脉和静脉细胞命运对于定义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，为了从干细胞工程化功能性脉管系统，重要的是概括脉管发育过程中的这些关键步骤。在这项提案中，我们计划调查的假设，同时分化成动脉和静脉内皮细胞干细胞，并进一步保持他们的身份是建立一个功能性血管网络的关键。本研究的目标是建立一个模型系统，使我们能够测试这一假设，并确定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，能够提供对干细胞龛的按需控制。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，重现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格的影响。基于这些研究，我们期望确定差异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三维动静脉环，以支持来自胚胎干细胞的灌注微血管网络的生长。我们的研究通过关注如何使用新型细胞打印技术将胚胎干细胞分化为适当的血管谱系，朝着工程化功能性血管系统的目标迈出了第一步。此外，我们的研究将建立一个体外模型的人类血管发育，可以揭示一些详细的信息，早期发育事件，并可能提供新的见解，各种形式的血管异常的机制。 公共卫生相关性：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，并进一步将其组装成具有灌注的三维血管网络，这是工程化人体组织用于治疗目的的关键步骤。此外，这项研究还将揭示与早期血管发育过程相关的信息，并可能为深入了解动静脉畸形的机制提供帮助。

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.