Generate Glucose-Responsive, Insulin-Producing Cells from Human Embryonic Stem Cells in 3D Cultures

本项目旨在建立三维人胚胎干细胞（hES）分化系统，并通过基因表达谱和谱系追踪实验来阐明人胰腺发生的机制。具体目标是：(1)建立3D胶原支架hES细胞培养系统，并证明3D培养的hES细胞产生葡萄糖反应，产生胰岛素的细胞；(2)表征3D培养的hES细胞向细胞的谱系特异性分化。初期的研究将集中在体外实验上，后续的研究将通过胰岛移植来表征糖尿病动物模型中胰腺细胞的体内功能。从这些研究中获得的知识将用于设计3D培养系统，以指导hES细胞分化为胰岛移植所需的葡萄糖反应细胞和胰岛素产生细胞。这是实现PI长期职业目标的下一步，即使用3D培养来了解3D环境中he细胞的生物学行为，这反过来将允许设计各种3D培养系统，以指导谱系特异性he细胞分化为各种治疗细胞，用于细胞基础治疗。这项工作的发现将为胚胎干细胞生物学提供新的见解，特别是在3D微环境中调控胚胎干细胞分化、增殖、粘附、迁移等机制。通过该项目开发的技术将为胰岛移植提供可再生细胞来源，可以恢复1型或晚期2型糖尿病患者接近生理的胰岛素分泌能力。该项目还将为干细胞工程和再生医学的本科生和研究生提供良好的教育和培训机会。

CBET-0756455YeThe objectives of this project are to develop a 3D human embryonic stem (hES) cell differentiation system and to elucidate the mechanisms of human pancreatogenesis through gene expression profiling and lineage tracing experiments. The specific aims are: (1) Establish a 3D collagen scaffolding hES cell culture system and demonstrate the generation of glucose-responsive, insulin-producing cells from hES cells in 3D cultures, and (2) Characterize the lineage-specific differentiation of hES cells into cells in 3D cultures. Initial studies will focus on in vitro experiments and subsequent studies will be designed to characterize in vivo functions of pancreatic cells in diabetic animal models through islet transplantation. Knowledge gained from these studies will be utilized to design a 3D culture system for directing the differentiation of hES cell into glucose-responsive and insulin-producing cells for islet transplantation. This is the next step to achieving the PI's long-term career goal of using 3D cultures to gain an understanding of biological behaviors of hES cells in 3D environments, which, in turn, will allow the designing of various 3D culture systems to direct lineage-specific hES cell differentiation into a variety of therapeutic cells for cell-based therapy. Findings from this work will offer new insights into hES cell biology, especially mechanisms that govern ES cell differentiation, proliferation, adhesion, migration, etc. in a 3D microenvironment. Technologies developed through this project will offer renewable cell sources for islet transplantation that can restore near-physiological insulin secretion capacity in type-1 or later stage type-2 diabetic patients. This project will also provide excellent opportunities for education and training of undergraduate and graduate students in stem cell engineering and regenerative medicine.