Microsphere-mediated differentiation of embryonic stem cells

描述(申请人提供)：针对各种退行性疾病和创伤的干细胞疗法可能能够恢复细胞丢失和刺激组织再生。多能胚胎干细胞(ESCs)能够分化为功能神经元、心肌细胞和胰岛β细胞，是一系列不同再生细胞疗法发展的强大细胞来源。尽管多能干细胞具有明显的潜力，但一个关键的限制是无法以类似于胚胎发育的方式在空间和时间上控制形态因子向ESCs的分子输送，从而有效地指导细胞向特定表型的分化。由不同生物材料制成的微球可以被设计成局部控制生物活性分子的释放，从而同时提供对生物分子向细胞和组织输送的空间、时间和剂量依赖的控制。将工程微球集成到干细胞微环境中，在干细胞分化过程中控制、局部地向干细胞输送形态原，是发展定向干细胞再生疗法的一种新的、可翻译的方法。这项建议的目的是：1)研究将不同类型的微球掺入正在分化的被称为类胚体(EBS)的ESCs聚集体中的影响；2)确定在EB微环境中独立传递形态因子(如Wnt3a和BMP2)的时空效应；以及3)研究从EBS中结合的微球中连续传递不同形态因子(Wnt3a和BMP2)的影响。这些研究的完成将产生新的信息，即通过基于生物材料的递送方法，在空间和时间上控制细胞外微环境调控ESC分化的分子组成的能力。此外，这种方法代表了一种通过在局部控制分子因子的呈现来指导干细胞体外分化的根本新途径，这可能是干细胞技术发展中广泛适用的原则。公共卫生相关性：寻求将存活细胞恢复到患病或受伤组织中的干细胞疗法可以促进功能性组织再生，但目前受到无法有效控制干细胞分化的限制。这一建议旨在通过在空间和时间上控制来自3D干细胞微环境中的微球中调节干细胞分化的形态因子的呈现来提高多潜能干细胞分化的效率和同质性。

DESCRIPTION (provided by applicant): Stem cell therapies for various degenerative diseases and traumatic injuries may be capable of restoring cell loss and stimulating tissue regeneration. Pluripotent embryonic stem cells (ESCs) are capable of differentiating into functional neurons, cardiomyocytes and pancreatic beta cells, thus representing a robust cell source for the development of an array of different regenerative cellular therapies. Despite the clear potential of pluripotent stem cells, a critical limitation is the inability to spatially and temporally control the molecular delivery of morphogenic factors to ESCs in a manner similar to embryonic development that efficiently directs the differentiation of the cells to specific phenotypes. Microspheres made of different biomaterials can be engineered to locally control the release of bioactive molecules, thereby simultaneously providing spatial, temporal and dose-dependent control of biomolecular delivery to cells and tissues. The integration of engineered microspheres into stem cell microenvironments for controlled, local delivery of morphogens to stem cells during the course of differentiation is a novel and translatable approach for the development of directed stem cell regenerative therapies. The objectives of this proposal are to 1) examine the effects of incorporating different types of microspheres within aggregates of ESCs undergoing differentiation referred to as embryoid bodies (EBs), 2) determine the spatiotemporal effects of independently delivering morphogens, such as Wnt 3a and BMP2, within EB microenvironments, and 3) examine the effects of sequential delivery of different morphogens (Wnt3a and BMP2) from microspheres incorporated within EBs. The completion of these studies will yield novel information about the ability to spatially and temporally control the molecular composition of the extracellular microenvironment regulating ESC differentiation via biomaterials-based delivery methods. In addition, this approach represents a fundamentally new route to directing the differentiation of stem cells in vitro through the controlled presentation of molecular factors locally, which may be a broadly applicable principle in the development of stem cell technologies. PUBLIC HEALTH RELEVANCE: Stem cell-based therapies seeking to restore viable cells to diseased or wounded tissues could promote functional tissue regeneration, but are currently limited by an inability to efficiently control the differentiation of the stem cells. This proposal seeks to improve the efficiency and homogeneity of pluripotent stem cell differentiation by spatially and temporally controlling the presentation of morphogenic factors regulating stem cell differentiation from microspheres incorporated within 3D stem cell microenvironments.