Engineering neural tissue using pluripotent stem cells

使用多能干细胞改造神经组织

• 批准号: RGPIN-2017-04044
• 负责人: Willerth, Stephanie
• 金额: $ 1.75万
• 依托单位: University of Victoria
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=679696
• 关键词: Engineering; neural; tissue; using; pluripotent

Different kind of cells make up the brain and spinal cord. Replicating these complex structures provides us with a significant opportunity to discover how such tissues form in the body. Such engineered neural tissues can be used for applications in pharmacological screening instead of donated human tissues. One popular strategy for tissue engineering uses biomaterial scaffolds to deliver signals that promote stem cells to differentiate into neural tissue. My group works with human induced pluripotent stem cells (hiPSCs), adult cells reprogrammed into a state where they can become any type of cell found in an organism. This property makes them an excellent cell source for tissue engineering. My group identified a number of chemical and physical cues that drive the differentiation of hiPSCs into neural tissue, which serves as a starting point for this research program. However, current methods for engineering neural tissue using hiPSCs require lengthy, labor intensive protocols. The overarching goal of this research program is to engineer functional neural tissues from hiPSCs by developing bioactive scaffolds that present the necessary chemical and physical cues for promoting rapid differentiation that can be translated into bioprinting applications. This research program consists of two different aims for achieving this goal, and the results of this research program will be used to generate neural tissue in a rapid and high throughput manner compared to current methods. ******The first aim investigates how to modify the properties of 3D fibrin scaffolds so that they can generate functional neural tissue from hiPSCs. We can manipulate the mechanical properties of these scaffolds using cross-linking agents to increase their stability. We also can enhance their chemical properties by functionalizing them with bioactive cues like peptides that promote neuronal differentiation of hiPSCs. We will then determine the influence of these two types of cues on hiPSC differentiation by using these scaffolds as bioink for 3D printing of functional neural tissues. ******The second aim will elucidate how the controlled release of novel chemical cues from microspheres can rapidly differentiate hiPSCs into neurons. These chemical cues include the transcription factor Ascl1 (shown to efficiently produce neurons) functionalized with intracellular protein delivery technology, purmorphamine (shown to enhance motor neuron differentiation), and guggulsterone (shown to enhance dopaminergic neuron differentiation). Currently, no existing drug delivery systems can generate controlled release of these molecules all of which promote rapid differentiation of hiPSCs into neural tissue. We will demonstrate how different combinations and concentrations of these drug-releasing microspheres can be used to engineer two different types of neural tissue, which could then be used for drug screening applications.

不同种类的细胞构成大脑和脊髓。复制这些复杂的结构为我们提供了一个重要的机会，可以发现这种组织如何形成体内。这种工程的神经组织可用于在药理学筛查中应用，而不是捐赠的人体组织。组织工程的一种流行策略使用生物材料支架传递信号，以促进干细胞分化为神经组织。我的小组与人类诱导的多能干细胞（HIPSC）一起工作，成年细胞重编程为可以成为生物体中发现的任何类型细胞的状态。该特性使它们成为组织工程的绝佳细胞来源。我小组确定了许多化学和物理线索，这些化学和物理线索将hipsc的分化为神经组织，这是该研究计划的起点。但是，使用HIPSC进行工程神经组织的当前方法需要冗长的劳动密集型方案。该研究计划的总体目标是通过开发生物活性脚手架来设计来自HIPSC的功能性神经组织，这些脚手架呈现出必要的化学和物理线索，以促进可以将快速分化的化学和物理提示转化为生物打印应用。该研究计划包括实现这一目标的两个不同的目的，与当前方法相比，该研究计划的结果将用于快速和高通量的方式生成神经组织。 ******第一个目的研究了如何修改3D纤维蛋白支架的特性，以便它们可以从HIPSC中产生功能性神经组织。我们可以使用交联剂来操纵这些支架的机械性能，以提高其稳定性。我们还可以通过用生物活性提示（如促进HIPSC神经元分化的肽）功能化来增强其化学特性。然后，我们将通过将这些支架用作生物界的3D打印功能性神经组织来确定这两种线索对HIPSC分化的影响。 *****第二个目标将阐明微球从微球中受控释放如何将HIPSC迅速区分为神经元。这些化学提示包括用细胞内蛋白质递送技术，呼吸肾上腺素（显示可增强运动神经元分化）和guggulsterone（显示可增强多巴胺能神经元差异）功能化的转录因子ASCL1（显示有效产生神经元）。目前，现有的药物输送系统可以产生这些分子的受控释放，所有这些分子均可以促进HIPSC快速分化为神经组织。我们将展示这些释放药物的微球的不同组合和浓度如何用于设计两种不同类型的神经组织，然后可以将其用于药物筛查。