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.

不同种类的细胞组成大脑和脊髓。复制这些复杂的结构为我们提供了一个重要的机会来发现这些组织是如何在体内形成的。这种工程神经组织可代替捐赠的人体组织用于药理学筛选。组织工程的一种流行策略使用生物材料支架来传递促进干细胞分化为神经组织的信号。我的团队研究人类诱导多能干细胞（hiPSC），这些成体细胞被重新编程为可以成为生物体中发现的任何类型细胞的状态。这一特性使它们成为组织工程的极佳细胞来源。我的小组确定了许多推动 hiPSC 分化为神经组织的化学和物理线索，这是该研究计划的起点。然而，目前使用 hiPSC 改造神经组织的方法需要冗长、劳动密集型的方案。该研究计划的总体目标是通过开发生物活性支架来改造 hiPSC 的功能性神经组织，该支架提供必要的化学和物理线索，以促进快速分化，并可转化为生物打印应用。该研究计划包含实现这一目标的两个不同目标，与现有方法相比，该研究计划的结果将用于以快速、高通量的方式生成神经组织。 ******第一个目标是研究如何修改 3D 纤维蛋白支架的特性，以便它们能够从 hiPSC 生成功能性神经组织。我们可以使用交联剂来控制这些支架的机械性能，以提高其稳定性。我们还可以通过使用促进 hiPSC 神经元分化的肽等生物活性线索对其进行功能化，从而增强其化学特性。然后，我们将使用这些支架作为功能性神经组织 3D 打印的生物墨水，确定这两种类型的线索对 hiPSC 分化的影响。 ******第二个目标将阐明微球中新型化学信号的受控释放如何能够快速将 hiPSC 分化为神经元。这些化学信号包括通过细胞内蛋白质递送技术功能化的转录因子 Ascl1（被证明可以有效产生神经元）、嘌吗啡胺（被证明可以增强运动神经元分化）和古古甾酮（被证明可以增强多巴胺能神经元分化）。目前，没有现有的药物递送系统可以控制这些分子的释放，所有这些分子都促进 hiPSC 快速分化为神经组织。我们将演示如何使用这些药物释放微球的不同组合和浓度来设计两种不同类型的神经组织，然后将其用于药物筛选应用。