Microfluidic Platform for Genetic Guidance of Stem Cell Differentiation

胚胎和成体干细胞是再生医学的基础。他们独特的能力 进行自我更新，平衡产生多种分化细胞类型的能力，有望 彻底改变退行性疾病的治疗方法。它们广泛翻译使用的一个核心挑战是 识别环境线索和基因调控网络，管理自我更新和 将干细胞定向分化为感兴趣的组织。确定潜在的调控基因 起作用的电路是引导干细胞分化的关键一步。此外，调节和调节的能力 监测干细胞以测试预测的基因调控网络关系是评估的必要步骤 合理的、可重复的、有效的分化遗传指导方法。 在此，我们建议将我们现有的微技术平台应用于多路复用器的动态成像 理解和评估干细胞中基因调控网络作用的微环境 差异化。已知的基因激活剂的引入与细胞谱系记者的结合提供了一种 独特的读数，潜在地使用遗传方法来指导干细胞分化。通过病毒 外源遗传因子的表达和荧光记者的监测我们将评估基因 小鼠胚胎干细胞分化为胰腺内胚层的调控网络， 牙齿和心脏瓣膜组织作为SysCODE财团的一部分。此外，微技术的使用 工具提供了可暴露于各种独立扰动的微环境，并且 以高时间和空间分辨率监测响应，允许评估许多基因 沿着特定的细胞分化途径进行的输入。 我们技术平台的中等吞吐量特性与潜力的产生相结合 通过SysCODE联盟的基因调控网络为评估提供了一个强大的试验台 网络预测和开发组织体外方法和治疗的最终潜在靶点 再生和器官发生。

Stem cells, embryonic and adult, represent the foundation for regenerative medicine. Their unique ability to undergo self-renewal balanced with the capacity to generate numerous differentiated cell types promises to revolutionize therapy of degenerative disorders. A central challenge to their widespread translational use is the identification of environmental cues and genetic regulatory networks that govern the self-renewal and directed differentiation of stem cells into a tissue of interest. Identification of the underlying gene regulatory circuitry at work is a key step for directing stem cell differentiation. Moreover, the ability to modulate and monitor stem cells to test predicted gene regulatory network relationships is a necessary step in evaluating plausible, reproducible and efficient methods for the genetic guidance of differentiation. Here we propose to adapt our existing microtechnology platform for dynamic imaging of multiplexed microenvironments to understanding and evaluating the role of gene regulatory networks in stem cell differentiation. The introduction of known genetic activators combined with cell lineage reporters provides a unique readout for the potential use of genetic methods to guide stem cell differentiation. Through the viral expression of exogenous genetic factors and monitoring of fluorescence reporters we will evaluate gene regulatory networks underlying the differentiation of murine embryonic stem cells into pancreatic endoderm, tooth and heart valve tissue as part of the SysCODE consortium. Furthermore, the use of microtechnology tools provide microenvironments that can be exposed to a variety of independent perturbations and the response monitored with high temporal and spatial resolution, permitting the evaluation of many genetic inputs along a specific cellular differentiation pathway. The moderate throughput nature of our technology platform in combination with the generation of potential gene regulatory networks through the SysCODE consortium represent a powerful testbed for the evaluation of network predictions and ultimately potential targets for developing in vitro methods and therapies for tissue regeneration and organogenesis.