Microfluidic Platform for Stem Cell Applications

 描述（由申请人提供）：干细胞疗法的新兴领域有可能永远改变医学。然而，将干细胞疗法带给患者的一个主要瓶颈是缺乏足够的体外测定来研究干细胞质量。关键测试标准是分化过程之前的功效（多能性）和分化后植入干细胞衍生组织之前的安全性（缺乏致瘤性）。可用于评估干细胞质量的最简单的测定是胚状体（EB）测定。然而，该测定不能支持组织生长足够长的时间以实现畸胎瘤的完全发育。因此，目前测试干细胞质量的黄金标准依赖于体内测试：将干细胞制剂注射到免疫缺陷小鼠体内。这种所谓的“畸胎瘤测定”评估干细胞的多能性，即发育成源自所有三个胚胎胚层的细胞类型的能力。不幸的是，这种体内测定具有显着的缺点：它需要大量动物，成本过高，耗时，劳动密集型，并且结果取决于手术技巧。该项目的目标是开发一种基于微流控芯片的体外测定方法，该芯片包含组织工程、血管化、人性化的微环境，用于测试干细胞多能性。使用 Nortis 技术获得的初步数据表明，所提出的体外模型可以比体内畸胎瘤测定更经济地进行多能性测试，并且在更短的时间内进行。此外，我们的数据表明，融入干细胞环境的灌注微脉管系统是人类畸胎瘤组织长期生存和分化的关键。在第一阶段，我们将开发微流体硬件和组织工程方案（具体目标 1）。此外，我们计划证明该测定的可行性，其质量和稳健性符合评估干细胞多能性的要求（具体目标 2）。我们将直接将我们的畸胎瘤芯片的性能与当前可用的方法、EB 测定和体内畸胎瘤测定进行比较。第一阶段的最低可行性要求是以显着降低的成本满足体内测定的性能质量和运行时间。在第二阶段，我们将致力于主要研发来验证检测方法并提高通量能力。最终，拟议的产品将为学术界和工业界的研究人员提供强大的新型体外工具，推动突破性干细胞疗法及其临床转化的发展。

 DESCRIPTION (provided by applicant): The emerging field of stem-cell therapy has the potential to transform medicine forever. However, a major bottleneck for bringing stem-cell therapies to the patient is the lack of adequate in-vitro assays for the study of stem-cell quality Critical test criteria are efficacy (pluripotency) prior to the differentiation process and safety lack of tumorigenicity) after differentiation prior to implantation of stem-cell derived tissues. The simplest assay available to assess stem-cell quality is the embryoid body (EB) assay. However, this assay is not able to support tissue growth long enough to achieve complete teratoma development. Therefore, the present gold standard for testing stem-cell quality relies on in vivo testing: by injecting stem-cell preparations into immunodeficient mice. This so-called "teratoma assay" assesses the stem cells' pluripotency, the ability to develop into cell types derived from all three embryonic germ layers. Unfortunately, this in-vivo assay has significant drawbacks: it requires a large number of animals, is prohibitively expensive, time consuming, labor- intensive, and results are dependent on surgical skills. The proposed project's objective is to develop an in-vitro assay based on a microfluidic chip containing a tissue-engineered, vascularized, humanized microenvironment for testing stem-cell pluripotency. Preliminary data obtained with the Nortis technology suggest that the proposed in-vitro model can perform the pluripotency test much more economically, and in a much shorter time frame than the in-vivo teratoma assay. Additionally, our data indicate that perfused microvasculature incorporated into stem-cell environment is key to long-term viability and differentiation of human teratoma tissue. During Phase I we will develop the microfluidic hardware and tissue-engineering protocols (Specific Aim 1). Additionally, we plan to demonstrate feasibility that the assay can be performed with a quality and robustness that complies with the requirements for assessing stem-cell pluripotency (Specific Aim 2). We will compare the performance of our teratoma chip directly with currently available methods, the EB assay and in-vivo teratoma assay. Minimum feasibility requirements for Phase I are to meet the performance quality and run time of the in- vivo assay, at significantly reduced costs. During Phase II we will dedicate major R&D to validate the assay and increase throughput capabilities. Ultimately, the proposed product will provide researchers in academia and industry with a powerful new in-vitro tool that will fuel the development of groundbreaking stem-cell therapies and their clinical translation.