Microfluidic Platform for Stem Cell Applications

 描述(由申请者提供)：干细胞治疗的新兴领域有可能永远改变医学。然而，将干细胞疗法引入患者的一个主要瓶颈是缺乏足够的体外分析来研究干细胞质量关键测试标准是分化过程之前的有效性(多能性)和安全性(在干细胞来源的组织植入之前分化后缺乏致瘤性)。评估干细胞质量的最简单的方法是类胚体(EB)试验。然而，这种试验不能支持组织生长足够长的时间来实现完全畸胎瘤的发展。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中。这种所谓的“畸胎瘤分析”评估了干细胞的多能性，即从所有三个胚胎生殖层发育成细胞类型的能力。不幸的是，这种体内试验有很大的缺点：它需要大量的动物，昂贵得令人望而却步，耗时长，劳动密集型，结果依赖于手术技能。这项拟议的项目的目标是开发一种基于微流控芯片的体外测试，该芯片包含一个组织工程化、血管化、人性化的微环境，用于测试干细胞的多能性。用Nortis技术获得的初步数据表明，所提出的体外模型可以更经济地进行多能性测试，并且比体内畸胎瘤测试的时间要短得多。此外，我们的数据表明，融合到干细胞环境中的灌流微血管是人类畸胎瘤组织长期存活和分化的关键。在第一阶段，我们将开发微流控硬件和组织工程方案(具体目标1)。此外，我们计划证明该检测的可行性，该检测的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目前可用的方法-EB检测和体内畸胎瘤检测-进行比较。第一阶段的最低可行性要求是以显著降低的成本满足体内试验的性能质量和运行时间。在第二阶段，我们将致力于主要的研发，以验证该分析并提高生产能力。最终，拟议中的产品将为学术界和工业界的研究人员提供一种强大的体外新工具，将推动突破性干细胞疗法的发展及其临床翻译。

 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.