Microfluidic systems for 3D cell culture and micro tissue constructs

这项研究计划专注于为组织工程和药物筛选应用创建三维(3D)微尺度组织模型的方法的开发。我们将使用微流控系统来生成包含支持嵌入细胞的细胞外基质(ECM)元素的组织模型。我们还将研究细胞悬浮液的喷墨打印的流体力学，以了解在组织构图过程中导致细胞分配不可靠的现象和参数。我们将使用粒子图像测速仪和高速成像来表征打印过程中喷墨喷嘴内部的流场。 另一个主要的研究目标将是开发微流控平台，用于在细胞培养过程中在空间和时间上控制微环境因素，如氧分压。我们将开发有选择地在我们的微流体设备的特定区域涂上保形涂层的方法，以使聚二甲基硅氧烷(PDMS)在保持透氧性的同时，不渗透培养介质中的营养物质或信号分子。由此产生的系统将被设计为提供缺氧和低氧培养条件，通过减少氧气控制通道和细胞培养室之间的扩散距离，我们将创造循环低氧条件，循环时间在几分钟内。为了监测组织结构内的氧气水平，我们将开发生物相容的微米大小的颗粒，负载氧气敏感染料。这些颗粒将悬浮并分散在细胞水凝胶珠中，允许光学、非侵入性、比率法原位测量组织结构内部的氧气水平。最后，我们还将开发方法来获得这些组织的高质量图像。通过几百微米厚的组织切片进行成像将需要减少光散射的技术。通过在微流控平台上集成细胞培养环境，我们将能够监控细胞周期进展和细胞外基质重塑。 通过这个研究计划，我们将解决3D细胞培养、模型组织的生成和组织表征所面临的一些最紧迫的挑战。从长远来看，这项工作的总体结果包括为基础科学和工业工具开发微型组织模型。这些结果的应用包括研究低氧对细胞周期和药物敏感性的影响，用于高通量、高含量的药物毒性测试和药物筛选。这里开发的方法也将适用于广泛的组织共培养模型和日益复杂的组织。

This research program focuses on the development of methods to create three dimensional (3D) microscale tissue models for tissue engineering and drug screening applications. We will use microfluidic systems to generate tissue models comprising elements of the extracellular matrix (ECM) which will support the embedded cells. We will also investigate hydrodynamics of inkjet printing of cell suspensions in order to understand the phenomena and parameters which contribute to unreliable cell dispensing during tissue patterning. We will use particle image velocimetry and high-speed imaging to characterize the flow field inside inkjet nozzles during the printing process. Another major research goal will be to develop microfluidic platforms for controlling microenvironmental factors such as oxygen tension, both spatially and temporally, during cell culture. We will develop methods to selectively coat specific regions of our microfluidic devices with conformal coatings in order to render the polydimethylsiloxane (PDMS) impermeable to absorption of nutrients or signaling molecules from the culture media, while maintaining oxygen permeability. The resulting systems will be designed to provide anoxic as well as hypoxic culture conditions, and by reducing diffusion distances between oxygen control channels and the cell culture chambers, we will create cyclic hypoxic conditions with cycle times on the order minutes. In order to monitor O2 levels within the tissue constructs, we will develop biocompatible, micron-sized particles loaded with oxygen-sensitive dyes. These particles will be suspended and dispersed within the cell-laden hydrogel beads, permitting optical, non-invasive, ratiometric measurement in situ of oxygen levels on the interior of the tissue constructs. Finally, we will also develop methods to obtain high quality images of these tissues. Imaging through tissue sections with several hundred micron thickness will require techniques to reduce light scattering. By integrating the cell culture environment on a microfluidic platform, we will be able to monitor cell cycle progression together with ECM remodeling. Through this research program we will address some of the most pressing challenges facing 3D cell culture, generation of model tissues, and tissue characterization. In the long term, the overall results from this work include development of microscale tissue models for basic science as well as tools for industry. Applications of these results include studying the effect of hypoxia on cell cycle and drug sensitivity, for high-throughput, high-content drug toxicity testing and drug screening. The methods developed here will also be applicable to a wide range of tissue co-culture models and tissues of increasing complexity.