Microfluidic systems for 3D cell culture and micro tissue constructs

该研究计划的重点是开发用于组织工程和药物筛选应用的三维（3D）微尺度组织模型的方法。我们将使用微流体系统来生成组织模型，该组织模型包括将支持嵌入的细胞的细胞外基质（ECM）的元素。我们还将研究细胞悬浮液的喷墨打印的流体动力学，以了解在组织图案化过程中导致不可靠的细胞分配的现象和参数。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体平台，用于在细胞培养过程中控制微环境因素，如空间和时间上的氧张力。我们将开发方法来选择性地用保形涂层涂覆我们的微流体装置的特定区域，以使聚二甲基硅氧烷（PDMS）不渗透培养基中的营养物质或信号分子的吸收，同时保持氧气渗透性。所得到的系统将被设计为提供缺氧以及低氧培养条件，并且通过减少氧控制通道和细胞培养室之间的扩散距离，我们将创建循环时间为分钟量级的循环低氧条件。为了监测组织结构内的O2水平，我们将开发生物相容性的、载有氧敏感染料的微米级颗粒。这些颗粒将悬浮并分散在载有细胞的水凝胶珠粒内，从而允许光学、非侵入性、比率测量法原位测量组织构建体内部的氧水平。最后，我们还将开发获得这些组织的高质量图像的方法。通过具有几百微米厚度的组织切片成像将需要减少光散射的技术。通过在微流体平台上整合细胞培养环境，我们将能够监测细胞周期进展以及ECM重塑。 通过这项研究计划，我们将解决3D细胞培养、模型组织生成和组织表征所面临的一些最紧迫的挑战。从长远来看，这项工作的总体成果包括为基础科学开发微尺度组织模型以及为工业开发工具。这些结果的应用包括研究缺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.