A High-Throughput Workflow to Study Remodeling of Extracellular Matrix-Based Microtissues
A High-Throughput Workflow to Study Remodeling of Extracellular Matrix-Based Microtissues
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DOI：
10.1089/ten.tec.2018.0290
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发表时间：
2019-01-01
影响因子：
3
通讯作者：
Wood, David K.
中科院分区：
文献类型：
--
作者：
Cummins, Katherine A.;Crampton, Alexandra L.;Wood, David K.
细胞微环境的变化是许多病理学的组成特征，包括癌症、纤维化和自身免疫性疾病。目前可用于研究三维组织重塑的体外方法不适合高通量研究，因为它们不适合大规模实验。结合液滴微流体和图案化的低粘附培养表面，我们设计了一个工作流程，将细胞-细胞外基质（ECM）相互作用纳入一个通用的高通量平台，该平台与现有的高通量液体处理系统兼容，能够进行长期实验（>1个月），非常适合传统和新型生物测量。通过我们的平台，我们证明了胶原微组织作为组织水平功能的一种应用的高通量ECM重塑研究的可行性。在这项研究中，我们使用我们的工作流程来检查ECM重塑在组织，细胞和亚细胞水平，利用从免疫组织化学和活细胞成像，增殖和收缩测定的测定。凭借我们独特的培养系统，我们可以随着时间的推移跟踪单个结构，并在多个尺度上评估大群体的重塑。最后，我们证明了冷冻保存我们的微组织的能力，同时保持高活力和细胞功能，这是一种非常宝贵的方法，可以在大规模生产后传播和冷冻微组织。使用这些方法，我们的基于ECM的系统成为一个可行的平台，用于模拟以组织重组为特征的疾病，以及一种可扩展的方法来进行体外基于细胞的药物筛选和高通量生物发现的测定。
Changes to the cellular microenvironment are an integral characteristic of numerous pathologies, including cancer, fibrosis, and autoimmune disease. Current in vitro methodologies available to study three-dimensional tissue remodeling are ill-suited for high-throughput studies as they are not scalable for large-scale experiments. Combining droplet microfluidics and patterned low-adhesion culture surfaces, we have engineered a workflow to incorporate cell-extracellular matrix (ECM) interactions in a versatile and high-throughput platform that is compatible with existing high-throughput liquid handling systems, enables long-term experiments (>1 month), and is well suited for traditional and novel biological measurements. With our platform, we demonstrate the feasibility of high-throughput ECM remodeling studies with collagen microtissues as one application of a tissue-level function. In this study, we use our workflow to examine ECM remodeling at the tissue, cell, and subcellular levels, leveraging assays ranging from immunohistochemistry and live cell imaging, to proliferation and contraction assays. With our unique culture system, we can track individual constructs over time and evaluate remodeling on several scales for large populations. Finally, we demonstrate the ability to cryopreserve our microtissues while retaining high viability and cell function, an invaluable method that could allow for dissemination and freezing of microtissues after mass production. Using these methods, our ECM-based system becomes a viable platform for modeling diseases characterized by tissue reorganization as well as a scalable method to conduct in vitro cell-based assays for drug screening and high-throughput biological discovery.