The Laboratory for Therapeutic 3D Bioprinting, Department of Orthopaedic Surgery, Massachusetts General Hospital , Harvard Medical School, Boston, Massachusetts.
Tissue Eng Part C Methods. 2018 Apr;24(4):205-213. doi: 10.1089/ten.TEC.2017.0499. Epub 2018 Mar 27.
Collective cell migration, in which cells migrate as a group, is fundamental in many biological and pathological processes. There is increasing interest in studying the collective cell migration in high throughput. Cell scratching, insertion blocker, and gel-dissolving techniques are some methodologies used previously. However, these methods have the drawbacks of cell damage, substrate surface alteration, limitation in medium exchange, and solvent interference. The superhydrophobic surface, on which the water contact angle is greater than 150 degrees, has been recently utilized to generate patterned arrays. Independent cell culture areas can be generated on a substrate that functions the same as a conventional multiple well plate. However, so far there has been no report on superhydrophobic patterning for the study of cell migration. In this study, we report on the successful development of a robotically patterned superhydrophobic array for studying collective cell migration in high throughput. The array was developed on a rectangular single-well cell culture plate consisting of hydrophilic flat microwells separated by the superhydrophobic surface. The manufacturing process is robotic and includes patterning discrete protective masks to the substrate using 3D printing, robotic spray coating of silica nanoparticles, robotic mask removal, robotic mini silicone blocker patterning, automatic cell seeding, and liquid handling. Compared with a standard 96-well plate, our system increases the throughput by 2.25-fold and generates a cell-free area in each well non-destructively. Our system also demonstrates higher efficiency than conventional way of liquid handling using microwell plates, and shorter processing time than manual operating in migration assays. The superhydrophobic surface had no negative impact on cell viability. Using our system, we studied the collective migration of human umbilical vein endothelial cells and cancer cells using assays of endpoint quantification, dynamic cell tracking, and migration quantification following varied drug treatments. This system provides a versatile platform to study collective cell migration in high throughput for a broad range of applications.
细胞集体迁移是指细胞作为一个整体进行迁移，它在许多生物学和病理学过程中都起着基础性作用。目前，人们越来越感兴趣的是在高通量水平上研究细胞的集体迁移。细胞划痕、插入阻断器和凝胶溶解技术是之前使用过的一些方法。然而，这些方法存在细胞损伤、基底表面改变、介质交换受限和溶剂干扰等缺点。超疏水表面的水接触角大于 150 度，最近已被用于生成图案阵列。在与传统多孔板功能相同的基底上，可以生成独立的细胞培养区域。然而，到目前为止，还没有关于超疏水图案化用于研究细胞迁移的报道。在本研究中，我们报告了成功开发了一种机器人图案化的超疏水阵列，用于高通量研究细胞的集体迁移。该阵列是在一个由亲水平底微井通过超疏水表面分隔的矩形单孔细胞培养板上开发的。制造过程是机器人化的，包括使用 3D 打印对基底进行离散保护掩模的图案化、机器人喷涂二氧化硅纳米粒子、机器人掩模去除、机器人微型硅酮阻断器图案化、自动细胞播种和液体处理。与标准的 96 孔板相比，我们的系统将通量提高了 2.25 倍，并在每个孔中产生了非破坏性的无细胞区域。与使用微孔板的传统液体处理方法相比，我们的系统效率更高，与手动操作相比，迁移分析的处理时间更短。超疏水表面对细胞活力没有负面影响。使用我们的系统，我们通过终点定量分析、动态细胞跟踪和不同药物处理后的迁移定量分析，研究了人脐静脉内皮细胞和癌细胞的集体迁移。该系统为广泛的应用提供了一个高通量研究细胞集体迁移的多功能平台。
