A multi-material platform for imaging of single cell-cell junctions under tensile load fabricated with two-photon polymerization
A multi-material platform for imaging of single cell-cell junctions under tensile load fabricated with two-photon polymerization
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DOI：
10.1007/s10544-022-00633-z
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发表时间：
2022-10
影响因子：
2.8
通讯作者：
J. Rosenbohm;Grayson Minnick;Bahareh Tajvidi Safa;A. M. Esfahani;Xiaowei Jin;Haiwei Zhai;N. Lavrik;Ruiguo Yang
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文献类型：
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作者：
J. Rosenbohm;Grayson Minnick;Bahareh Tajvidi Safa;A. M. Esfahani;Xiaowei Jin;Haiwei Zhai;N. Lavrik;Ruiguo Yang
我们之前报道了一种由 IP-S 光树脂和双光子聚合 (TPP) 制成的单细胞粘附微拉伸测试仪 (SCAμTT)，用于研究在限定拉伸载荷下单细胞-细胞连接的力学。该平台的一个主要限制是 IP-S（用于 TPP 制造的光树脂）的自发荧光，这会显着增加背景信号并使拉伸细胞的荧光成像变得困难。在这项研究中，我们报告了一种新的 SCAμTT 平台的设计和制造，该平台可以减轻自发荧光，并证明其在单细胞对相互连接拉伸时成像的能力。通过采用 IP-S 和 IP-Visio（一种具有减少自发荧光的光树脂）的两种材料设计，我们发现该平台的自发荧光显着减少。此外，通过将孔径集成到带有金涂层的基底上，自发荧光对成像的影响几乎完全减轻。借助这个新平台，我们展示了对一对上皮细胞拉伸至 250% 应变时进行成像的能力，使我们能够观察连接破裂和 F-肌动蛋白收缩，同时记录连接处超过 800 kPa 的应力累积。这里介绍的平台和方法有可能能够详细研究细胞-细胞连接的力学和机械转导，并改进机械生物学应用中其他 TPP 平台的设计。图形摘要
We previously reported a single-cell adhesion micro tensile tester (SCAμTT) fabricated from IP-S photoresin with two-photon polymerization (TPP) for investigating the mechanics of a single cell-cell junction under defined tensile loading. A major limitation of the platform is the autofluorescence of IP-S, the photoresin for TPP fabrication, which significantly increases background signal and makes fluorescent imaging of stretched cells difficult. In this study, we report the design and fabrication of a new SCAμTT platform that mitigates autofluorescence and demonstrate its capability in imaging a single cell pair as its mutual junction is stretched. By employing a two-material design using IP-S and IP-Visio, a photoresin with reduced autofluorescence, we show a significant reduction in autofluorescence of the platform. Further, by integrating apertures onto the substrate with a gold coating, the influence of autofluorescence on imaging is almost completely mitigated. With this new platform, we demonstrate the ability to image a pair of epithelial cells as they are stretched up to 250% strain, allowing us to observe junction rupture and F-actin retraction while simultaneously recording the accumulation of over 800 kPa of stress in the junction. The platform and methodology presented here can potentially enable detailed investigation of the mechanics of and mechanotransduction in cell-cell junctions and improve the design of other TPP platforms in mechanobiology applications.Graphical abstract