Institute of Analytical and Bioanalytical Chemistry, University of Ulm, Albert-Einstein-Allee 11, 89081 Ulm, Germany.
Analyst. 2012 Nov 21;137(22):5208-14. doi: 10.1039/c2an36001b. Epub 2012 Sep 14.
Mechanical forces affect biological systems in their natural environment in a widespread manner. Mechanical stress may either stimulate cells or even induce pathological processes. Cells sensing mechanical stress usually respond to such stressors with proliferation or differentiation. Hence, for in vitro studies, the ability to impose a controlled mechanical stress on cells combined with appropriate analytical tools providing an immediate answer is essential to understand such fundamental processes. Here, we present a novel uniaxial motorized cell stretching device that has been integrated into a combined fluorescence microscope (FM)-atomic force microscope (AFM) system, thereby enabling high-resolution topographic and fluorescent live cell imaging. This unique tool allows the investigation of mechanotransduction processes, as the cells may be exposed to deliberately controlled mechanical stress while simultaneously facilitating fluorescence imaging and AFM studies. The developed stretching device allows applying reproducible uniaxial strain from physiologically relevant to hyperphysiological levels to cultured cells grown on elastic polydimethylsiloxane (PDMS) membranes. Exemplarily, stretching experiments are shown for transfected squamous cell carcinoma cells (SCC-25) expressing fluorescent labeled cytokeratin, whereby fluorescence imaging and simultaneously performed AFM measurements reveal the cytokeratin (CSK) network. Topographical changes and mechanical characteristics such as elasticity changes were determined via AFM while the cells were exposed to mechanical stress. By applying a cell deformation of approx. 20%, changes in the Young's modulus of the cytoskeletal network due to stretching of the cells were observed. Consequently, integrating a stretching device into the combined atomic force-fluorescence microscope provides a unique tool for dynamically analyzing structural remodeling and mechanical properties in mechanically stressed cells.
机械力在其自然环境中以广泛的方式影响生物系统。机械应力既可以刺激细胞，甚至可以诱导病理过程。感知机械应力的细胞通常会通过增殖或分化来响应这种应激源。因此，对于体外研究，将受控机械应力施加到细胞上的能力与提供即时答案的适当分析工具相结合，对于理解这些基本过程至关重要。在这里，我们展示了一种新颖的单轴电动细胞拉伸装置，该装置已集成到组合荧光显微镜（FM）-原子力显微镜（AFM）系统中，从而能够实现高分辨率的地形和荧光活细胞成像。这种独特的工具允许研究机械转导过程，因为细胞可以暴露于故意控制的机械应力，同时促进荧光成像和 AFM 研究。开发的拉伸装置允许将从生理相关到超生理水平的可重复的单轴应变应用于在弹性聚二甲基硅氧烷（PDMS）膜上生长的培养细胞。例如，展示了用于表达荧光标记细胞角蛋白的转染鳞状细胞癌细胞（SCC-25）的拉伸实验，其中荧光成像和同时进行的 AFM 测量揭示了细胞角蛋白（CSK）网络。通过 AFM 确定了拓扑变化和机械特性，例如弹性变化，而细胞则暴露于机械应力下。通过施加约 20%的细胞变形，观察到由于细胞拉伸导致细胞骨架网络的杨氏模量发生变化。因此，将拉伸装置集成到组合原子力-荧光显微镜中为动态分析机械应激细胞中的结构重塑和机械特性提供了独特的工具。
