Department of Mechanical Engineering, University of Toledo, 1650 N.Westwood Avenue, Toledo, OH 43606-3390, USA.
J Biomech. 2013 May 31;46(9):1524-30. doi: 10.1016/j.jbiomech.2013.03.023. Epub 2013 Apr 28.
The aim of this study was to obtain nanomechanical properties of living cells focusing on human amniotic fluid stem (hAFS) cell using nanoindentation techniques. We modified the conventional method of atomic force microscopy (AFM) in aqueous environment for cell imaging and indentation to avoid inherent difficulties. Moreover, we determined the elastic modulus of murine osteoblast (OB6) cells and hAFS cells at the nucleus and cytoskeleton using force-displacement curves and Hertz theory. Since OB6 cell line has been widely used, it was selected to validate and compare the obtained results with the previous research studies. As a result, we were able to capture high resolution images through utilization of the tapping mode without adding protein or using fixation methods. The maximum depth of indentation was kept below 15% of the cell thickness to minimize the effect of substrate hardness. Nanostructural details on the surface of cells were visualized by AFM and fluorescence microscopy. The cytoskeletal fibers presented remarkable increase in elastic modulus as compared with the nucleus. Furthermore, our results showed that the elastic modulus of hAFS cell edge (31.6 kPa) was lower than that of OB6 cell edge (42.2 kPa). In addition, the elastic modulus of nucleus was 13.9 kPa for hAFS cell and 26.9 kPa for OB6 cells. Differences in cell elastic modulus possibly resulted from the type and number of actin cytoskeleton organization in these two cell types.
本研究旨在利用纳米压痕技术获得活细胞的纳米力学特性，重点关注人羊水干细胞（hAFS）。我们对传统的原子力显微镜（AFM）在水相环境中的细胞成像和压痕方法进行了改进，以避免固有困难。此外，我们通过力-位移曲线和赫兹理论确定了核和细胞骨架中鼠成骨细胞（OB6）和 hAFS 细胞的弹性模量。由于 OB6 细胞系已被广泛应用，因此选择其用于验证并将获得的结果与先前的研究进行比较。结果，我们能够在不添加蛋白质或使用固定方法的情况下，通过利用轻敲模式获得高分辨率的图像。压痕的最大深度保持在细胞厚度的 15%以下，以最小化基底硬度的影响。细胞表面的纳米结构细节通过 AFM 和荧光显微镜进行可视化。与核相比，细胞骨架纤维的弹性模量显著增加。此外，我们的结果表明，hAFS 细胞边缘的弹性模量（31.6 kPa）低于 OB6 细胞边缘的弹性模量（42.2 kPa）。此外，hAFS 细胞的核弹性模量为 13.9 kPa，OB6 细胞的核弹性模量为 26.9 kPa。两种细胞类型的细胞弹性模量的差异可能是由于肌动蛋白细胞骨架组织的类型和数量不同所致。
