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首页> 外文期刊>Journal of Biomechanics >Estimation of single stress fiber stiffness in cultured aortic smooth muscle cells under relaxed and contracted states: Its relation to dynamic rearrangement of stress fibers.
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Estimation of single stress fiber stiffness in cultured aortic smooth muscle cells under relaxed and contracted states: Its relation to dynamic rearrangement of stress fibers.

机译：在松弛和收缩状态下培养的主动脉平滑肌细胞中单应力纤维刚度的估计：与应力纤维的动态重排的关系。

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For a quantitative analysis of intracellular mechanotransduction, it is crucial to know the mechanical properties of actin stress fibers in situ. Here we measured tensile properties of cultured aortic smooth muscle cells (SMCs) in a quasi-in situ tensile test in relaxed and activated states to estimate stiffness of their single stress fibers (SFs). An SMC cultured on substrates was held using a pair of micropipettes and detached from the substrate while maintaining its in situ cell shape and cytoskeletal integrity. Stretching up to approximately 15% followed by unloading was repeated three times to stabilize their tension-strain curves in the untreated (relaxed) and 10 microM-serotonin-treated (activated) condition. Cell stiffness defined as the average slope of the loading limb of the stable loops was approximately 25 and approximately 40 nN/% in relaxed and activated states, respectively. It decreased to approximately 10 nN/% following SF disruption with cytochalasin D in both states. The number of SFs in each cell measured with confocal microscopy decreased significantly upon serotonin activation from 21.5+/-3.8 (mean+/-SD, n=80) to 17.5+/-3.9 (n=77). The dynamics of focal adhesions (FAs) were observed in adherent cells using surface reflective interference contrast microscopy. FAs aligned and elongated along the cell major axis following activation and then merged with each other, suggesting that the decrease in SFs was caused by their fusion. Average stiffness of single SFs estimated by the average decrease in whole-cell stiffness following SF disruption divided by the average number of SFs in each cell was approximately 0.7 and approximately 1.6 nN/% in the relaxed and activated states, respectively. Stiffening of single SFs following SF activation was remarkably higher than stiffening at the whole-cell level. Results indicate that SFs stiffen not only due to activation of the actomyosin interaction, but also due to their fusion, a finding which would not be obtained from analysis of isolated SFs.
机译：对于细胞内机械转导的定量分析，至关重要的是就地了解肌动蛋白应力纤维的机械性能。在这里，我们在松弛和激活状态下的准原位拉伸测试中测量了培养的主动脉平滑肌细胞（SMC）的拉伸性能，以估计其单应力纤维（SFs）的刚度。使用一对微量移液管固定在底物上培养的SMC，并与底物分离，同时保持其原位细胞形状和细胞骨架完整性。在未处理（松弛）和10 microM血清素处理（激活）的条件下，将拉伸至大约15％的位置重复拉伸三遍，以稳定其张力-应变曲线。单元刚度定义为稳定环的加载分支的平均斜率在松弛和激活状态下分别约为25和40 nN /％。在两种状态下，用细胞松弛素D破坏SF后，其降至约10 nN /％。血清素激活后，用共聚焦显微镜测量的每个细胞中的SF数量从21.5 +/- 3.8（平均+/- SD，n = 80）显着降低到17.5 +/- 3.9（n = 77）。使用表面反射干涉对比显微镜观察粘附细胞中粘着斑的动力学。激活后，FAs沿细胞主轴排列并伸长，然后彼此融合，表明SFs的降低是由于它们的融合引起的。通过SF破裂后全细胞刚度的平均降低除以每个细胞中SF的平均数来估计的单个SF的平均刚度在松弛和活化状态下分别为约0.7和约1.6nN /％。 SF激活后单个SF的增强明显高于全细胞水平的增强。结果表明，SF不仅由于肌动球蛋白相互作用的激活而变硬，而且还由于它们的融合而变硬，这一发现不能从分离的SF的分析中获得。

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