Characterizing the mechanics of cultured cell monolayers
Characterizing the mechanics of cultured cell monolayers
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DOI：
10.1073/pnas.1213301109
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发表时间：
2012-10-09
影响因子：
11.1
通讯作者：
Charras, Guillaume T.
中科院分区：
文献类型：
--
作者：
Harris, Andrew R.;Peter, Loic;Charras, Guillaume T.
单细胞厚的单层细胞是多细胞生物中最简单的组织，但它们在发育和正常生理中起着关键作用。在早期发育中，胚胎形态发生主要是由于内部产生的力引起的单层重排和变形。后来，单层作为物理屏障将内部环境与外部分开，并且必须承受外部施加的力。虽然抵抗和产生机械力是单层功能的重要组成部分，但缺乏简单的实验方法来表征单层机械性能。在这里，我们描述了一种用于自由悬浮培养单层的拉伸测试系统，该系统能够在多细胞、单细胞和亚细胞尺度上检查其机械行为。使用这个系统，我们提供单层弹性的测量，并表明，这是两个数量级大于其孤立的细胞成分的弹性。单分子层在细胞间连接破裂之前可以承受两倍以上的长度。在断裂应力的测量，使第一次估计的平均所需的力量，以真正成熟的单层细胞，大约九倍以上的分离细胞对测量。在单细胞中，单层的机械性能强烈依赖于肌动蛋白细胞骨架，肌球蛋白，和细胞间的粘附界面相邻细胞的完整性。高放大率成像显示，角蛋白丝在延伸过程中逐渐拉伸，表明它们参与单层力学。这种多尺度研究单层响应变形使我们的设备提供了第一个定量调查单层生物学和力学之间的联系。
One-cell-thick monolayers are the simplest tissues in multicellular organisms, yet they fulfill critical roles in development and normal physiology. In early development, embryonic morphogenesis results largely from monolayer rearrangement and deformation due to internally generated forces. Later, monolayers act as physical barriers separating the internal environment from the exterior and must withstand externally applied forces. Though resisting and generating mechanical forces is an essential part of monolayer function, simple experimental methods to characterize monolayer mechanical properties are lacking. Here, we describe a system for tensile testing of freely suspended cultured monolayers that enables the examination of their mechanical behavior at multi-, uni-, and subcellular scales. Using this system, we provide measurements of monolayer elasticity and show that this is two orders of magnitude larger than the elasticity of their isolated cellular components. Monolayers could withstand more than a doubling in length before failing through rupture of intercellular junctions. Measurement of stress at fracture enabled a first estimation of the average force needed to separate cells within truly mature monolayers, approximately ninefold larger than measured in pairs of isolated cells. As in single cells, monolayer mechanical properties were strongly dependent on the integrity of the actin cytoskeleton, myosin, and intercellular adhesions interfacing adjacent cells. High magnification imaging revealed that keratin filaments became progressively stretched during extension, suggesting they participate in monolayer mechanics. This multiscale study of monolayer response to deformation enabled by our device provides the first quantitative investigation of the link between monolayer biology and mechanics.