Dimensional and mechanical dynamics of active and stable edges in motile fibroblasts investigated by using atomic force microscopy

Abstract

The atomic force microscope (AFM) was employed to investigate the extension and retraction dynamics of protruding and stable edges of motile 3T3 fibroblasts in culture. Such dynamics closely paralleled the results of earlier studies employing video microscopy that indicated that the AFM force-mapping technique does not appreciably perturb these dynamics. Force scans permitted height determinations of active and stable edges. Whereas the profiles of active edges are f lat with average heights of 0.4-0.8 m, stable edges smoothly ascend to 2-3 m within about 6 m of the edge. In the region of the leading edge, the height f luctuates up to 50% (SD) of the mean value, much more than the stable edge; this f luctuation presumably ref lects differences in underlying cytoskeletal activity. In addition, force mapping yields an estimate of the local Young's modulus or modulus of elasticity (E, the cortical stiffness). This stiffness will be related to ''cortical tension,'' can be accurately calculated for the stable edges, and is Ϸ12 kPa in this case. The thinness of the leading edge precludes accurate estimation of the E values, but within 4 m of the margin it is considerably smaller than that for stable edges, which have an upper limit of 3-5 kPa. Although blebbing cannot absolutely be ruled out as a mechanism of extension, the data are consistent with an actin polymerization and͞or myosin motor mechanism in which the average material properties of the extending margin would be nearly constant to the edge. Because the leading edge is softer than the stable edge, these data also are consistent with the notion that extension preferentially occurs in regions of lower cortical tension.

Measuring Cell Viscoelastic Properties Using a Microfluidic Extensional Flow Device

“…For proof-of-principle studies, we focused on NIH 3T3 fibroblasts, which have been mechanically characterized by AFM (46,47), optical stretching (48,49), and MPA (50). To explore the potential of our cross-slot device for novel discovery, we also characterized GBM TICs, a stem-like subpopulation of GBM tumors thought to drive tumor initiation, recurrence, and therapeutic resistance (26,51,52).…”

Imaging and measuring the rituximab-induced changes of mechanical properties in B-lymphoma cells using atomic force microscopy

“…In the past decade, the atomic force microscopy (AFM) has been proven to be a powerful, versatile and easy-to-use tool for investigating the mechanical properties of single cells [13][14][15][16][17][18][19][20][21][22][23]. First, AFM can work in fluids and this enables the researchers to perform the experiments on living cells in situ.…”

“…Third, AFM is easy to control and the sample preparation is simple. With AFM nanoindentation technique, the mechanical properties of various cells have been measured, including breast cancer cells [14], fibroblasts [15,16,22], microbial cells [17], leukemia cells 0006-291X/$ -see front matter Ó 2010 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2010.12.043 [18,19], normal rat kidney (NRK) cell [20], and lymphocytes [21], providing novel information for understanding the cellular physiological activities.…”

AFM as a tool to probe and manipulate cellular processes

“…To date, very few experiments have been designed to evaluate the mechanical contributions of individual cells to specific physiological responses. The importance of the mechanical responses and behaviour of single cells is especially relevant in dynamic physiological processes such as tissue remodelling, cell motility and signal transduction in various mechanically sensitive cells such as those involved in hearing, bone remodelling and pressure sensing in vascular systems [36,37,39,[41][42][43][44][45][46]. Several new concepts, describing mechanical responses in cells, have emerged since the development of AFM-thanks to its ability to apply or record minute forces on individual cells.…”

“…AFM force experiments on living cells have also opened up new vistas in the investigation of the mechanical properties of individual cells and have demonstrated the importance of these mechanical properties to various physiological and physiopathological processes [17,19,[35][36][37][38][39][40]. This review will discuss some recent contributions of the AFM to cell physiology with a focus on the probing and nanomanipulation of structures in living cell.…”