https://www.seas.upenn.edu/%7Edischer/pdfs/Cell_on_Gel-CELLpaper.pdf

Matrix elasticity directs stem cell lineage specification.

http://experimentationlab.berkeley.edu/sites/default/files/AFMImages/butt_AFM.pdf

Force measurements with the atomic force microscope: Technique, interpretation and applications

The morphology and cytoskeletal structure of fibroblasts, endothelial cells, and neutrophils are documented for cells cultured on surfaces with stiffness ranging from 2 to 55,000 Pa that have been laminated with fibronectin or collagen as adhesive ligand. When grown in sparse culture with no cell-cell contacts, fibroblasts and endothelial cells show an abrupt change in spread area that occurs at a stiffness range around 3,000 Pa. No actin stress fibers are seen in fibroblasts on soft surfaces, and the appearance of stress fibers is abrupt and complete at a stiffness range coincident with that at which they spread. Upregulation of 5 integrin also occurs in the same stiffness range, but exogenous expression of 5 integrin is not sufficient to cause cell spreading on soft surfaces. Neutrophils, in contrast, show no dependence of either resting shape or ability to spread after activation when cultured on surfaces as soft as 2 Pa compared to glass. The shape and cytoskeletal differences evident in single cells on soft compared to hard substrates are eliminated when fibroblasts or endothelial cells make cell-cell contact. These results support the hypothesis that mechanical factors impact different cell types in fundamentally different ways, and can trigger specific changes similar to those stimulated by soluble ligands. Cell Motil. Cytoskeleton 60:24 –34, 2005. © 2004 Wiley-Liss, Inc.

/pdf/effects-of-substrate-stiffness-on-cell-morphology-59za0lba3e.pdf

Effects of substrate stiffness on cell morphology, cytoskeletal structure, and adhesion

Change in cell stiffness is a new characteristic of cancer cells that affects the way they spread1,2. Despite several studies on architectural changes in cultured cell lines1,3, no ex vivo mechanical analyses of cancer cells obtained from patients have been reported. Using atomic force microscopy, we report the stiffness of live metastatic cancer cells taken from the body (pleural) fluids of patients with suspected lung, breast and pancreas cancer. Within the same sample, we find that the cell stiffness of metastatic cancer cells is more than 70% softer, with a standard deviation over five times narrower, than the benign cells that line the body cavity. Different cancer types were found to display a common stiffness. Our work shows that mechanical analysis can distinguish cancerous cells from normal ones even when they show similar shapes. These results show that nanomechanical analysis correlates well with immunohistochemical testing currently used for detecting cancer.

Nanomechanical analysis of cells from cancer patients

Contractile myocytes provide a test of the hypothesis that cells sense their mechanical as well as molecular microenvironment, altering expression, organization, and/or morphology accordingly. Here, myoblasts were cultured on collagen strips attached to glass or polymer gels of varied elasticity. Subsequent fusion into myotubes occurs independent of substrate flexibility. However, myosin/actin striations emerge later only on gels with stiffness typical of normal muscle (passive Young's modulus, E approximately 12 kPa). On glass and much softer or stiffer gels, including gels emulating stiff dystrophic muscle, cells do not striate. In addition, myotubes grown on top of a compliant bottom layer of glass-attached myotubes (but not softer fibroblasts) will striate, whereas the bottom cells will only assemble stress fibers and vinculin-rich adhesions. Unlike sarcomere formation, adhesion strength increases monotonically versus substrate stiffness with strongest adhesion on glass. These findings have major implications for in vivo introduction of stem cells into diseased or damaged striated muscle of altered mechanical composition.

/pdf/myotubes-differentiate-optimally-on-substrates-with-tissue-2s499cqco3.pdf

Myotubes differentiate optimally on substrates with tissue-like stiffness: pathological implications for soft or stiff microenvironments

Drug-Induced Changes of Cytoskeletal Structure and Mechanics in Fibroblasts: An Atomic Force Microscopy Study

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 flat with average heights of 0.4–0.8 μm, stable edges smoothly ascend to 2–3 μm within about 6 μm of the edge. In the region of the leading edge, the height fluctuates up to 50% (SD) of the mean value, much more than the stable edge; this fluctuation presumably reflects 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 ≈12 kPa in this case. The thinness of the leading edge precludes accurate estimation of the E values, but within 4 μm 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 and/or 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.

https://www.pnas.org/content/pnas/96/3/921.full.pdf

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

Afm imaging and elasticity measurements on living rat liver macrophages

Investigating the Cytoskeleton of Chicken Cardiocytes with the Atomic Force Microscope

We have used an atomic force microscope (AFM) to measure locally electrostatic interaction between the AFM's tip and a charged surface We investigated small amphiphilic positively charged bilayer patches adsorbed to the negatively charged mica surface in various electrolytes at different concentrations Thus we could determine the lateral resolution of the AFM in detecting repulsive and attractive electrostatic forces to be better than 25 nm We have investigated the electrostatic interactions between the tip and the sample in a variety of mono- and bivalent electrolytes In addition, when imaging these structured surfaces with the AFM, we could observe contrast reversal at low forces, which can be explained by the electrostatic forces between tip and sample

Christian Rotsch

Papers

Drug-Induced Changes of Cytoskeletal Structure and Mechanics in Fibroblasts: An Atomic Force Microscopy Study

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

Afm imaging and elasticity measurements on living rat liver macrophages

Investigating the Cytoskeleton of Chicken Cardiocytes with the Atomic Force Microscope

Mapping Local Electrostatic Forces with the Atomic Force Microscope