Combining AFM and Acoustic Probes to Reveal Changes in the Elastic Stiffness Tensor of Living Cells

Abstract

Knowledge of how the elastic stiffness of a cell affects its communication with its environment is of fundamental importance for the understanding of tissue integrity in health and disease. For stiffness measurements, it has been customary to quote a single parameter quantity, e.g., Young's modulus, rather than the minimum of two terms of the stiffness tensor required by elasticity theory. In this study, we use two independent methods (acoustic microscopy and atomic force microscopy nanoindentation) to characterize the elastic properties of a cell and thus determine two independent elastic constants. This allows us to explore in detail how the mechanical properties of cells change in response to signaling pathways that are known to regulate the cell's cytoskeleton. In particular, we demonstrate that altering the tensioning of actin filaments in NIH3T3 cells has a strong influence on the cell's shear modulus but leaves its bulk modulus unchanged. In contrast, altering the polymerization state of actin filaments influences bulk and shear modulus in a similar manner. In addition, we can use the data to directly determine the Poisson ratio of a cell and show that in all cases studied, it is less than, but very close to, 0.5 in value.

Myosin II Activity Softens Cells in Suspension

“…Our observation of cell stiffening for myosin-inhibited cells deformed at timescales of seconds stands in clear contrast to data published on cells adherent to a substrate (26)(27)(28)(29). In these publications, the cells were reported to soften upon myosin inhibition.…”

“…In summary, cells, once suspended and free from any physical attachment to the surrounding, stiffen and become more solidlike after myosin II inhibition-contrary to reports of cell softening with adherent cells (26)(27)(28)(29). Measurements on adherent cells are generally done on cells adhering to plastic tissue culture dishes.…”

“…Phorbol 12,13-dibutyrate, on the other hand, activates the protein kinase C (24,25) and therefore increases myosin II activity. So far, there exists a range of data on mechanical property measurements of adherent cells treated with these drugs (26)(27)(28)(29), which have shown that inhibition of myosin activity leads to a loss of cell prestress and a subsequent lower measured elastic modulus of the cells.…”

Are cancer cells really softer than normal cells?

“…Because we focus here on microrheology experimental studies that explicitly compare individual cancer cells with normal cells (for more general reviews on rheology and cell mechanics see for instance (Wirtz, ; Yao et al., ; Haase & Pelling, )), computational approaches (see Katira et al., 2013 for a review) or techniques that have been used in the field of cell mechanics but not in the context of cancer are not detailed in the following. This is the case for instance of the uniaxial single cell microplate rheometer (Thoumine & Ott, ; N. Desprat, ), the recent method using magnetic nanowires (Chevry et al., ; Berret, ) or acoustic microscopy (Nijenhuis et al., ). Techniques that measure the stiffness of a tissue and do not resolve individual cells, such as ultrasound elastography (Tanter et al., ; Chamming's et al., ) or magnetic resonance elastography (MRE) (Pepin et al., ) will also not be discussed in detail here.…”

Atomic force microscopy studies on cellular elastic and viscoelastic properties

“…where υ is the Poisson ratio of cell; cells are often considered as incompressible material and thus υ=0.5 (Vargas-Pinto et al, 2013;Nijenhuis et al, 2014;Hecht et al, 2015). F is the applied loading force of AFM probe; δ is the indentation depth; E is the Young's modulus of the celll θ is the half-opening angle of the conical tip; and R is the radius of spherical tip.…”