Mechanical Phenotyping of Mouse Embryonic Stem Cells: Increase in Stiffness with Differentiation

Abstract

Atomic force microscopy (AFM) has emerged as a promising tool to characterize the mechanical properties of biological materials and cells. In our studies, undifferentiated and early differentiating mouse embryonic stem cells (mESCs) were assessed individually using an AFM system to determine if we could detect changes in their mechanical properties by surface probing. Probes with pyramidal and spherical tips were assessed, as were different analytical models for evaluating the data. The combination of AFM probing with a spherical tip and analysis using the Hertz model provided the best fit to the experimental data obtained and thus provided the best approximation of the elastic modulus. Our results showed that after only 6 days of differentiation, individual cell stiffness increased significantly with early differentiating mESCs having an elastic modulus two-to threefold higher than undifferentiated mESCs, regardless of cell line (R1 or D3 mESCs) or treatment. Singletouch (indentation) probing of individual cells is minimally invasive compared to other techniques. Therefore, this method of mechanical phenotyping should prove to be a valuable tool in the development of improved methods of identification and targeted cellular differentiation of embryonic, adult, and induced-pluripotent stem cells for therapeutic and diagnostic purposes.

Cellular Stiffness as a Novel Stemness Marker in the Corneal Limbus

“…Overall, mechanical comparisons of stem cells and their progeny have revealed that cells with epithelial-like morphologies are softer than cells with mesenchymal-like morphologies (22,23). The epithelial-to-mesenchymal transition observed during LEC differentiation, coupled with the finding presented here of a corresponding stiffness increase, further supports the contention that cells with a mesenchymal morphology are stiffer than epithelial cells.…”

“…Recently, various cell mechanical properties, including stiffness (i.e., Young's modulus) and size, have been shown to distinguish stem cell phenotypes from differentiated cells at the single-cell level for various stem cell types (20)(21)(22)(23)(24). Changes in biomechanical properties during stem cell differentiation could enable microfluidic approaches for on-the-fly analysis and sorting of stem cells from differentiated cells.…”

Effect of membrane stiffness and cytoskeletal element density on mechanical stimuli within cells: an analysis of the consequences of ageing in cells

“…For example, Pillarisetti et al (2011) measured non linear strain hardening in mouse embryonic cells using AFM indentation. In this study, a rigid conical indenter with a contact angle of 141 degrees was used to indent the cell model.…”

Differential regulation of morphology and stemness of mouse embryonic stem cells by substrate stiffness and topography

“…By contrast, rigid substrates promote the induction and differentiation of cardiomyocytes while mESCderived cardiac foci display functional synchrony with native cardiomyocytes [22]. Stemness preservation is also attributed to the low elastic modulus of undifferentiated mESCs placed on a stiff tissue culture dish compared to that for differentiated mESCs [23]. On the other hand, external mechanical stimuli can also manipulate mESC stemness.…”