Abstract
The cell biomechanical properties play a key role in the determination of the changes during the essential cellular functions, such as contraction, growth, and migration. Recent advances in nano-technologies have enabled the development of new experimental and modeling approaches to study cell biomechanics, with a level of insights and reliability that were not possible in the past. The use of atomic force microscopy (AFM) for force spectroscopy allows nanoscale mapping of the cell topography and mechanical properties under, nearly physiological conditions. A proper evaluation process of such data is an essential factor to obtain accurate values of the cell elastic properties (primarily Young's modulus). Several numerical models were published in the literature, describing the depth sensing indentation as interaction process between the elastic surface and indenting probe. However, many studies are still relying on the nowadays outdated Hertzian model from the nineteenth century, or its modification by Sneddon. The lack of comparison between the Hertz/Sneddon model with their modern modifications blocks the development of advanced analysis software and further progress of AFM promising technology into biological sciences. In this work, we applied a rationalized use of mechanical models for advanced postprocessing and interpretation of AFM data. We investigated the effect of the mechanical model choice on the final evaluation of cellular elasticity. We then selected samples subjected to different physicochemical modulators, to show how a critical use of AFM data handling can provide more information than simple elastic modulus estimation. Our contribution is intended as a methodological discussion of the limitations and benefits of AFM-based advanced mechanical analysis, to refine the quantification of cellular elastic properties and its correlation to undergoing cellular processes in vitro.
Highlights
The determination of the biomechanical properties of cells and their surrounding extracellular matrix (ECM) unveils fundamental insights to understand the development and the features of healthy and pathological conditions (Yim and Sheetz, 2012)
We performed two-way repeated measures analysis of variance (ANOVA), with matching on the indentation levels, to assess whether using different models on the same samples will lead to statistically different results (Figure 2E)
We performed repeated measures one-way ANOVA using all the values calculated across the cell volumes, categorizing them only by the model used
Summary
The determination of the biomechanical properties of cells and their surrounding extracellular matrix (ECM) unveils fundamental insights to understand the development and the features of healthy and pathological conditions (Yim and Sheetz, 2012). Cellular mechanics is a dynamic process originating from the disposition and interaction of the cytoskeletal proteins (mainly actin microtubules and intermediate filaments) (Ahmed et al, 2015; Huber et al, 2015), and by its coupling to the cell nucleus (Li et al, 2014). A common regulation mechanism resides in the formation of focal adhesions between the actinintegrin transmembrane complex and target ECM binding sites (Huang et al, 2004). Focal adhesion formation induces the upregulation of different biomolecular cascades, including the Hippo pathway through YAP and TAZ transcription factors (Benham-Pyle et al, 2015). The alteration of cytoskeletal homeostasis is common in many pathological processes. Endothelial cells can respond to altered fluid dynamics and increased shear-stress with hypertrophy and tissue stiffening (Lu and Kassab, 2011)
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