Pericellular Brush and Mechanics of Guinea Pig Fibroblast Cells Studied with AFM

Abstract

The atomic force microscopy (AFM) indentation method combined with the brush model can be used to separate the mechanical response of the cell body from deformation of the pericellular layer surrounding biological cells. Although self-consistency of the brush model to derive the elastic modulus of the cell body has been demonstrated, the model ability to characterize the pericellular layer has not been explicitly verified. Here we demonstrate it by using enzymatic removal of hyaluronic content of the pericellular brush for guinea pig fibroblast cells. The effect of this removal is clearly seen in the AFM force-separation curves associated with the pericellular brush layer. We further extend the brush model for brushes larger than the height of the AFM probe, which seems to be the case for fibroblast cells. In addition, we demonstrate that an extension of the brush model (i.e., double-brush model) is capable of detecting the hierarchical structure of the pericellular brush, which, for example, may consist of the pericellular coat and the membrane corrugation (microridges and microvilli). It allows us to quantitatively segregate the large soft polysaccharide pericellular coat from a relatively rigid and dense membrane corrugation layer. This was verified by comparison of the parameters of the membrane corrugation layer derived from the force curves collected on untreated cells (when this corrugation membrane part is hidden inside the pericellular brush layer) and on treated cells after the enzymatic removal of the pericellular coat part (when the corrugations are exposed to the AFM probe). We conclude that the brush model is capable of not only measuring the mechanics of the cell body but also the parameters of the pericellular brush layer, including quantitative characterization of the pericellular layer structure.

Knock Down of Plakophillin 2 Dysregulates Adhesion Pathway through Upregulation of miR200b and Alters the Mechanical Properties in Cardiac Cells

“…Other models could be used for this task. One fairly new and accurate is the "Brush Model" developed by Sokolov et al [40][41][42], which can be useful to isolate the mechanical response of the cell from deformation of the pericellular layer surrounding the cell itself. The model used in the present study does not consider the brush but was chosen as (i) it is the most commonly used model, (ii) the same protocol/methodology has been used for all cells and indentations.…”

Atomic Force Microscopy Detects the Difference in Cancer Cells of Different Neoplastic Aggressiveness via Machine Learning

“…Five micrometer silica AFM probes were used in this study. The probes were prepared by utilizing tipless AFM cantilevers (Bruker, Inc., Santa Barbara, CA, a nominal spring constant of 0.06 N m −1 ; the exact spring constant was defined using thermal tuning method before gluing the silica probe) as described, e.g., in the study by Dokukin et al [ 32 ] Force maps (16 × 16 and 32 × 32 pixels) with a vertical ramping size of 5 μm were recorded for each cell and cell's cluster correspondingly. Vertical speed in all experiments was set to 5 μm s −1 , allowed to record one pixel within 1.5 s. For an example of 16 × 16 pixels, the time of acquisition of such a map will be 16 × 16 s, which is a bit more than 6 min; acquisition of 32 × 32 pixel image will require 26 min.…”

“…This model was suggested in the study by Sokolov et al, [ 30 ] and later developed in previous studies. [ 9,31,32 ] As was shown, considering the pericellular layer and the use of a large colloidal probe are essential for self‐consistency of the Hertz model, which is used to extract the effective Young's modulus of the cell body (note that the Hertz model is used because there is no adhesion between the AFM probe and cell surface measured in liquids). This model is described in detail in the references mentioned earlier and recently overviewed.…”

Biomechanical measurement and analysis of colchicine-induced effects on cells by nanoindentation using an atomic force microscope

“…Nikkhah et al (2011) reported that nonmalignant breast epithelial cells had significant higher Young's moduli than their malignant counterparts. Dokukin et al (2016) reported that the force signature of the pericellular brush layer of Guinea pig fibroblast cells was significantly changed after a treatment with hyaluronidase. Kasas et al (2013) investigated the nanomechanical properties (Young's modulus, deformability and adhesion) of biological samples (mammalian cells, plant cells, yeast cells, bacteria and viruses).…”