Scanning Probe Microscopy Techniques for Studying the Cell Glycocalyx

Abstract

The glycocalyx is a brush-like layer that covers the surfaces of the membranes of most cell types. It consists of a mixture of carbohydrates, mainly glycoproteins and proteoglycans. Due to its structure and sensitivity to environmental conditions, it represents a complicated object to investigate. Here, we review studies of the glycocalyx conducted using scanning probe microscopy approaches. This includes imaging techniques as well as the measurement of nanomechanical properties. The nanomechanics of the glycocalyx is particularly important since it is widely present on the surfaces of mechanosensitive cells such as endothelial cells. An overview of problems with the interpretation of indirect data via the use of analytical models is presented. Special insight is given into changes in glycocalyx properties during pathological processes. The biological background and alternative research methods are briefly covered.

Keywords: atomic force nanoindentation; cell nanomechanics; glycocalyx; scanning probe microscopy.

Figure 3

(A) Principal of atomic force nanoindentation technique for determination of glycocalyx nanomechanical properties; (B) example of obtained force curve with (1) and without (2) glycocalyx. Region I corresponds to glycocalyx layer while region II to cell body; (C) Definition of geometrical parameters. Taken from [57] with permission.

Figure 1

AFM images of PC12 cells obtained in peak force tapping mode. Left: surface of native PC12 cells; right: surface of PC12 cells after enzymatic removal of glycocalyx. Modified from [43].

Figure 2

Distribution of (a) disconnection distance obtained in new AFM ringing mode and (b) height signal simultaneously recorded in peak force tapping mode on A375 human melanoma skin epithelial cells. Taken from [45] under Creative Commons license.

Figure 4

Comparison between morphological and nanomechanical properties of endothelial cells grown in static and flow conditions. (A,B) AFM images of cells, (C) glycocalyx stiffness, and (D) glycocalyx thickness. Statistics: (#) different at p = 0.05, (ns) non-significant. Modified from [60].

Figure 5

Alterations in NO production by rat fat pad endothelial cells after selective 10 min of pulling on the different components of the glycocalyx using the AFM probe. Right: scheme of cantilever functionalization. Statictics: (*) p < 0.05 compared to paired unstimulated regions Modified from [61].

Figure 6

Comparison between glycocalyx length (A) and effective coverage (B) of endothelial cells in ex vivo mouse aorta with diabetes progression. Statistics: (*) p < 0.0001; (ns) non-significant. Modified from [62].

Figure 7

Schematic representation of the differences in the structure of the pericellular layer of nonmalignant and cancerous cells and their changes after enzymatic treatment. Taken from [64] under Creative Commons license.