Role of endothelial cell-substrate contact area and fibronectin-receptor affinity in cell adhesion to HEMA/EMA copolymers

Abstract

The objective of this study was to examine the effect of substrate hydrophobicity on cell-substrate contact area and the affinity between adsorbed fibronectin (Fn) and its receptor. Homo- and copolymer films of hydrophobic ethyl methacrylate (EMA) and hydrophilic hydroxyethyl methacrylate (HEMA) were spun-cast onto glass slides. Bovine aortic endothelial cells (BAEC) were plated for 2 h in serum-free medium onto polymers preadsorbed with Fn. Cells were fixed, labeled, and examined by total internal reflection fluorescence microscopy (TIRFM) to determine the topography of the basal surface as a function of distance from the substrate. Phase contrast microscopy was used to examine the total projected area of adherent cells. The cumulative contact area was greatest on cells attached to surfaces prepared from 0% HEMA and lowest on surfaces with the highest HEMA content. An equilibrium adhesion model used these data together with the critical force for detachment and the Fn density (Burmeister et al., J Biomed Mater Res 1996;30:13-22) to determine the affinity between Fn and its receptor and the bond strength. The affinity and force per bond decreased with increasing HEMA content. These results indicate that differences in the strength of endothelial cell adhesion to polymers are influenced by the conformation of the adsorbed adhesion proteins.

Effects of surface properties and bioactivation of biomaterials on endothelial cells

“…In the same way, Fn adsorbed on a hydrophilic surface supports higher EC adhesion than when adsorbed on a hydrophobic surface and enhances cell retention (64,75). The protein exposes a higher density of cell binding sites when exposed to a hydrophilic substrate, consequently, cells may form more bonds with the underlying surface resulting in a higher adhesion strength (75)(76)(77). Again, surface hydrophilicity does not correlate well with cell responses (Evan Dubiel, Yves Martin, Patrick Vermette, manuscript in preparation).…”

Total Internal Reflection Fluorescence Microscopy: Applications in Biophysics

“…VA-TIRF has been used to map out depth-dependent concentration profiles of fluorescein and fluorescein-labeled IgG next to surfaces [85], pyrene in thin polymer films [96], and fluorescein and acridine orange next to surfaces [97]. This method has also been used to map out the topology of cell-substrate contact regions by applying the technology to adherent cells with fluorescently labeled cytoplasms or membranes [89,94,[98][99][100][101][102][103][104]. The goal in these studies has been to determine not only the separation distances between substrates and adjacent cell membranes, but also the nature of focal adhesion contacts.…”

“…In a series of papers, TIRFM was used to monitor the regions of contact between endothelial cells and various types of surfaces coated with fibronectin or RGD peptides while the cells detach under stress induced by flow. These measurements have application to the design of nonthrombogenic synthetic vascular grafts [101,104,198,199].…”

Large‐Scale Protein Arrays Generated with Interferometric Lithography for Spatial Control of Cell‐Material Interactions

“…Focal adhesions are on the order of 10 nm to 10 μm and serve as crucial outside-to-inside signaling gateways that are necessary for proper cell function [2]. The micrometer-and nanometer-scale organization of surface proteins is expected to play a critical role in adhesion complex formation and function [3][4][5][6][7]. Seminal work in the area of cell adhesion studies in Whitesides' laboratory patterned cell adhesive domains on the cellular scale (10s of microns), demonstrating that by controlling the shape and size of the adhesive domain, the shape and degree of physical interaction between the surface and the cell could be controlled [8][9][10][11].…”