Functional Incorporation of Integrins into Solid Supported Membranes on Ultrathin Films of Cellulose: Impact on Adhesion

Abstract

Biomimetic models of cell surfaces were designed to study the physical basis of cell adhesion. Vesicles bearing reconstituted blood platelet integrin receptors alpha(IIb)beta(3) were spread on ultrathin films of cellulose, forming continuous supported membranes. One fraction of the integrin receptors, which were facing their extracellular domain toward the aqueous phase, were mobile, exhibiting a diffusion constant of 0.6 micro m(2) s(-1). The functionality of receptors on bare glass and on cellulose cushions was compared by measuring adhesion strength to giant vesicles. The vesicles contained lipid-coupled cyclic hexapeptides that are specifically recognized by integrin alpha(IIb)beta(3). To mimic the steric repulsion forces of the cell glycocalix, lipids with polyethylene glycol headgroups were incorporated into the vesicles. The free adhesion energy per unit area deltag(ad) was determined by micro-interferometric analysis of the vesicle's contour near the membrane surface in terms of the equilibrium of the elastic forces. By accounting for the reduction of the adhesion strength by the repellers and from measuring the density of receptors one could estimate the specific receptor ligand binding energy. We estimate the receptor-ligand binding energy to be 10 k(B)T under bioanalogue conditions.

Force-induced growth of adhesion domains is controlled by receptor mobility

“…1b). Because integrin mobility is the only difference between the two systems, the role of receptor-mobility can be clearly elucidated.Studies on similar models (11,13,(15)(16)(17)(18)(19) have contributed to understanding the initial stages of cell adhesion in the absence of force. The adhering vesicle assumes a truncated oblate shape where a given fraction of the membrane is parallel to the substrate forming a contact zone (Fig.…”

“…In traditional reflection interference contrast microscopy (RICM) (18)(19)(20), the height of the vesicle membrane above the substrate is determined and micrometer-sized domains with densely packed bonds are identified (11,20). Dy-RICM exploits the vertical resolution of the set-up, which, at 5 nm, is an order of magnitude higher than the lateral resolution.…”

“…Although it has been mooted previously that, at shorter time scales, the response is dominated by the physical properties of the membrane (1,10), the force-response of cells over subsecond time scales has rarely been explored. At the same time, several studies have shown that, even in the absence of force, the adhesion is enhanced if both the ligands and the receptors mediating the binding are mobile (11,12), in comparison with the case when only one of the binding partners diffuses in the membrane. Nevertheless, most recent experiments on mechanoresponse/sensing have been on cells adhering to a substrate decorated with immobilized receptors, the assumption being that receptor mobility does not seriously affect the force response.…”

“…Our model (11,13) consists of a giant unilamellar vesicle with mobile RGD-peptide-carrying lipids (14), which interact with a supported bilayer doped with ␣ IIb ␤ 3 integrins (14). Depending on the specific deposition technique used (11,13), the integrins are either fixed (immobile system; Fig.…”

Bioactive compounds immobilized on Ti and TiNbHf: AFM-based investigations of biofunctionalization efficiency and cell adhesion

“…Nevertheless, proteins are sensitive to pH and temperature changes, which disrupt the weak bonds between chains, causing proteins to lose their secondary structure and denature. Even the contact to surfaces can induce their denaturation, which in turn decreases or abolishes their bioactive function [27]. In order to avoid full-length protein structures that are prone to denaturation, an alternative is to mimic the adhesive function of ECM proteins by synthesizing their particular adhesion motifs.…”