We present a synergistic combination of simulations and experimental data on the dynamics of membrane adhesion. We show that a change in either the density or the strength of the bonds results in very different dynamics. Such behavior is explained by introducing an effective binding affinity that emerges as a result of the competition between the strength of the chemical bonds and the environment defined by the fluctuating membrane.
Prior to establishing tight contact with the endothelium, cells such as leukocytes or cancer cells use the recognition between sialyl-LewisX ligands and E-selectin receptors to establish weak, reversible adhesion and to roll along the vessel wall. We study the physical aspects of this process by constructing a mimetic system that consists of a giant fluid vesicle with incorporated lipid-anchored sialyl-LewisX molecules that bind to E-selectin that is immobilized on the flat substrate. The vesicles also carry a certain fraction of repelling PEG2000 molecules. We analyze the equilibrium state of adhesion in detail by means of reflection interference contrast microscopy and find that the adhesion process relies purely on the formation of one or more adhesion domains within the vesicle-substrate contact zone. We find that the content of ligands in the vesicle must be above 5 mol % to establish specific contacts. All concentrations of sialyl-LewisX above 8 mol % provide a very similar final state of adhesion. However, the size and shape of the adhesion domains strongly depend on both the concentrations of E-selectin (0-3500 molecules/microm2) and PEG2000 (0-5 mol %). At 3500 E-selectin molecules/microm2 and small concentrations of PEG2000, the vesicle-substrate contact is maximized and fully occupied by a single adhesion domain. At concentrations of 5 mol %, PEG2000 completely impedes the specific binding to any substrate. Lastly, an increase in the adhesion strength is observed in systems with identical compositions if the reduced volume of the vesicles is larger.
We establish a model of cell-tissue interaction consisting of vesicles carrying lipopolymers (to mimic the glycocalix) and mobile specific ligands of the blood platelet integrin α IIb β3 covering the substrate. We find the phase diagram with a first-order transition between a gravity-controlled weak state of the vesicle-substrate adhesion and a strong-adhesion state governed by receptor-ligand interaction. Adhesion energy ε adh is measured as a function of ligand and repeller concentration by interferometric contour analysis on the basis of a new refined model of soft shell adhesion (accounting for the membrane bending and stretching at the adhesion rim of the ellipsoidal vesicle). At ligand densities comparable to integrin density, ε adh decreases sharply. Increasing the repeller content weakens the adhesion strength.
We report the deformation and unbinding of weakly adhering giant vesicles under hydrodynamic shear forces. Linear shear fields are generated in a flat cell and vesicle adhesion onto a supported membrane is generated by electrostatic forces between oppositely charged lipids. The hydrodynamic flow in the aqueous medium near the outer side of the vesicle, within the vesicle and the tanktreading motion in the membrane are observed by tracing small markers attached to the vesicles or suspended in the medium by confocal laser scanning microscopy. The lift force generated by the rotational flow in the vesicle was estimated to be at least two orders of magnitude larger than predicted by the simple theory.
By use of a model system consisting of giant vesicles adhering to flat substrates, we identified, both experimentally and theoretically, two new control mechanisms for antagonist-induced deadhesion. Adhesion is established by specific binding of surface-grafted E-selectin and vesicle-carrying oligosaccharide Lewis(X). Deadhesion is achieved by controlled titration of monoclonal antibodies against E-selectin. The first mechanism is characterized by a considerable retraction of the contact zone resulting in a loss of contact area between the vesicle and the substrate. Within the developed theoretical framework, the observed equilibrium state is understood as a balance between the spreading pressure of the vesicle and the antagonist-induced lateral pressure at the edge of the contact zone. In the second mechanism, the antibodies induce unbinding by penetrating the contact zone without significantly affecting its size. This process reveals the decomposition of the adhesion zone into microdomains of tight binding separated by strongly fluctuating sections of the membrane. Both experiment and theory show a sigmoidal decrease of the number of bound ligands as a function of the logarithm of antagonist concentration. The work presented herein also provides a new method for the determination of the receptor binding affinity of either the surface-embedded ligands or the competing antagonist molecules.
Integrins are adhesion receptors that mediate cell adhesion and play an important function in many biological processes such as morphogenesis and tissue remodeling. These membrane proteins specifically interact with a short tripeptide sequence, RGD (Arg-Gly-Asp), present in numerous extracellular macromolecules. Model systems have been developed in order to understand how membrane adhesion is induced by this specific RGD peptide ligand/integrin recognition system. We have previously shown that RGD giant vesicles selectively adhere to endothelial cells by formation of pinning centers. Nevertheless, the nature of the lipids located in the adhesion contact zone is unknown. One hypothesis is that the lipidic ligands migrate to the contact zone where they are confined after binding to the receptor. To study the possible formation of ligand domains within the vesicle bilayer, we synthesized a fluorescently labeled RGD lipid that can be easily incorporated in giant vesicles. Adhesion of giant RGD vesicles onto an integrinfunctionalized surface was followed simultaneously by reflection interference contrast microscopy and fluorescence microscopy. For the first time, it was possible to observe the microsegregation of RGD lipids in the contact zone during adhesion. Additionally, we observed interesting photosensitive properties of the chalcone chromophore that could lead to a new method of analyzing the lipid organization within the membrane during adhesion and to the design of new ligand lipids and vesicle vectors for cell targeting.
A constrained cyclic ArgGly-Asp-D-Phe-Lys, abbreviated as cyclo(-RGDfK-), lipopeptide has been synthesized and incorporated into artificial membranes such as giant vesicles with DOPC and solid-supported lipid bilayers. The selective adhesion and spreading of endothelial cells of the human umbilical cord on solids functionalized by membranes with this RGD-lipopeptide have been observed. Furthermore, we have demonstrated strong selective adhesion of giant vesicles to endothelial cells through local adhesion domains by combined application of hydrodynamic flow field and reflection interference contrast microscopy (RICM). The adhesion can be inhibited by competition with a water-soluble RGD peptide. We suggest that this strategy could improve the efficiency of liposomes targeting used as vectors or as drug carriers to cells.
We have developed "vertical" magnetic tweezers capable of exerting controlled pico and subpico Newton forces. Using this apparatus, we apply a point-like force to a vesicle that is adhered by means of specific bonds between the vesicle-carrying oligosaccharide sialyl LewisX and the surface-grafted E-selectin. An exponential decrease of the bound vesicle area with the decay rate that is insensitive to the force and the composition of the system is observed. We measure an equilibrium under force that is associated with an increased binding in the center of the contact zone. We also show that the determination of the shape is potentially sufficient to determine the number of formed specific bonds.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.