Polymer Mixing and the Thermodynamics of Cell Adhesion at Fluid Interfaces

Abstract The mechanism of adhesion of cells to other cells or to non-cellular surfaces is a central problem in cell biology and biotechnology. The present studies were carried out to investigate the relationships between cell surface thermodynamics, the kinetics of cell adhesive behaviour and the molecular and morphological structure of the cell surface, when cell adhesion is elicited by defined physiological stimuli. The surface thermodynamic studies are applications of classical capillarity in which we measure the wetting of cell surfaces (the surface affinity or work of adhesion) by aqueous phase separated polymer solutions of dextran and poly (ethylene glycol), before and after exposure of the cells to the adhesive stimuli. For the phase system used here (4%/4% w/w dextran mol. wt. 2 × 106/poly (ethylene glycol) mol. wt. 2 × 104 in HEPES buffered physiological saline), the measured interfacial free energy was 4.02 × 10−6 J m−2. Isolated leukocytes were exposed to adhesion-promoting che-moattractant stimuli (for example, the tripeptide N-formyl methionyl leucyl phenylalanine or serum complement activated yeast particles). We observed increases in the work of cell adhesion to the dextran phase which were proportional to the stimulus dose; at the maximally effective doses, the cell-liquid-liquid contact angles changed by about 50°, corresponding to 2 × l06J m−2 changes in the work of adhesion. In parallel with the thermodynamic measurements we measured the kinetics of cell activation to provide the time scale of cell behaviour which is missing from the equilibrium surface energy measurements. Infrared photometry (optical density and perpendicular light scattering) provided information on the kinetics of cell adhesion and morphological responses to stimuli; simultaneous measurements of oxygen free radical-dependent chemiluminescence and spectrophotometric measurements of enzyme activity defined the kinetics of biochemical activation. The thermodynamic studies were carried out during the plateau of the time-response curves; under these conditions both the kinetic and thermodynamic measurements have similar stimulus dose-response curves, suggesting that both types of process are manifestations of the same cell surface events. The exact relation of the surface affinity changes to the molecular mechanism(s) of cell adhesion is not yet clear, however. Synthetic structural studies (model membrane reconstitution) indicate that the cell surface/polymer phase affinity is a function of the concentration, molecular weight and conformational state of the cell surface glycopolymers (the “glycocalyx”). Analytic studies have shown correlations between surface affinity changes and the molecular weight/concentration profiles of isolated cell membrane glycoproteins. Modelistic considerations suggest that the cell surface glycoproteins which determine the phase wetting behaviour have concentrations in the micromolar range. If these aqueous phase reactive glycocalyx proteins are the molecules which mediate cell adhesion elicited by small specifically recognised ligands, then it may be necessary to modify the current model of cell adhesion which proposes that adhesion results from a balance between “specific bonding and non-specific repulsion.” For example, in stimulus induced leukocyte adhesion a “specific induction, non-specific execution” model may be more useful.