Donald J.L. McIver

Determination of cell/medium interfacial tensions from contact angles in aqueous polymer systems

The contact angles on cell layers of a series of polymeric droplets from aqueous two-phase systems of dextran and poly(ethylene glycol) have been used to determine the critical or limiting interfacial tension for spreading on the cell layers. Test droplets of the denser dextran-rich phase were formed in the lighter poly(ethylene glycol)-rich phase. The interfacial tensions, gamma, between the phases were determined with the pendant drop method, and a linear relationship was found between gamma-1/2 and the cosine of the angle the droplets made with the cell layers (Good-Girifalco plot). We were thus able to determine the limiting or critical interfacial tension, gamma c, for spreading on the cell layers. The value of gamma c is a measure of the interfacial energy of the cell/bathing medium interface. Values of gamma c obtained by this method include the following: 0.65 and 0.84 microN . m-1 for human erythrocytes and neutrophils, respectively; 0.93 microN . m-1 for porcine pulmonary macrophages; 0.75--3.60 microM . m-1 for various transformed murine lymphoid cell lines, and 2.53 microN . m-1 for Balb/c murine spleen lymphocytes. Exposure to various agents has differing effects on gamma c. Concanavalin A reduces gamma c, and bacterial lipopolysaccharide increases gamma c of murine spleen lymphocytes. The calcium ionophore, A23187, increases gamma c of both porcine pulmonary macrophages and murine spleen lymphocytes. This new method provides a quantitative approach to the cell surface energy and hydrophobicity which are thought to play an important role in membrane-mediated phenomena and in cell adhesion.

Activated neutrophils impair rabbit heart recovery after hypothermic global ischemia

Cardiopulmonary bypass is known to cause neutrophil activation, and activated neutrophils appear to be of importance in myocardial reperfusion injury. This study examined the effect of a preischemic infusion of activated neutrophils on the recovery of myocardial function after 40 minutes of hypothermic global ischemia. Studies were carried out in three groups of Langendorff-perfused rabbit hearts: control, control (unactivated) neutrophil infusion, and phorbol myristate acetate-activated neutrophil infusion. The activated neutrophil group showed significant deterioration in function during the activated neutrophil infusion. All three groups demonstrated significant depression of function initially after reperfusion, but the two control groups subsequently recovered to baseline levels. The activated neutrophil group, however, showed a persistent significant depression in ventricular force, rate of ventricular tension development, and rate of ventricular relaxation as well as a significant increase in coronary vascular resistance. It is concluded that activated neutrophils depress myocardial function and contribute to impaired recovery of function after global hypothermic ischemia.

Interfacial tensions in healthy and atherosclerotic rabbit aortae

Interfacial tensions at the saline/arterial wall interface were determined by measuring contact angles between various test fluid droplets and the walls of rabbit aortae immersed in physiological saline. These contact angles and the interfacial tensions of the test fluid/bathing fluid interface (measured by the Du Noüy ring method) were converted to saline/arterial wall interfacial tensions by applying Neumanns equation of state. Four diseased animals, fed an atherogenic diet for 6-8 weeks and 6 controls formed the experimental group. A significantly higher interfacial tension (P < 0.001), was determined for lesion surfaces in atherosclerotic arteries (0.36 +/- 0.08 (SEM) mM . M-1, n = 13) compared to both the surrounding undisturbed regions (0.035 +/- 0.01 mN . m-1, n = 14) and the intact surface of control vessels (0.060 +/- mN . m-1, n = 48). This increase may reflect a change in the strength of hydrophilic interactions associated with the lesion surface in atherogenesis.

Interfacial tensions at lipid—water interfaces. comparison of equation-of-state predictions with direct experimental measurements

Studies of interfacial tensions in solid/liquid/vapor systems usually take as their starting point the equation derived by Young in 1805. However, both the derivation and the application of the equation of state have so far been limited to solid/liquid/vapor interfaces in 2-component, 3-phase systems. There are a number of solid/liquid/liquid systems where determination of vapor/solid interfaces would be of considerable interest. There appears to be no intuitive objection to the applicability of the equation of state at such interfaces, but it is possible, for example, that entropic effects at an oil-water interface could introduce errors which play little role at the more well-behaved solid/vapor interface. Accordingly, the application of the equation of state at lipid/water/oil interface has been investigated where interfacial tensions may be measured directly from deformation measurements and compared with indirect estimates from contact angle data and the equation of state. 10 references.

A synthetic emulsion reproduces the functional properties of pulmonary surfactant

An effective substitute for pulmonary surfactant must be able to reduce water-vapour surface tensions to under 1 mN/m and it must spread rapidly and spontaneously at the air interface from the aqueous phase. Pure dipalmitoylphosphatidylcholine meets the former requirement, but not the latter. A synthetic surfactant is described which meets both of these criteria. The surfactant is prepared as a DPPC monolayer stabilizing an aqueous emulsion of an inert fluorocarbon oil; it spreads rapidly at the air/liquid interface, lowers the surface tension to under 1 mN/m during compression at the air/liquid interface and restores normal pressure-volume characteristics to surfactant-depleted lungs.

Membrane glycoprotein and surface free energy changes in hypoxic fibroblast cells

Hypoxia affects the biochemistry of mammalian cells and thus alters their sensitivity to subsequent chemo- and radiotherapy. When V79 Chinese hamster lung fibroblasts were grown under conditions of extreme hypoxia (less than 10 ppm O2) there was a significant shift in the membrane glycoprotein composition. Scanning electron microscopy revealed altered cell surface morphology including loss of pseudopodial projections. Experiments to determine changes in interfacial free energy of these cells using equilibrium two phase systems of poly(ethylene glycol) (PEG) and dextran were carried out. Test fluid droplets of the denser dextran-rich phase were formed on layers of cells in the PEG-rich phase as the bathing medium, and the contact angles the droplets made with the cell layers were measured from photomicrographs. The contact angles on cells in the plateau phase increased significantly with time of exposure to hypoxia, from 25 degrees (zero time) to 35 degrees (6 h) to 60 degrees (9 h). Contact angles on cells in the exponential phase increased from 80 degrees (zero time) to 150 degrees after 20 h of hypoxia. It appears that the altered contact angles reflect changes in cell surface hydrophobicity that may, in part, reflect alterations in the membrane glycoprotein composition.

Pressure distortion of an artificial membrane and the effect of ligand/protein binding

Abstract Fourier transform infrared spectroscopy was employed to study configuration changes induced in a model membrane exposed to very high (to 25 kbar) hydrostatic pressures. The model membrane consisted of 25% dimyristoyl- l -α-phosphatidylcholine (DMPC) and 12.5% human albumin suspended in 62.5% heavy water (D 2 O). Clofibrate (10 −2 M), which binds avidly to albumin, was used as a ligand. The suspension formed a lipid bilayer to which albumin was attached by hydrogen bonding of a side chain to a carbonyl group of DMPC. The protein structure remained mainly helical with some β-sheeting. Hydrogen bond strength within the protein was increased by lipid-protein interactions. Clofibrate bound to the lipid-albumin complex and reduced somewhat the β-sheet / α-helix ratio. Increasing pressure increased the intermolecular interactions between albumin and DMPC as indicated by the increasing intensity of a band near 1587 cm −1 . Clofibrate reduced this effect by preventing pressure-enhanced protein interactions with DMPC. The results indicate that model membrane physical chemistry is altered by pressure and this may have consequences for ligand-receptor interactions in biological systems.

A Polymer Mixing Approach to Chemoattractant-Stimulated Neutrophil Adhesion

We have used photometric and aqueous phase polymer affinity techniques to investigate the kinetics and surface thermodynamics of neutrophilic leukocyte responses to the soluble bacterial chemotaxin N-formyl methionyl leucyl phenylalanine (fMLP). Unlike the relatively slow responses of macrophages to particles, neutrophils respond to chemoattractants within seconds and display a complex series of time-dependent changes which in part reflect cellular adaptation to the chemotaxin by surface receptor internalisation [1,2]. The kinetics of these changes can be resolved by photometric methods, but not by the relatively slow techniques of macroscopic polymer phase wetting. We have therefore used the fungal metabolite cytochalasin B to prevent reversal of the rapid neutrophil responses to fMLP. Under these conditions, wetting of the neutrophils by dextran/PEG phases is essentially independent of time, and depends only on the dose of chemoattractant. The dose-response curves for fMLP induced cell aggregation and increased dextran phase affinity are similar, each with an ED50 of 2 x 10−8 M, suggesting that the two processes may have the same basis, for example in specific granule glycoproteins which become expressed on the cell surface during aggregation. By contrast, chemotactic cell surface ruffling (assessed either by digital image analysis of photomicrographs or by perpendicular light scattering from the cell suspensions) and oxygen radical-dependent chemiluminescence have dose-response curves with values of ED50 ten times lower and higher, respectively, than aggregation and phase affinity changes.

Macrophage Surface Affinity Changes During Phagocytosis

We have used a combination of surface thermodynamic, spectro-photometric and morphometric techniques to investigate the mechanism of particle ingestion (phagocytosis) by isolated alveolar macrophages. We studied the time and dose-dependence of the effects on macrophages of both structurally specific particles (serum complement-opsonised zymosan) and non-specific particles (non-opsonised silica). Particle exposure increased the cell affinity for the dextran phase of 4% Dextran 2M/4% PEG 20K phases in a dose-and opsonin-dependent manner which correlated well with biochemical and morphological assessment of cell activation. We do not yet know the structural basis of the cell surface changes which are responsible for the alterations in polymer phase affinity, but likely candidates are phagocytosis-induced changes in the amount, conformation or molecular weight of membrane glycoproteins.