Contact-mediated triggering of lamella formation by Dictyostelium amoebae on solid surfaces.

Amoebae of the slime mould Dictyostelium discoideum form broad ultrathin cytoplasmic lamellae by a centripetal contractile process soon after they have spread on certain solid surfaces. We have investigated the surface requirements for initial triggering of this contact-mediated signalling system. The lamellar response is not normally evoked by glass, but is seen on glass covalently derivatized with paraffinic chains, as well as on glass covalently derivatized with amine groups and on glass bearing adsorbed polylysine. We have recorded the frequency of the lamellar response on these surfaces as a function of ionic strength and pH, and have measured the electrostatic potentials of the surfaces by the streaming potential method. Using these data we have concluded that the general trigger for the lamellar response is not a 'simple' physical or chemical property of the substrata: it is not dependent on specific chemical groups, degree of hydrophobicity, electrostatic potential, or charge density, taken as isolated factors. It seems likely that triggering is dependent on the overall energetics of cell-substratum interaction.

Cell adhesion to hydroxyl groups of a monolayer film.

We have studied cells on chemically defined monomolecular films of the long-chain alcohol docosanol. Langmuir-Blodgett films of the alcohol were deposited on glass coverslips, previously made hydrophobic with octadecyl groups. This gives films in which the alcohol headgroups face outwards to the water. Molecular orientation and film integrity were shown by a fluorescence adsorption test. Cell contacts on the films were observed in media without proteins by interference reflection microscopy (IRM) and the mechanics of detachment were examined by hydrodynamic shearing in a flow chamber. Cell contact with docosanol was compared with that on an adjacent area of octadecyl glass without a monolayer. Dictyostelium amoebae settled and spread on both docosanol and octadecyl glass, but little or no locomotion was seen on docosanol. On octadecyl glass the amoebae moved actively, forming ultrathin cytoplasmic lamellae, which look dark under IRM, and left distinctive trails of membranous debris. Hydrodynamic shearing showed that the amoebae stuck strongly to both surfaces and could not be removed from either at the maximum attainable wall shear stress of 6Nm-2. Red blood cells also adhered to both surfaces and removal from both occurred between 1 and 3Nm-2. IRM and scanning electron microscopy (SEM) studies indicated that this force leads to a minimal measure of red cell adhesion, since removal often involved the breakage of cytoplasmic tethers. Our results show that alcoholic -OH groups, in a two-dimensional array, provide a surface that is strongly adhesive for cells. No other method has made it possible to demonstrate cell adhesion purely to -OH groups, in a known orientation and density, and in the absence of any other functional groups on the interface.

Inhibition of cell adhesion by a synthetic polymer adsorbed to glass shown under defined hydrodynamic stress.

A co-polymer with hydrophobic and hydrophilic segments was allowed to adsorb from aqueous solution onto glass previously made hydrophobic by derivatization with octadecyl dimethylchlorosilane. The polymer is thought to adsorb via its hydrophobic segments, leaving the hydrophilic segments free to extend into the water. After allowing cells to settle on the treated surface, the shear stress at the chamber wall required to remove red blood cells, Dictyostelium discoideum amoebae and Escherichia coli was determined in a calibrated laminar flow chamber. On octadecyl glass a shear stress of 2-3 Nm-2 evicts 50% of adherent red cells and E. coli. No D. discoideum amoebae could be removed at 5Nm-2. In striking contrast, the lowest experimentally obtainable shear stress of 0.03 Nm-2 removes 97.0-99.5% of cells of all three types from the polymer-treated surface, even after a cell residence time of 1 h without flow in the absence of free polymer. The minimum shear stress of 0.03Nm-2 corresponds to only approximately equal to 20 times the force of gravity on a red cell. The mechanism of action of the polymer and the implications of the results are discussed.

Photosynthetic electron carriers at a heptane-water interface.

Surface properties of monomolecular films of chlorophyll (Chl), plastocyanin (Pc), cytochrome c (Cyt) and ferridoxin (Fd) were measured at a heptane-water interface. Mixed films of Chl and the other components were examined in darkness and in light. The area/molecule, A, for Fd (from Cl. pasteurianum) is much larger at a heptane-water than Fd (from spinach) at an air-water interface. This difference in A may be the source of the Fd or the extent of denaturation at the different interfaces. There appears to be a photoreaction between Chl and Fd in the presence of ascorbate. The A for Cyt is much larger at a heptane-water than at an air-water interface. In mixed films there is a strong interaction between Chl and reduced Cyt in the dark. No such interaction is observed between Chl and oxidized Cyt. With mixed films of Chl and reduced Pc there is a significant, reversible, light induced change in deltaV.