SHEAR STRESS ENHANCED SOLUTE TRANSPORT ACROSS MEMBRANES

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The proposed research plan is focused on the study of calcium ion influx through cellular membranes as a function of fluid shear stress. Blood platelets and endothelial cells will be studied independently to determine the impact of shear stress-enhanced transmembrane calcium ion flux on intracellular free calcium ion concentration. The effect of shear stress on the diffusional and convective transport of other biologically important substances will also be examined. Insight into the initiating steps of fluid mechanical cellular activation will be sought. The influence of elongational stress on fluid dynamic platelet activation will be evaluated. Measurements will be made continuously with a cone and plate viscometer, a parallel plate flow chamber or an 'elongational flow rheometer'. each modified for fluorescence and luminescence measurement. Intracellular calcium ion concentration, intracellular pH, membrane potential and platelet dense granule release will be monitored continuously during fluid stress exposure. The rotational chamber geometry permits application of a unifom, well-described shear field to the entire sample volume. The parallel plate apparatus can apply well-described shear stress to anchorage-dependent cells attached to one wall of the flow chamber. The 'elongational flow rheometer' produces a flow of uniform, constant acceleration which results in the application of a constant fluid dynamic stretching force. Proposed studies will utilize unilamellar liposomes as model cell membranes. Use of liposomes will decouple the calcium ion flux measurement in a clear and unambiguous way from the normal cellular processes of calcium ion homeostasis. Tbe role of platelet membrane proteins in the management of calcium ion flux will be investigated through preparation of liposomes from isolated platelet membranes. Tbe effect of cardiovascular fluid stress on the efflux kinetics of pharmaceuticals from liposomes will also be studied. The results obtained from experiment will be incorporated into a mathematical model of shear stress enhanced transport through cell membranes. A predictive scheme will be developed to estimate the degree of shear stress induced transport as a function of physical system parameters and fluid dynamic exposure.