The Central hypothesis to be tested in the proposed research is that the differences in blood reactivity among materials arise primarily from the propensity of each material to cause the surface concentration and surface induced degree of transition of adsorbed, platelet adhesive proteins to vary. Testing this hypothesis will lead to a fundamental understanding of certain primary events occurring upon exposure of foreign materials to blood, and thus provide an improved basis for the development of materials with increased blood compatibility.
The specific aims of the proposed research are: 1) The transitions that occur in proteins after their adsorption to polymeric substrates will be studied with series of proteins, surfaces and techniques in order to establish the generality, nature, and biological significance of these transitions. The amount of adsorption of these proteins from plasma and blood will also be measured. Proteins to be studied include fibrinogen, fibronectin, and vitronectin. The techniques to be used include detergent elutability, antibody binding, displaceability with other proteins, and Fourier transform infrared (FTIR) spectroscopy. Studies will be conducted on five separate, well characterized series of materials (polyurethanes; NHLBI reference materials; hydroxyethylmethacrylate-ethylmethacrylate (HEMA-EMA) copolymers; and polyethylene oxides(PEO)). 2) The effect of the protein transitions on platelet interactions with the same series of polymers will be characterized. We will use an epifluorescent video microscopy(EVM) system to monitor platelet interactions with surfaces in situ continuously using flowing platelet/red cell suspensions and whole blood. In addition, quantitative adhesion measurements will be done with 111 In labeled platelets. Platelet activation will also be characterized using morphology, granule release, an antibody specific to activated platelets, and by measuring fluorescence generation due to intracellular calcium mobilization. Flow cytometry as well as in situ EVM examination will be developed to assess platelet activation in situ and in non-adherent platelets. 3) The role of specific platelet receptors in adhesion to these materials will be studied with a series of antibodies to several of the platelet receptors in order to delineate the mechanisms by which platelets become adherent to biomaterials surfaces.
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