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Surface chemical studies of human platelets Chiu, Basil


The purpose of this project is to investigate the surface properties of platelet discocytes, echinocytes and spherocytes. Normal "non-sticky" discoid shaped platelets (discocytes) can be transformed by ADP into irregularly shaped echinocytes which are "sticky" and aggregate easily in media containing Ca⁺⁺ and fibrinogen. A model is examined here in which an echinocyte attains its "sticky" properties by evagination of a surface-connected canalicular system. Platelets also evaginate this canalicular system upon hypotonic shock, in which case the platelets swell up to form spherocytes. By comparing the properties of the different geometric forms of platelets insight into the nature of "stickiness" was sought. The surface areas of the discocyte and spherocyte measured microscopically were found to be 16.4 and 36.7x10⁻⁸ cm² respectively while that of the echinocyte was estimated to be 23.7x10⁻⁸ cm² using surface chemical analysis. Electron microscopic examination showed that the canalicular system may not be totally evaginated in the echinocyte. Although it was found that the spherocyte could still be agglutinated passively by ristocetin it had completely lost its ability to aggregate. Microelectrophoretic studies revealed 8 and 6 fold increases in the density of Ca⁺⁺ and Mg⁺⁺ binding sites respectively on the echinocyte surface relative to the discocyte. The spherocyte on the other hand had lost most of its Ca binding sites. Electrokinetic analysis of live, fixed and neuraminidase or alkaline phosphatase treated platelets showed major differences in charge as well as amino, sialic acid and phosphate group densities among the discocyte, echinocyte and spherocyte. The evaginated canalicular membrane surfaces of the latter two were also different. SDS-PAGE of platelets radiolabelled via lactoperoxidase iodination, periodate-borohydride tritiation or neuraminidase/galactose oxidase-borohydride tritiation failed to show any difference in the gel patterns between the three platelet forms. No new glycoprotein species appeared during the transformations. The presence of fibrinogen interferes in a concentration related manner with lactoperoxidase iodination of GP-III on the echinocyte surface. An overall picture is presented here showing differences between the surface properties of platelet discocytes, echinocytes and spherocytes. The accumulated evidence suggests that changes in the whole platelet surface occur during activation and the model of a cloistered "sticky" membrane may be an oversimplification.

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