The Anti-aggregating Peptide KRDS Impairs a-granule Release, Whereas RGDS Does Not.
The effects on platelet activation of two different tetrapeptides, KRDS present on human lactotransferrin and RGDS present on adhesive proteins such as human fibrinogen α-chain, were compared by a combination of morphological and functional techniques. Ultrastructural observations of α-thrombin stimulated platelets (0.05 U/ml), show strong platelet aggregation and full α-granule release. In the presence of RGDS (0.1-1 mM) aggregation was impaired but secretion was not blocked and platelets had released their α-granule contents. Platelets appeared uniformly degranulated with a dense central meshwork of microfilaments. In the presence of KRDS (0.5-1 mM), the platelets were activated with shape change and pseudopod formation. Aggregation was also impaired, but to a lesser extent since RGDS is active at a concentration as low as 0.1 mM, and, in contrast to RGDS, secretion was severely reduced. Electron microscopy showed that numerous α-granules were still scattered in the cytoplasmic matrix or often gathered in the centre of the platelet, but the majority of the open canalicular system cisternae remained clear. An immunoelectron microscopic study using immunogold and monospecific antibodies directed against fibrinogen and the a-granule membrane protein P-selectin (GMP 140) was performed. In the presence of RGDS, fibrinogen was released and P-selectin was translocated to the platelet surface; in contrast, in the presence of KRDS, fibrinogen remained localized in the α-granule, and the P-selectin associated with the a-granule. These observations were accompanied by some functional results: thrombin-induced platelet aggregation was inhibited by both peptides, and in contrast to RGDS, secretion was severely reduced in the presence of KRDS: serotonin release from dense granule was reduced by 73% compared to the control. These results show that these two tetrapeptides, in spite of some structural similarities, act differently in impairing platelet function. KRDS interfering with both the dense and α-granule release reaction may be a useful tool for a better understanding of the platelet secretion mechanism.
Institut des Vaisseaux et du Sang, Service d'Hématologie and INSERM U348, Hopital Lariboisiere, 8 rue Guy Patin, Paris, 75010, France.
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Mazoyer
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JP
Caen
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Tenza
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EM
Cramer