Thrombin Receptor Occupancy Modulates Aggregation Efficiency and Platelet Surface Expression of vWF and Thrombospondin at Low Thrombin Concentrations

Ana Kasirer-Friede; Chantal Legrand; Mony M. Frojmovic

doi:10.1055/s-0037-1614608

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 1999; 81(06): 967-975
DOI: 10.1055/s-0037-1614608

Letters to the Editor

Schattauer GmbH

Thrombin Receptor Occupancy Modulates Aggregation Efficiency and Platelet Surface Expression of vWF and Thrombospondin at Low Thrombin Concentrations

Ana Kasirer-Friede

²From the Depts of Experimental Medicine and Physiology, McGill University, Montreal, Canada

,

Chantal Legrand

¹Unité 353 INSERM, Hôpital St. Louis, Paris, France

,

Mony M. Frojmovic

²From the Depts of Experimental Medicine and Physiology, McGill University, Montreal, Canada

› Author Affiliations

Abstract

Summary

Previous studies evaluating requirements for occupancy of thrombin receptors in normal platelet secretion and aggregation, using the thrombin antagonists hirudin and PPACK (D-Phe-Pro-Arg-chloromethylke-tone), have suggested that at low thrombin activating concentrations (0.025–0.13 U/ml), occupancy was required only in the first 45–60 s following activation. In our study, we differentiate between thrombin receptor occupancy requirements for surface expression of secreted adhesive proteins, for activation of GPIIb-IIIa receptors, and for aggregation of washed platelets (WP) in laminar shear flow. Platelets activated with 0.05 U/ml thrombin for 10 min to allow maximal secretion (hereafter referred to as “pre-activated platelets”), then sheared, showed a 50–70% decrease in platelet counts after 60 s of shear. Treatment of pre-activated platelets with hirudin or PPACK produced a 65% reduction of capture efficiencies, α_G (reflecting experimental/theoretical initial rates of aggregation), as well as a 30–40% decrease in the surface expression of von Willebrand factor (vWF) and thrombospondin (TSP). However, α-granule membrane P-selectin expression and numbers of activated GPIIb-IIIa receptors were comparable for treated and non-treated platelets. No significant difference in any of the parameters tested was observed when platelets were similarly pre-activated with 0.2 U/ml thrombin, due to treatment with thrombin antagonists. Binding of soluble FITC-vWF (GRGDSP-sensitive) to pre-activated, thrombin antagonist treated platelets, was greatly reduced (≥80%). Soluble Fg was shown to bind to antagonist-treated pre-activated platelets, but could not significantly enhance platelet aggregation. Although occupancy of thrombin receptors by catalytically active thrombin is required transiently for secretion and activation of platelets, there is a further requirement for thrombin occupancy at low thrombin concentrations, for optimizing initial rates of platelet aggregation, surface expression of vWF and TSP, and activated GPIIb-IIIa ligand recognition.

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References

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