Abstract

In this study, we evaluated the influence of antithrombin on platelet adhesion onto immobilized fibrinogen using an in vitro system simulating venous and arterial flow. Platelets in anticoagulated whole blood (PPACK, 40μM) were labeled with mepacrine (10μM). Adhesion of platelets onto fibrinogen-coated glass cover slips was assessed in a rectangular flow chamber (shear rates of 13 s−1 to 1500 s−1). Platelets were visualized at 15 sec, 1 and 5 min following perfusion using a fluorescence laser-scan microscope. In parallel, the effects of supraphysiological supplementation of blood with antithrombin (2.8 IU/ml of blood) on platelet adhesion rates was evaluated. During perfusion, platelet adhesion onto fibrinogen linearly increased with exposure time and shear rates. Within the first min of perfusion, an inverse correlation between platelet adhesion and plasma antithrombin activity was observed at shear rates of 13 s−1 and 50 s−1 (r=−0.48 and r=−0.7, p each <0.05). Significant differences in platelet adhesion (1786±516 U vs. 823±331 U, p<0.05) related to low (92±3.3%) and high (117±4.1%) antithrombin activity was observed at a flow rate of 13s−1 within first minute. Further supplementation of anticoagulated whole blood with antithrombin (activity up to 280 %) decreased the rate of platelets adhesion (ratio of adhesion at 1 and 5 min) about 35% when compared to nonsupplemented blood (1.25 ± 0.17 vs. 1.95 ± 0.4, p=0.008). Application of heparine as anticoagulant did not enhance the antiadhesion properties of antithrombin. Our findings are in accordance with the “low shear phenomenon” of arterial thrombus progression, i.e. thrombus enlargement at distal areas with reduced flow or even stasis. Moreover, the observation that antithrombin significantly suppressed platelet adhesion onto immobilized fibrinogen under low flow conditions is of therapeutic interest and needs further evaluation.

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