Thrombin facilitates primary platelet adhesion onto vascular surfaces in the absence of plasma adhesive proteins: studies under flow conditions

Background and Objectives. The effect of local and circulating thrombin on platelet adhesion onto vascular surfaces was explored in the absence of pla sma adhesive proteins using flow conditions. Design and Methods. To study the local effects of thrombin, denuded rabbit aorta segments were incubated with thrombin concentrations of 0,001, 0.01 a nd 0.1 U/mL. To evaluate the effects of circulating thrombin, the same conc entrations were added to perfusates consisting of washed platelets and wash ed red blood cells suspended in a human albumin solution (5%). In some expe riments, purified von Willebrand's factor (VWF) (Haemate-P) was added to th e perfusates (0.8 U/mL of vWF, final concentration). A humanized chimeric a ntibody to the GPI-Ib-IIIa complex (Reopro) was used to determine the role of this glycoprotein on platelet adhesion under the conditions described. T he effect of blocking GPIb was also assessed. Perfusions were carried out a t 800 s(-1) for 10 min, The interaction of platelet with the vessel surface was morphometrically evaluated and expressed as percentage of surface cove rage (%SC). Changes in the surface expression of the major platelet antigen s were also analyzed by flow cytometry, Results. Incubation of subendothelial surfaces with thrombin enhanced plate let deposition with respect to control levels (increases in SC of 64%, 79% and 86% with 0,001, 0.01 and 0.1 U/mL of thrombln, respectively). Low conce ntrations of thrombin (0.001 and 0.01 U/mL) incorporated In the perfusates resulted In a similar pro-adhesive effect (Increases in SC of 64% and 71%, respectively) while the highest concentration (0.1 U/mL) failed to produce a pro-adhesive effect due to the augmented formation of platelet aggregates with subsequent thrombocytopenia (15+/-1 vs. 160+/-5 x10(9) plt/L in the p erfusates), Similar results were obtained when VWF was present in the perfu sate, Reduction of platelet deposition by blockade of GPIIb-IIIa (to 5.3+/- 0.7%) was partially restored by thrombin, Blockade of GPlb prevented platel ets from adhering even when thrombin was present (%SC of 2,0+/-0,8%), No si gnificant changes in the distribution of platelet membrane glycoproteins du ring perfusion experiments were detected. Interpretation and Conclusions. Our results suggest that thrombin facilitat es primary platelet adhesion onto vascular surfaces even in the absence of plasma adhesive proteins. This effect seems to be mainly dependent on the G PIb/vWF axis. (C) 2000, Ferrata Storti Foundation.