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(Arteriosclerosis, Thrombosis, and Vascular Biology. 1996;16:64-71.)
© 1996 American Heart Association, Inc.

Articles

Adhesion of Blood Platelets Is Inhibited by VCL, a Recombinant Fragment (Leucine⁵⁰⁴ to Lysine⁷²⁸) of von Willebrand Factor

Jan J. Sixma; Martin J.W. IJsseldijk; George Hindriks; G. Henrita van Zanten; Marian Gorecki; Amos Panet; Leonard I. Garfinkel; Philip G. de Groot

From the Department of Haematology, University Hospital Utrecht (Netherlands) (J.J.S., M.J.W.I., G.H., G.H. van Z., P.G. de G.), and Bio-Technology General (Israel) Ltd, Kiryat Weizmann, Rehovot, Israel (M.G., A.P., L.I.G.).

Correspondence to Dr J.J. Sixma, Department of Haematology, University Hospital Utrecht, PO Box 85500, 3508 GA Utrecht, Netherlands.

Abstract VCL, fragment Leu⁵⁰⁴ to Lys⁷²⁸ of von Willebrand factor (vWF) expressed in Escherichia coli, contains the glycoprotein (GP) Ib–binding domain of vWF. This fragment inhibited ristocetin-induced platelet aggregation with an IC₅₀ of 0.2 µmol/L and botrocetin-induced aggregation with an IC₅₀ of 0.08 µmol/L. We studied the antiadhesive profile of VCL by adding it to blood that was circulated over various adhesive surfaces. VCL inhibited adhesion to endothelial cell matrix, which served as a model of the vessel wall. Maximal inhibition at a high shear rate of 1600 s^-1 was stronger (60%) than at a low shear rate of 300 s^-1 (40%). Half maximal inhibition was found to be 1.5 µmol/L at both shear rates. The role of various adhesive molecules was investigated in more detail by coating glass coverslips with collagen type I, laminin, fibronectin, or vWF. Fibrinogen was studied as well. Platelet adhesion to laminin and vWF was not inhibited by VCL. Adhesion to collagen, fibronectin, and fibrinogen was particularly inhibited at a high shear rate. VCL coated to a coverslip caused a concentration-dependent adhesion that was blocked by antibodies against GPIb, which block interaction with vWF. Binding studies showed a nonsaturable ristocetin binding of VCL to platelets that was blocked by vWF or inhibitory antibodies against GPIb. Binding to collagen was weak, and VCL did not inhibit binding of vWF at a 5000-fold excess. From these data, we conclude that VCL inhibits adhesion in all cases in which adhesion is vWF dependent by competing for vWF binding to activated GPIb. The lack of inhibition of adhesion to vWF as a single molecule may be explained by assuming that this adhesion is determined by interaction of nonactivated GPIb with vWF that has been changed in conformation by adsorption. Studies investigating thrombus formation on the connective tissue of an atherosclerotic plaque in a human coronary artery showed that VCL was able to partially prevent this thrombus formation. VCL may be of value in preventing adhesion and thrombus formation under conditions in which these processes are dependent on vWF.

Key Words: von Willebrand factor • blood platelets • adhesion

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