Adhesion of Blood Platelets Is Inhibited by VCL, a Recombinant Fragment (Leucine 504 to Lysine 728 ) of von Willebrand Factor

VCL, fragment Leu504 to Lys728 of von Willebrand factor (vWF) expressed inEscherichia 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.