Affiliations: [a] Cox Lab for Biomedical Engineering, Rice University, Houston, Texas, USA | [b] Department of Medicine, Baylor College of Medicine, Houston, Texas, USA
Note: [*] Reprint requests to: Joel L. Moake, M.D., Cox Laboratory for Biomedical Engineering, Rice University, Houston, TX 77251-1892, USA; Fax: (713) 285-5353; E-mail: [email protected]
Abstract: Shear stress-induced platelet aggregation requires von Willebrand factor (vWF), platelet glycoprotein (GP) Ib, GPlIb-lIla, Ca2+, and adenosine diphosphate (ADP). Recent reports using vWF labeled with either 125I or fluorescein isothiocyanate (FITC) have demonstrated that in shear-fields, vWF binds to both GPlb and GPIIb-IIIa. The sequence of the vWF binding to the two platelet receptors has not been precisely determined in these reports. In this study, a flow cytometry technique using a primary anti-vWF antibody and a secondary FITC IgG antibody was used to measure shear stress-induced vWF binding to platelets. Washed normal platelets suspended at 50,000/µl with purified large vWF multimers were exposed to laminar shear stresses of 15 to 120 dynes/cm2 for 30 sec. At this low platelet count, little or no aggregation occurred in the shear fields. A significant increase in post-shear vWF-positive platelets was consistently observed. Experiments with platelets from normal and severe von Willebrand’s disease (vWD) (which lack plasma and platelet α-granule vWF) demonstrated that exogenous vWF predominately contributed to the platelet-vWF binding. Blockade of platelet GPlb with the monoclonal anti-GPlb antibody, 6Dl, completely inhibited shear stress-induced platelet-vWF attachment. In contrast, blockade of GPIIb-IIIa with monoclonal anti-GPIIb-IIIa antibodies, 10E5 or c7E3, or with the GPIIb-IIIa-blocking tetrapeptide, RGDS, had little or no inhibitory effect on platelet-vWF binding. These data demonstrate that the binding of vWF to GPIb is likely to be the initial shear-induced platelet-ligand binding event.
Keywords: von Willebrand factor, glycoprotein (GP) Ib, GPIIb-IIla, shear stress
DOI: 10.3233/BIR-1997-34104
Journal: Biorheology, vol. 34, no. 1, pp. 57-71, 1997
