title: Shear stress-induced binding of von Willebrand factor to platelets.
abstract: Shear stress-induced platelet aggregation requires von Willebrand factor (vWF), platelet glycoprotein (GP) Ib, GPIIb-IIIa, 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 GPIb and GPIIb-IIIa. The sequence of the vWF finding 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/microliters 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 alpha-granule vWF) demonstrated that exogenous vWF predominately contributed to the platelet-vWF binding. Blockade of platelet GPIb with the monoclonal anti-GPIb antibody, 6D1, 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.
authors.affiliation: Cox Lab for Biomedical Engineering, Rice University, Houston, Texas 77251-1892, USA. konstant@biomed.rice.edu
authors.forename: K
authors.initials: K
authors.lastname: Konstantopoulos
authors.forename: T W
authors.initials: TW
authors.lastname: Chow
authors.forename: N A
authors.initials: NA
authors.lastname: Turner
authors.forename: J D
authors.initials: JD
authors.lastname: Hellums
authors.forename: J L
authors.initials: JL
authors.lastname: Moake
mesh_terms: D001792:Blood Platelets / Q000378:metabolism*; D005434:Flow Cytometry; D018056:Hemorheology; D006801:Humans; D010974:Platelet Aggregation; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000378:metabolism; D013314:Stress, Mechanical; D014841:von Willebrand Factor / Q000378:metabolism*
chemical_list: D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D019038:Platelet Glycoprotein GPIb-IX Complex; D014841:von Willebrand Factor