Inhibition of adhesive and signaling functions of the platelet GPIb-V-IX complex by a cell penetrating GPIbα peptide

Abstract

Background: Interaction between the platelet glycoprotein (GP)Ib-V-IX complex and von Willebrand factor (VWF) is critical for initiating platelet-vessel wall contacts, particularly under high shear conditions. This interaction also plays an important role in initiating platelet activation through the generation of intracellular signals resulting in platelet shape change and integrin αIIbβ3 activation. Objective: A cell-penetrating peptide strategy as used to study the role of the intracellular domain of the GPIb α subunit in VWF/GPIb-V-IX-dependent adhesion and activation. Methods: Peptides of 11-13 amino acids, covering the 557-610 region, were coupled to a nine-arginine permeating tag (R9) and the effects of their cell entry on VWF-dependent responses were analyzed. Results: The R9α 557 peptide corresponding to the 557-569 segment reduced platelet agglutination in response to VWF, while the other peptides had no effect. The decreased platelet agglutination appeared to be an indirect consequence of adenosine diphosphate release as a normal response was restored by apyrase or a P2Y1 receptor antagonist. A more direct effect of R9α557 on GPIb VWF-dependent functions was observed in adhesion studies on a VWF matrix, where it decreased platelet adhesion and profoundly inhibited filopodia formation. In addition, cell adhesion was reduced and shape change absent when Chinese hamster ovary cells expressing the GPIb-IX complex were incubated with R9α557. Conclusion: This study performed in intact platelets suggests a functional role of the 557-569 domain of GPIbα in controlling VWF-dependent adhesion and signaling. © 2006 International Society on Thrombosis and Haemostasis.