PORCINE VON-WILLEBRAND-FACTOR BINDING TO HUMAN PLATELET GPIB INDUCES TRANSMEMBRANE CALCIUM INFLUX

Porcine von Willebrand factor (P-vWF) binds to human platelet glycopro tein (GP) Ib and, upon stirring (1500 rpm/min) at 37 degrees C, induce s, in a dose-dependent manner, a transmembrane flux of Ca2+ ions and p latelet aggregation with an increase in their intracellular concentrat ion. The inhibition of P-VWF binding to GP Ib, obtained with anti GP I b monoclonal antibody (LJ-Ib1), inhibits the increase of intracellular Ca2+ concentration ([Ca2+]i) and platelet aggregation. This effect is not observed with LJ-Ib10, an anti GP Ib monoclonal antibody which do es not inhibit the VWF binding to GP Ib. An anti GP IIb-IIIa monoclona l antibody (LJ-CP8) shown to inhibit the binding of both vWF and fibri nogen to the GP IIb-IIIa complex, had only a slight effect on the [Ca2 +]i rise elicited by the addition of P-vWF. No inhibition was also obs erved with a different anti GP IIb-IIIa monoclonal antibody (LJ-PS), s hown to block the binding of vWF and not that of fibrinogen to the GP IIb-IIIa complex. PGE(1), apyrase and indomethacin show a minimal effe ct on [Ca2+]i rise, while EGTA completely blocks it. The GP Ib occupan cy by recombinant VWF fragment rvWF(445-733) completely inhibits the i ncrease of [Ca2+]i and large aggregates formation. Our results suggest that, in analogy to what is seen with human vWF under high shear stre ss, the binding of P-VWF to platelet GP Ib, al low shear stress and th rough the formation of aggregates of an appropriate size, induces a tr ansmembrane flux of Ca2+, independently from platelet cyclooxygenase m etabolism, perhaps through a receptor dependent calcium channel. The i ncrease in [Ca2+]i may act as an intracellular message and cause the a ctivation of the GP IIb-IIIa complex.