Effects of leukocyte-derived cathepsin G on platelet membrane glycoprotein Ib-IX and IIb-IIIa complexes: a comparison with thrombin.

Cathepsin G is a serine, chymotrypsin-like protease released by activated polymorphonuclear leukocytes (PMN) that may act as a platelet agonist. The effect of this enzyme on platelet surface glycoproteins (Gp) Ib and IIb-IIIa was evaluated by means of a cytofluorimetric assay, using fluorescein isothiocyanate-labeled monoclonal antibodies (MoAbs) directed at the alpha chain of Gp Ib (SZ2), at Gp IX or at the complex Gp IIb-IIIa (P2), and the fibrinogen-receptor-specific MoAb PAC-1. In human washed platelets, cathepsin G increased the binding of P2 and PAC-1, decreased the binding of SZ2, but only slightly affected the binding of anti-Gp IX. SZ2 binding decrease was more rapid in cathepsin G- than in thrombin-stimulated platelets, whereas the increase of P2 and PAC-1 binding occurred to a comparable extent with either agonist. In paraformaldehyde (PFA)-fixed and energy-depleted platelets, no effect on either Gp Ib or Gp IIb-IIIa complex was observed with thrombin. At variance, cathepsin G was still able to reduce binding of SZ2, whereas increased binding of P2 or PAC-1 antibodies was not observed. Triton X-100 permeabilization of cathepsin G-treated, PFA-fixed platelets did not restore SZ2 binding at variance with thrombin. Moreover, platelet incubation with cathepsin G resulted in the loss of ristocetin-induced agglutination in the presence of the von Willebrand factor and in the appearance of Gp Ib-derived proteolytic products in supernatants. After dissociation by EDTA pretreatment of surface Gp IIb-IIIa complexes, cathepsin G still induced increased binding of P2. Aspirin and an adenosine diphosphate scavenger system had only a slight but not significant effect on changes in antibody binding induced by cathepsin G. All these data would indicate that cathepsin G, like thrombin, interacts with platelet-surface Gp, inducing the exposure of the intracellular pool of the Gp IIb-IIIa complex with concomitant expression of a functional fibrinogen receptor. Moreover, it induces a loss of antigenic sites on Gp Ib, but the mechanism involved, a proteolytic cleavage of Gp Ib, is substantially different from that of thrombin. These changes, induced by a product of activated PMN, might reduce the reactivity of platelets to the subendothelium, while increasing their ability to undergo aggregation and release reaction.