Inherited Human gp91 phox Deficiency Is Associated With Impaired Isoprostane Formation and Platelet Dysfunction

Abstract

Object— Platelet isoprostane 8-ISO-prostaglandin F2α (8-iso-PGF2α), a proaggregating molecule, is believed to derive from nonenzymatic oxidation of arachidonic acid. We hypothesized that NADPH is implicated in isoprostane formation and platelet activation. Methods and Results— We studied 8-iso-PGF2α in platelets from 8 male patients with hereditary deficiency of gp91 phox , the catalytic subunit of NADPH oxidase, and 8 male controls. On stimulation, platelets from controls produced 8-iso-PGF2α, which was inhibited −8% by aspirin and −58% by a specific inhibitor of gp91 phox . Platelets from patients with gp91 phox hereditary deficiency had normal thromboxane A 2 formation but marked 8-iso-PGF2α reduction compared with controls. In normal platelets incubated with a gp91 phox inhibitor or with SQ29548, a thromboxane A 2 /isoprostane receptor inhibitor, platelet recruitment, an in vitro model of thrombus growth, was reduced by 44% and 64%, respectively; a lower effect (−17%) was seen with aspirin. Moreover, thrombus formation under shear stress (blood perfusion at the wall shear rate of 1500 s −1 ) was reduced in samples in which isoprostane formation was inhibited by NADPH oxidase inhibitors. In gp91 phox -deficient patients, agonist-induced platelet aggregation was within the normal range, whereas platelet recruitment was reduced compared with controls. Incubation of platelets from gp91 phox -deficient patients with 8-iso-PGF2α dose-dependently (1 to 100 pmol/L) increased platelet recruitment by mobilizing platelet Ca 2+ and activating gpIIb/IIIa; a further increase in platelet recruitment was detected by platelet coincubation with l -NAME, an inhibitor of NO synthase. Conclusion— This study provides the first evidence that platelet 8-iso-PGF2α maximally derives from gp91 phox activation and contributes to platelet recruitment via activation of gpIIb/IIIa.