Inherited human gp91phox deficiency is associated with impaired isoprostane formation and platelet dysfunction.
OBJECTIVE
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.


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.


CONCLUSIONS
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.
Department of Internal Medicine and Medical Specialties, I Clinica Medica, University of Rome La Sapienza, Viale del Policlinico 155, Rome, Italy.
Pasquale
P
Pignatelli
Roberto
R
Carnevale
Serena
S
Di Santo
Simona
S
Bartimoccia
Valerio
V
Sanguigni
Luisa
L
Lenti
Andrea
A
Finocchi
Loredana
L
Mendolicchio
Anna Rosa
AR
Soresina
Alessandro
A
Plebani
Francesco
F
Violi
D000328:Adult; D001241:Aspirin / Q000494:pharmacology; D001792:Blood Platelets / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D016022:Case-Control Studies; D003677:Deficiency Diseases / Q000378:metabolism* / Q000473:pathology; D015237:Dinoprost / Q000031:analogs & derivatives / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D028421:Isoprostanes / Q000378:metabolism*; D008297:Male; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000172:deficiency* / Q000235:genetics; D009249:NADP / Q000378:metabolism; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000172:deficiency* / Q000235:genetics; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism
D004791:Enzyme Inhibitors; D028421:Isoprostanes; D008562:Membrane Glycoproteins; D010975:Platelet Aggregation Inhibitors; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; C075750:8-epi-prostaglandin F2alpha; D009249:NADP; D015237:Dinoprost; D019001:Nitric Oxide Synthase; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D001241:Aspirin; D002118:Calcium; D019331:NG-Nitroarginine Methyl Ester