NADH/NAD(P)H oxidase activity and expression.

(A) NADH/NAD(P)H oxidase activity in stimulated platelets. Addition of NADH led to a dose-dependent increase in O$2−$ production in lysates of platelets (n = 3). When platelets had been stimulated with collagen, it was further increased compared to lysates of control platelets. NAD(P)H as a cosubstrate caused less O$2−$ production (n = 3). (B) NAD(P)H oxidase subunit expression in platelets. Immunoblots of platelet protein expression (PLT) of the NAD(P)H oxidase subunits p47^phox and p67^phox compared to neutrophil protein (PMN) as a positive control. As described in “Materials and methods,” 40 μg of protein lysed was subjected to SDS–polyacrylamide-gel-electrophoresis, blotted on nitrocellulose membranes, and visualized using polyclonal antibodies against the indicated proteins.