Reduced Nicotinamide Adenine Dinucleotide Phosphate Inhibits Platelet Function

Aim: To investigate the effects of reduced nicotinamide adenine dinucleotide phosphate(NADPH)on platelet function and its potential mechanisms.Methods: In vitro studies,the effects of different concentrations of NADPH on platelet aggregation induced by ADP(10 μM),thrombin(0.05 U/m L)or AA(0.5 m M)were determined and the inhibitory effects of NAD+,NADH,NADP+ and NADPH on platelet aggregation were compared.By examining the concentration of NADPH in platelets,whether NADPH could enter into platelets was explored.In vivo studies,the effects of NADPH on tail bleeding time,clotting time and ferric chloride-induced thrombosis model were determined in mice or rats.The p38 MAPK activator anisomycin and inhibitor SB203580 were utilized to explore whether NADPH inhibiting platelet aggregation was related to the phosphorylation of p38.Results: NADPH could concentration-dependently inhibit platelet aggregation induced by ADP(10 μM)or thrombin(0.05 U/m L),but not AA(0.5 m M).NADPH might not have enough time to enter into platelets and mainly play its role outside platelets.When the inhibitory effects of NAD+,NADH,NADP+ and NADPH on platelet aggregation were compared,NADPH demonstrated the best inhibitory effect.NADPH transiently prolonged the tail bleeding time in mice at 30 min after the injection of NADPH(7.5 mg/kg),while aspirin(15 mg/kg)significantly prolonged the tail bleeding time in mice at all time points examined.As the effects of NADPH or aspirin on clotting time in rats were detected,both of them had no effect.In Fe Cl3-induced abdominal aorta injury thrombosis model,administration of NADPH significantly delayed the onset of vessel occlusion,while aspirin almost completely prevented the vessel occlusion.With microscope examination the thrombi in injured vessel sections of rats received NADPH were much smaller and looser than rats received vehicle treatment and the thrombi in injured vessel sections of rats given aspirin were fewest.Furthermore,both ADP and thrombin induced phosphorylation of p38 and the effect was markedly inhibited by NADPH.The inhibitory effects of NADPH on platelet aggregation were mimicked by the p38 inhibitor SB 203580 and were slightly antagonized by the p38 activator anisomycin.Conclusions: The current results demonstrated that NADPH inhibited platelet aggregation and its mechanism might relate to p38 MAPK pathway.NADPH could delay the onset of vessel occlusion to some extent and had mild effects on bleeding time and clotting time.These suggested that NADPH might be a novel compound for management of high risk of cardiovascular disease.To our knowledge,this is the first study on the effect of NADPH on platelet function.