Novel role for low molecular weight plasma thiols in nitric oxide-mediated control of platelet function

Abstract

Nitric oxide (NO) is a powerful antiplatelet agent, but its notoriously short biological half-life limits its potential to prevent the activation of circulating platelets. Here we used diethylamine diazeniumdiolate (DEA/NO) as an NO generator to determine whether the antiplatelet effects of NO are prolonged by the formation of a durable, plasma-borne S-nitrosothiol reservoir. Preincubation of both platelet rich plasma (PRP) and washed platelets (WP) with DEA/NO (2 μM) for 1 min inhibited collagen-induced platelet aggregation by 82 ± 5 and 91 ± 2%, respectively. After 30 min preincubation with DEA/ NO, NO was no longer detectable in either preparation, but aggregation remained markedly inhibited (72 ± 7%) in PRP. In contrast, the inhibitory effect in WP was almost completely lost at this time (5 ± 3%) but was partially restored (39 ± 10%) in WP containing human serum albumin (1%) and fully restored by co-incubation with albumin and the low molecular weight (LMW) thiols, glutathione, (5 μM), cysteinyl-glycine (10 μM), or cysteine (10 μM). This NO-mediated effect was not seen with LMW thiols in the absence of albumin and was associated with S-nitrosothiol formation. Our results demonstrate that LMW thiols play an important role in both the formation and activation of an S-nitrosoalbumin reservoir that significantly prolongs the duration of action of NO.