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

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 DEAINO (2 muM) 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 MTP 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 (LAW) thiols, glutathione, (5 muM), cysteinyl-glycine (10 muM), or cysteine (10 muM). 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 LAW 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.