Antiplatelet properties of protein S-nitrosothiols derived from nitric oxide and endothelium-derived relaxing factor.

Abstract

5-Nitrosothiols may serve as carriers in the mechanism of action of endothelium-derived relaxing factor (EDRF) by stabilizing the labile nitric oxide (NO) radical from inactivation by reactive species in the physiological milieu and by delivering NO to the heme activator site of guanylyl cyclase. Low-molecularweight thiols, such as cysteine and glutathione, form 5-nitrosothiol adducts with vasodilatory and antiplatelet properties, and protein thiols can interact in the presence of NO and/or EDRF to form uniquely stable 5-nitroso-proteins. We now show that the 5-nitroso-proteins, 5-nitroso-albumin, 5-nitroso-tissue type plasminogen activator, and 5-nitroso-cathepsin B, have potent antiplatelet effects with an ICso of approximately 1.5 fiM. In the dog, 5-nitroso-albumin inhibits ex vivo platelet aggregation and significantly prolongs the template bleeding time from 2.15±0.13 (mean±SEM) to 9.70±1.24 minutes. The antiplatelet action of 5-nitroso-proteins is associated with the stimulation of guanylyl cyclase and a significant decrease in flbrinogen binding to platelets. 5-Nitroso-proteins undergo thiol-nitrosothiol exchange with low-molecular-weight thiols to form low-molecular-weight 5-nitroso-thiols, and they also interact directly with the platelet surface, both of which processes facilitate generation of NO. These data suggest that 5-nitroso-proteins are potent antiplatelet agents and may be intermediates in the antiplatelet mechanism of EDRF action. {Arteriosclerosis and Thrombosis 1993;13:791-799) KEY WORDS • 5-nitrosothiols • thionitrites • cyclic GMP • thiols • sulfhydryl groups

Evidence for S -nitrosothiol-dependent changes in fibrinogen that do not involve transnitrosation or thiolation

“…There have been several reports on RSNO-dependent inhibition of blood clot formation (18) and platelet aggregation (20)(21)(22)(23). In all of these cases, the mechanism of inhibition has been attributed to either the NO-mimicking activity of RSNOs in triggering the NO͞cGMP pathway or has involved the transnitrosation or S-thiolation of an essential free thiol(s) on the proteins participating in thrombosis or platelet activation.…”

Kinetics and equilibria of S‐nitrosothiol—thiol exchange between glutathione, cysteine, penicillamines and serum albumin

“…S-Nitrosothiols (RSNO) can elicit many of the various physiological regulatory functions attributed to nitric oxide/endothelium-derived relaxing factor, including smooth muscle relaxation [l-4], platelet deactivation [5,6], immunosupression [7] and neurotransmission [8]. RSNO occur in human venous plasma mainly as S-nitroso-albumin (7 PM) [9], and 0.3 ,uM GSNO has been measured in human bronchial lavage fluid [lo].…”

Interactions between cell surface protein disulphide isomerase and S-nitrosoglutathione during nitric oxide delivery

“…This suggests a discrepancy between the metabolism phenomenon, and subsequent signalling events. It has been documented that NO delivery into cells from protein RSNO molecules, such as albSNO, requires a prior transfer of NO to a low molecular weight thiol, via transnitrosation [8,9]. Our experiments were performed in the absence of added thiol and this may explain why metabolism of albSNO was not accompanied by cGMP accumulation or inhibition of platelet aggregation.…”

“…Several studies have reported delivery of NO into cells via transnitrosation reactions, which involve transfer of NO + from one thiol to another [5][6][7]. For protein RSNOs (such as Snitrosoalbumin) to mediate platelet inhibition, prior exchange of NO onto a low molecular weight thiol appears necessary [8,9]. Free thiols are available on the platelet surface itself [10,11] and S-nitrosation of these thiols may be a further possible pathway of anti-platelet action.…”