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Oxidation, Glutathione leads to S-Glutathionylation, eNOS
Key Event Relationship Overview
AOPs Referencing Relationship
Life Stage Applicability
|All life stages||High|
Key Event Relationship Description
Under oxidative stress, reduced glutathione (GSH) is oxidized to glutathione disulfide (GSSG), which then induces S-glutathionylation of eNOS at its cysteine residues, resulting in eNOS uncoupling (Chen et al., 2010).
Evidence Supporting this KER
In an in vitro system, GSSG induced S-glutathionylation of human eNOS in a dose-dependent manner, which was reversed by reducing agents 2-mercaptoethanol and dithiothreitol (Chen et al., 2010). Cysteine residues 689 and 908 were identified as the critical sites for S-glutathionylation of eNOS since their mutatation resisted glutathionylation. These results were confirmed in bovine aortic endothelial cells (BAECs) and in aortae of spontaneously hypertensive (SHR) rats. Treatment with 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), an inhibitor of glutathione reductase, induced an increase in GSSG and eNOS S-glutathionylation as confirmed by mass spectrometry in which Cys 689 was shown to be more than 50% S-glutathionylated. Aortae of SHR rats showed S-glutathionylation via immunohistology whereas normotensive rats had little S-glutathionylation. In human aortic endothelial cells (HAECs) exposed to ultrafine particles (UFP), GSSG levels were increased in parallel with protein S-glutathionylation, which was confirmed by ELISA to be eNOS S-glutathionylation (Du et al., 2013). This was recapitulated in LDLR-null mice exposed to UFP. Additional evidence was observed in BAECs undergoing hypoxia and reoxygenation where eNOS S-glutathionylation increased by three-fold compared to control cells and GSSG levels were increased (De Pascali et al., 2014). These effects were reversed with treatment of N-acetyl-l-cysteine, which increased cellular concentration of GSH. These results support strong biological plausibility for this key event relationship.
Uncertainties and Inconsistencies
No uncertainties or inconsistencies were found for this KER.
Known modulating factors
Known Feedforward/Feedback loops influencing this KER
Domain of Applicability
The evidence supporting this key event relationship are from BAECs, HAECs, LDLR-null mice, and SHR rats (Chen et al., 2010; De Pascali et al., 2014; Du et al., 2013).
Chen, C.-A., Wang, T.-Y., Varadharaj, S., Reyes, L.A., Hemann, C., Talukder, M.A.H., Chen, Y.-R., Druhan, L.J., and Zweier, J.L. (2010). S-glutathionylation uncouples eNOS and regulates its cellular and vascular function. Nature 468, 1115–1118.
De Pascali, F., Hemann, C., Samons, K., Chen, C.-A., and Zweier, J.L. (2014). Hypoxia and reoxygenation induce endothelial nitric oxide synthase uncoupling in endothelial cells through tetrahydrobiopterin depletion and S-glutathionylation. Biochemistry (Mosc.) 53, 3679–3688.
Du, Y., Navab, M., Shen, M., Hill, J., Pakbin, P., Sioutas, C., Hsiai, T.K., and Li, R. (2013). Ambient ultrafine particles reduce endothelial nitric oxide production via S-glutathionylation of eNOS. Biochem. Biophys. Res. Commun. 436, 462–466.