S-Nitrosothiols have been implicated to play key roles in a variety of physiological processes. The potential physiological importance of S-nitrosothiols prompted us to examine their reaction with thiols. We find that S-nitrosothiols can react with thiols to generate nitroxyl (HNO) and the corresponding disulfide. Further reaction of HNO with the remaining S-nitrosothiol and thiol results in the generation of other species including NO, sulfinamide, and hydroxylamine. Mechanisms are proposed that rationalize the observed products.

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Peroxynitrite (ONOO ؊ ), a potent oxidant formed by reaction of nitric oxide (NO ⅐ ) with superoxide anion, can activate guanylyl cyclase and is able to induce vasodilation or inhibit platelet aggregation and leukocyte adhesion, via thiol-dependent formation of NO ⅐ . Reaction of ONOO ؊ with thiols is thought to proceed through formation of a S-nitrothiol (thionitrate; RSNO 2 ) intermediate and yields low levels of S-nitrosothiols (thionitrites; RSNO), both of which are theoretical sources of NO (2, 3). An important component of NO ⅐ biochemistry involves the formation of thionitrite esters with cysteine or cysteine-residues (Snitrosothiols; RSNO), and low molecular weight S-nitrosothiols or S-nitrosothiol adducts in proteins such as albumin or hemoglobin are known to be generated in vivo (4 -7). Formation of S-nitrosothiols may provide a buffering system regulating the bioavailability of NO ⅐ and/or serve to increase its range of action, since S-nitrosothiols are more stable and can release NO ⅐ by site-specific regulatory mechanisms, via reactions with transition metal ions or other reducing systems such as thioredoxin or superoxide anion (7-10). Furthermore, S-nitrosation and transnitrosation reactions with protein cysteine residues may regulate the activity of many enzymes and represent an important signaling mechanism (11-13). Despite the growing interest in the role of S-nitrosothiols in the biological actions of NO ⅐ , there is still uncertainty regarding the mechanism of S-nitrosation in vivo, which most likely does not involve direct reaction of NO ⅐ with thiols but requires electrophilic activation to a nitrosyl cation (NO ϩ )-like intermediate. Metabolites formed during NO ⅐ (auto)oxidation, such as dinitrogen trioxide (N 2 O 3 ), are likely candidates in this respect (14 -17). Alternatively, NO ⅐ can be activated to intermediates with NO ϩ character via interaction with ferric heme proteins (18) or via formation of dinitrosyl-iron complexes (19), in both cases leading to more efficient S-nitrosation. Finally, thiols are important physiological targets for peroxynitrite (ONOO Ϫ ) 2 (20), a potent oxidant formed during simultaneous generation of NO ⅐ and superoxide anion (O 2 . ) (4, 21), and formation of S-nitrosothiols has been detected after reaction of ONOO Ϫ with thiols (22,23). This chemistry has been linked to an increasing number of findings that ONOO -is capable of activating guanylyl cyclase (23-25) and can induce vasodilation, inhibition of platelet aggregation, or leukocyte adhesion and activation (26 -29), actions involving (re)generation of NO ⅐ via reaction with * This study was supported by National Institutes of Health Grant HL57452 (to C. E. C.), the Cystic Fibrosis Foundation, the American Lung Association of California, and a Fellowship from the Parker B. Francis Foundation (to A. v. d. V.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Sectio...

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The arylhydrocarbon receptor (AhR) is known for its ability to bind aromatic-containing compounds, which starts a molecular cascade involving the induction of cytochrome P450s and inflammatory cytokines. Our hypothesis is that many inhaled environmental toxicant components activate these inflammatory pathways via an initial binding to the AhR. To test this possibility, we treated Clara cell-derived NCI-H441 cells with the AhR agonist, 2,3,7,8-tetrachlordibenzo-p-dioxin (TCDD), and demonstrated that AhR activation increased the expression of both cytochrome P450 s and inflammatory markers. We also found increased mucin 5AC production with TCDD treatment. Similar results were observed in NCI-H441 cells treated with urban dust particles. Mucin 5AC expression was highly correlated with increased-expression cyclooxygenase-2 and IL-1b, thus implicating these two inflammatory markers as possible conduits for AhR-mediated mucin production. We hypothesize that this increase in mucin 5AC production is a result of inflammation-induced differentiation of our epithelial cell to a mucin-producing cell. This theory is supported by morphological changes observed in the cells, as well as decreased expression of Clara cell secretory protein (CC10). In an in vivo C57BL/6 mouse model, TCDD increased expression of inflammatory cytokines, mucin 5AC, and a number of matrix metalloproteases in whole-lung samples. These changes were not seen in mice in which AhR signaling was repressed. These markers from the whole-lung samples have been correlated to onset of bronchitis, asthma, small airways disease, and fibrosis, and their increased expression further implicates AhR activation in producing the molecular environment for the development of lung injury to occur.

Protein S-thiolation, the formation of mixed disulphides of cysteine residues in proteins with low-molecular-mass thiols, occurs under conditions associated with oxidative stress and can lead to modification of protein function. In the present study, we examined the site of S-thiolation of the enzyme creatine kinase (CK), an important source of ATP in myocytes. Inactivation of this enzyme is thought to play a critical role in cardiac injury during oxidative stress, such as during reperfusion injury. Reaction of rabbit CK M isoenzyme with GSSG, used to model protein S-thiolation, was found to result in enzyme inactivation that could be reversed by GSH or dithiothreitol. Measurement of GSH that is released during the thiolation reaction indicated that the maximum extent of CK thiolation was approx. 1 mol of GSH/mol of protein, suggesting thiolation on one reactive cysteine residue. Accordingly, matrix-assisted laser-desorption ionization MS confirmed that the molecular mass of CK was increased, consistent with addition of one GSH molecule/molecule of CK. Using trypsin digestion, HPLC and MS analysis, the active-site cysteine residue (Cys(283)) was identified as the site of thiolation. Reversal of thiolation was shown to be rapid when GSH is abundant, rendering dethiolation of CK thermodynamically favoured within the cell. We conclude that S-glutathionylation of CK could be one mechanism to explain temporary reversible loss in activity of CK during ischaemic injury. The maintainance of GSH levels represents an important mechanism for regeneration of active CK from S-glutathionylated CK.

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Background Medication adherence is crucial for improving clinical outcomes in the treatment of patients. We evaluate the effect of short message service (SMS) reminder on medication adherence and serum hormones in patients with breast cancer on aromatase inhibitors. Methods An open-label, multi-centre, prospective randomised controlled trial of SMS versus Standard Care was conducted. Medication adherence was assessed via self-report using the Simplified Medication Adherence Questionnaire at baseline, 6 month, and 1 year. Androstenedione, estradiol, and estrone were measured at baseline and 1 year. The χ 2 test and mixed effects logistic regression was performed to compare medication adherence between groups. Difference in androstenedione and estrone levels were assessed using analysis of covariance, whereas χ 2 test and logistic regression was used for estradiol. Analysis was based on intention-to-treat. Results A total of 244 patients were randomised to receive weekly SMS reminder ( n = 123) or Standard Care ( n = 121) between May 2015 and December 2018. The odds of adherence was higher at 6-month in SMS (OR = 1.78, 95% CI 1.04–3.05, p = 0.034), and not significantly different at 1-year (OR = 1.15, 95% CI: 0.67–1.96 p = 0.617). Mixed effects logistic regression analysis showed higher odds of adherence in SMS over the 1-year period (OR = 2.35, 95% CI: 1.01–5.49, p = 0.048). There was no difference in serum hormone levels between groups. Conclusion SMS reminder improved medication adherence in the short-term but had no effect on serum hormones levels in the longer term. Future studies could investigate the use of tailored SMS intervention according to patient preference to improve its sustainability.