Original ContributionInterference of Carboxy-PTIO with Nitric Oxide- and Peroxynitrite-Mediated Reactions
Introduction
NO has several important biological functions, including the regulation of vascular tone, inhibition of platelet aggregation, modulation of synaptic transmission in the brain, and neurotransmission in the peripheral nervous system.[1] NO can undergo a variety of reactions within cells, and to understand NO-mediated regulatory effects it is necessary to study the rates of these reactions and the activity of their products.[2] The reactions with molecular oxygen, superoxide, and iron proteins are already known to be physiologically significant.[3] The reaction with oxygen is second order with respect to NO,4, 5, 6 and yields intermediates with potential cytotoxic and mutagenic properties.7, 8 It is not yet clear which of the possible nitrogen oxides are intermediates in this reaction.4, 9
One of the fastest reactions of NO is the nearly diffusion-controlled combination with superoxide to form peroxynitrite,10, 11 a potent oxidant of many biological molecules.[12] In a metal-catalyzed reaction, peroxynitrite nitrates several phenolic compounds including free and peptide tyrosine.13, 14, 15 Peroxynitrite is protonated with a pKa of 6.8, and the corresponding peroxynitrous acid rapidly decomposes to form a potent oxidant with hydroxyl radical-like properties.[10] The precise mechanism of peroxynitrite decomposition is not clear yet, but it appears that free radical intermediates are not involved.[12]
Several reports indicate that peroxynitrite has similar physiological effects to NO. It induces NO-like relaxation of vascular smooth muscle,16, 17 inhibits platelet aggregation,[17] and stimulates purified soluble guanylyl cyclase.[18] These effects may be due to formation of small amounts of NO during peroxynitrite decomposition[16] or result from S-nitrosation of thiols followed by release of free NO from the corresponding thionitrites.18, 19 Thus, selective NO scavengers are required to discriminate between free NO and other species exhibiting NO-like biological activity. Reduced hemoglobin was described as a potent and specific scavenger of NO,[20] but authentic peroxynitrite reacts rapidly with hemoglobin in a manner indistinguishable from NO-induced formation of methemoglobin.[21] Another class of NO scavengers are nitronyl nitroxides.[22] The prototypes of such stable free radicals are PTIO and its water-soluble derivative carboxy-PTIO, which react rapidly with NO to yield the corresponding imidazolineoxyl and free NO2 radical.[23] Based on susceptibility to carboxy-PTIO, it was reported that EDRF was identical with NO, whereas the nitrergic neurotransmitter producing relaxation of certain smooth muscles was not.23, 24 To find out whether carboxy-PTIO can be regarded as a specific NO scavenger, we have studied the effect of this drug on several NO- and peroxynitrite-mediated reactions.
Section snippets
Materials
Solutions of peroxynitrite (80–100 mM) were prepared as described[21] and diluted to 10 mM with H2O prior to experiments. GSNO and carboxy-PTIO were purchased from ALEXIS Corp., Switzerland. DEA/NO was from NCI Chemical Carcinogen Repository, Kansas City, MO; 10-fold concentrated stock solutions of the NO donor were prepared daily in 10 mM NaOH. SIN-1, a generous gift from Cassella-Riedel, Frankfurt, Germany, was dissolved at pH 5.0 prior to use. The monoclonal nitrotyrosine antibody was a
Results
The ability of carboxy-PTIO to scavenge NO was assessed by determination of NO-induced accumulation of cGMP in cultured porcine aortic endothelial cells in the absence and presence of the drug. Basal cGMP levels of unstimulated cells were 2.2 ± 0.7 pmol/106 cells. As shown in Fig. 1, 1 μM DEA/NO induced a 20-fold increase in cGMP (44 ± 12.5 pmol/106 cells) and this effect was inhibited by 0.1 mM carboxy-PTIO to 16.3 ± 4.8% of the control. In the presence of the Ca2+ ionophore A 23187 (0.3 μM),
Discussion
Our results on inhibition of NO-induced cGMP accumulation in cultured cells and inhibition of NO-stimulated soluble guanylyl cylcase confirm that carboxy-PTIO is a potent scavenger of NO, but the action profile of this compound may be more complex than hitherto assumed. Unexpectedly, carboxy-PTIO did not inhibit but potentiated the effect of SIN-1 on cGMP accumulation in endothelial cells. Decomposition of SIN-1 yields NO and superoxide,[36] suggesting that the sydnonimine may be a donor of
Acknowledgements
We thank an anonymous referee for his comments on oxidation of GSH by NO2 and Margit Rehn for excellent technical assistance. This work was supported by Grants 10655, 10859, 11478 (to B.M.), F712 (K.S.), and 11301 (to E.R.W.) of the Fonds zur Förderung der Wissenschaftlichen Forschung in Österreich.
References (43)
- et al.
Modulation of superoxide-dependent oxidation and hydroxylation reactions by nitric oxide
J. Biol. Chem.
(1996) - et al.
Kinetics of nitric oxide autoxidation in aqueous solution
J. Biol. Chem.
(1994) - et al.
Kinetics and mechanism of tetrahydrobiopterin-induced oxidation of nitric oxide
J. Biol. Chem.
(1995) - et al.
Peroxynitrite-mediated tyrosine nitration catalysed by superoxide dismutase
Arch. Biochem. Biophys.
(1992) - et al.
Kinetics of superoxide dismutase- and iron-catalyzed nitration of phenolics by peroxynitrite
Arch. Biochem. Biophys.
(1992) - et al.
Peroxynitrite-induced accumulation of cyclic GMP in endothelial cells and stimulation of purified soluble guanylyl cyclase. Dependence on glutathione and possible role of S-nitrosation
J. Biol. Chem.
(1995) - et al.
Decomposition of S-Nitrosoglutathione in the presence of copper and glutathione
Arch. Biochem. Biophys.
(1996) - et al.
Correlation between nitric oxide formation during degradation of organic nitrates and activation of guanylate cyclase
Eur. J. Pharmacol.
(1987) - et al.
Inhibition of nitric oxide synthesis by methylene blue
Biochem. Pharmacol.
(1993) - et al.
Analysis of nitrate, nitrite, and [15N]nitrate in biological fluids
Anal. Biochem.
(1982)
Induction of GTP cyclohydrolase I by bacterial lipopolysaccharide in the rat
FEBS Lett.
Autoxidation kinetics of aqueous nitric oxide
FEBS Lett.
Oxidation of nitrogen oxides by bound dioxygen in hemoproteins
J. Inorg. Biochem.
Reactions of nitrogen dioxide in aqueous model systems: Oxidation of tyrosine units in peptides and proteins
Arch. Biochem. Biophys.
Effect of calcium on endothelium-derived relaxing factor formation and cGMP levels in endothelial cells
Eur. J. Pharmacol.
Peroxynitrite oxidation of sulfhydryls. The cytotoxic potential of superoxide and nitric oxide
J. Biol. Chem.
Kinetics of nitrosation of thiols by nitric oxide in the presence of oxygen
J. Biol. Chem.
Nitric oxidePhysiology, pathophysiology, and pharmacology
Pharmacol. Rev.
EPR characterization of molecular targets for NO in mammalian cells and organelles
FASEB J.
Reactions of the bioregulatory agent nitric oxide in oxygenated aqueous media: Determination of the kinetics for oxidation and nitrosation by intermediates generated in the NO/O2 reaction
Chem. Res. Toxicol.
DNA deaminating ability and genotoxicity of nitric oxide and its progenitors
Science
Cited by (83)
Enhancing agroecosystem nitrogen management: microbial insights for improved nitrification inhibition
2023, Trends in MicrobiologyMagnesium hydride acts as a convenient hydrogen supply to prolong the vase life of cut roses by modulating nitric oxide synthesis
2021, Postharvest Biology and TechnologyCitation Excerpt :A schematic model is shown in Fig. 6, summarizing the vital role of NO for MgH2-prolonged vase life of cut roses. Although we can not exclude the possibility that both PTIO and tungstate used in this study might not specifically target NO and its synthesis (Pfeiffer et al., 1997; Xiong et al., 2012), the results obtained from these experiments with pharmacological manipulation of endogenous NO levels, clearly revealed that the function of MgH2 in prolonging vase life of cut rose flowers might be mediated by NO via NR. The results of this work revealed that the application of MgH2 was as efficient as HRW on postharvest preservation of cut flowers by reestablishing redox homeostasis.
Sialic acid–binding immunoglobulin-like lectin 8 (Siglec-8) is an activating receptor mediating β<inf>2</inf>-integrin–dependent function in human eosinophils
2018, Journal of Allergy and Clinical ImmunologyCardiac contractility in Antarctic teleost is modulated by nitrite through xanthine oxidase and cytochrome p-450 nitrite reductase
2015, Nitric Oxide - Biology and ChemistryCitation Excerpt :PTIO is currently used as a selective NO scavenger [35]. However, studies on its effects on NO- and peroxynitrite-mediated reactions showed the formation of bioactive reaction products, which may induce unexpected effects, such as an increased inhibition of dopamine uptake by NO donors [36,37]. Another important point to consider in interpreting PTIO results is the drug tissue distribution during heart perfusion.