On the chemical biology of the nitrite/sulfide interaction

The authors are grateful to the Susanne-Bunnenberg-Stiftung of the Düsseldorf Heart Center (to MK), the COST action BM1005 (European Network on Gasotransmitters), and the Faculty of Medicine, University of Southampton (to MF) for financial support.Sulfide (H2S/HS−) has been demonstrated to exert an astounding breadth of biological effects, some of which resemble those of nitric oxide (NO). While the chemistry, biochemistry and potential pathophysiology of the cross-talk between sulfide and NO have received considerable attention lately, a comparable assessment of the potential biological implications of an interaction between nitrite and sulfide is lacking. This is surprising inasmuch as nitrite is not only a known bioactive oxidation product of NO, but also efficiently converted to S-nitrosothiols in vivo; the latter have been shown to rapidly react with sulfide in vitro, leading to formation of S/N-hybrid species including thionitrite (SNO−) and nitrosopersulfide (SSNO−). Moreover, nitrite is used as a potent remedy against sulfide poisoning in the clinic. The chemistry of interaction between nitrite and sulfide or related bioactive metabolites including polysulfides and elemental sulfur has been extensively studied in the past, yet much of this information appears to have been forgotten. In this review, we focus on the potential chemical biology of the interaction between nitrite and sulfide or sulfane sulfur molecules, calling attention to the fundamental chemical properties and reactivities of either species and discuss their possible contribution to the biology, pharmacology and toxicology of both nitrite and sulfide.Publisher PDFPeer reviewe

Nitrosopersulfide (SSNO(-)) targets the Keap-1/Nrf2 redox system.

Nitric oxide (NO), hydrogen sulfide and polysulfides have been proposed to contribute to redox signaling by activating the Keap-1/Nrf2 stress response system. Nitrosopersulfide (SSNO(-)) recently emerged as a bioactive product of the chemical interaction of NO or nitrosothiols with sulfide; upon decomposition it generates polysulfides and free NO, triggering the activation of soluble guanylate cyclase, inducing blood vessel relaxation in vitro and lowering blood pressure in vivo. Whether SSNO(-) itself interacts with the Keap-1/Nrf2 system is unknown. We therefore sought to investigate the ability of SSNO(-) to activate Nrf2-dependent processes in human vascular endothelial cells, and to compare the pharmacological effects of SSNO(-) with those of its precursors NO and sulfide at multiple levels of target engagement. We here demonstrate that SSNO(-) strongly increases Nrf2 nuclear levels, Nrf2-binding activity and transactivation activity, thereby increasing mRNA expression of Hmox-1, the gene encoding for heme oxygenase 1, without adversely affecting cell viability. Under all conditions, SSNO(-) appeared to be more potent than its parent compounds, NO and sulfide. SSNO(-)-induced Nrf2 transactivation activity was abrogated by either NO scavenging with cPTIO or inhibition of thiol sulfuration by high concentrations of cysteine, implying a role for both persulfides/polysulfides and NO in SSNO(-) mediated Nrf2 activation. Taken together, our studies demonstrate that the Keap-1/Nrf2 redox system is a biological target of SSNO(-), enriching the portfolio of bioactivity of this vasoactive molecule to also engage in the regulation of redox signaling processes. The latter suggests a possible role as messenger and/or mediator in cellular sensing and adaptations processes

On the chemical biology of the nitrite/sulfide interaction

Sulfide (H2S/HS(-)) has been demonstrated to exert an astounding breadth of biological effects, some of which resemble those of nitric oxide (NO). While the chemistry, biochemistry and potential (patho)physiology of the cross-talk between sulfide and NO has received considerable attention lately, a comparable assessment of the potential biological implications of an interaction between nitrite and sulfide is lacking. This is surprising inasmuch as nitrite is not only a known bioactive oxidation product of NO, but also efficiently converted to S-nitrosothiols in vivo; the latter have been shown to rapidly react with sulfide in vitro, leading to formation of S/N-hybrid species including thionitrite (SNO(-)) and nitrosopersulfide (SSNO(-)). Moreover, nitrite is used as a potent remedy against sulfide poisoning in the clinic. The chemistry of interaction between nitrite and sulfide or related bioactive metabolites including polysulfides and elemental sulfur has been extensively studied in the past, yet much of this information appears to have been forgotten. In this review, we focus on the potential chemical biology of the interaction between nitrite and sulfide or sulfane sulfur molecules, calling attention to the fundamental chemical properties and reactivity of either species and discuss its possible contribution to the biology, pharmacology and toxicology of both nitrite and sulfide

Human red blood cells at work: identification and visualization of erythrocytic eNOS activity in health and disease

A nitric oxide synthase (NOS)-like activity has been demonstrated in human red blood cells (RBCs), but doubts about its functional significance, isoform identity and disease relevance remain. Using flow cytometry in combination with the NO-imaging probe DAF-FM we find that all blood cells form NO intracellularly, with a rank order of monocytes > neutrophils > lymphocytes > RBCs > platelets. The observation of a NO-related fluorescence within RBCs was unexpected given the abundance of the NO-scavenger oxyhemoglobin. Constitutive normoxic NO formation was abolished by NOS inhibition and intracellular NO scavenging, confirmed by laser-scanning microscopy and unequivocally validated by detection of the DAF-FM reaction product with NO using HPLC and LC-MS/MS. Employing immunoprecipitation, ESI-MS/MS-based peptide sequencing and enzymatic assay we further demonstrate that human RBCs contain an endothelial NOS (eNOS) that converts L-3H-Arginine to L-3H-Citrulline in a Ca2+/Calmodulin-dependent fashion. Moreover, in patients with coronary artery disease, red cell eNOS expression and activity are both lower than in age-matched healthy individuals and correlate with the degree of endothelial dysfunction. Thus, human RBCs constitutively produce NO under normoxic conditions via an active eNOS isoform the activity of which is compromised in patients with coronary artery disease

Анализ минеральных равновесий термальных вод Байкальской рифтовой зоны с использованием программных пакетов Hydrogeo и PHREEQC

В работе изложены результаты сравнительного анализа значений индекса насыщения термальных вод Байкальской рифтовой зоны к ряду породообразующих минералов. Получены интересные, но в то же время спорные, результаты расчетов с применением современных программных продуктов HydroGeo и PHREEQC. В первом случае было выявлено равновесие некоторых терм к кварцу, арагониту, кальциту, доломиту, флюориту, тогда как во втором - в основном к силикатам (кварцу, халцедону, хризотилу, сепиолиту и др.) и так же к флюориту. The paper presents the results of the comparative analysis index values of saturation of the thermal waters of the Baikal rift zone to some rock-forming minerals. Interesting, but at the same time controversial, the results of calculations with the use of modern software products HydroGeo and PHREEQC. In the first case revealed a certain equilibrium therm to quartz, aragonite, calcite, dolomite, fluorite, whereas the second is mostly silicates (quartz, chalcedony chrysotile, sepiolite, etc.) and fluorite

Left ventricular diastolic dysfunction in Nrf2 knock out mice is associated with cardiac hypertrophy, decreased expression of SERCA2a, and preserved endothelial function

AbstractIncreased production of reactive oxygen species and failure of the antioxidant defense system are considered to play a central role in the pathogenesis of cardiovascular disease. The transcription factor nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a key master switch controlling the expression of antioxidant and protective enzymes, and was proposed to participate in protection of vascular and cardiac function. This study was undertaken to analyze cardiac and vascular phenotype of mice lacking Nrf2. We found that Nrf2 knock out (Nrf2 KO) mice have a left ventricular (LV) diastolic dysfunction, characterized by prolonged E wave deceleration time, relaxation time and total diastolic time, increased E/A ratio and myocardial performance index, as assessed by echocardiography. LV dysfunction in Nrf2 KO mice was associated with cardiac hypertrophy, and a downregulation of the sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) in the myocardium. Accordingly, cardiac relaxation was impaired, as demonstrated by decreased responses to β-adrenergic stimulation by isoproterenol ex vivo, and to the cardiac glycoside ouabain in vivo. Surprisingly, we found that vascular endothelial function and endothelial nitric oxide synthase (eNOS)-mediated vascular responses were fully preserved, blood pressure was decreased, and eNOS was upregulated in the aorta and the heart of Nrf2 KO mice. Taken together, these results show that LV dysfunction in Nrf2 KO mice is mainly associated with cardiac hypertrophy and downregulation of SERCA2a, and is independent from changes in coronary vascular function or systemic hemodynamics, which are preserved by a compensatory upregulation of eNOS. These data provide new insights into how Nrf2 expression/function impacts the cardiovascular system

Biological hydropersulfides and related polysulfides – a new concept and perspective in redox biology

The chemical biology of thiols (RSH, e.g., cysteine and cysteine‐containing proteins/peptides) has been a topic of extreme interest for many decades due to their reported roles in protein structure/folding, redox signaling, metal ligation, cellular protection, and enzymology. While many of the studies on thiol/sulfur biochemistry have focused on thiols, relatively ignored have been hydropersulfides (RSSH) and higher order polysulfur species (RSSnH, RSSnR, n > 1). Recent and provocative work has alluded to the prevalence and likely physiological importance of RSSH and related RSSnH. RSSH of cysteine (Cys‐SSH) has been found to be prevalent in mammalian systems along with Cys‐SSH‐containing proteins. The RSSH functionality has not been examined to the extent of other biologically relevant sulfur derivatives (e.g., sulfenic acids, disulfides, etc.), whose roles in cell signaling are strongly indicated. The recent finding of Cys‐SSH biosynthesis and translational incorporation into proteins is an unequivocal indication of its fundamental importance and necessitates a more profound look into the physiology of RSSH. In this Review, we discuss the currently reported chemical biology of RSSH (and related species) as a prelude to discussing their possible physiological roles

Long-lasting blood pressure lowering effects of nitrite are NO-independent and mediated by hydrogen peroxide, persulfides, and oxidation of protein kinase G1α redox signalling

Aims Under hypoxic conditions, nitrite (NO2-) can be reduced to nitric oxide (NO) eliciting vasorelaxation. However, nitrite also exerts vasorelaxant effects of potential therapeutic relevance under normal physiological conditions via undetermined mechanisms. We, therefore, sought to investigate the mechanism(s) by which nitrite regulates the vascular system in normoxia and, specifically, whether the biological effects are a result of NO generation (as in hypoxia) or mediated via alternative mechanisms involving classical downstream targets of NO [e.g. effects on protein kinase G1 alpha (PKG1 alpha)]. Methods and results Ex vivo myography revealed that, unlike in thoracic aorta (conduit vessels), the vasorelaxant effects of nitrite in mesenteric resistance vessels from wild-type (WT) mice were NO-independent. Oxidants such as H2O2 promote disulfide formation of PKG1 alpha, resulting in NO- cyclic guanosine monophosphate (cGMP) independent kinase activation. To explore whether the microvascular effects of nitrite were associated with PKG1 alpha oxidation, we used a Cys42Ser PKG1 alpha knock-in (C42S PKG1 alpha KI; 'redox-dead') mouse that cannot transduce oxidant signals. Resistance vessels from these C42S PKG1 alpha KI mice were markedly less responsive to nitrite-induced vasodilation. Intraperitoneal (i.p.) bolus application of nitrite in conscious WT mice induced a rapid yet transient increase in plasma nitrite and cGMP concentrations followed by prolonged hypotensive effects, as assessed using in vivo telemetry. In the C42S PKG1 alpha KI mice, the blood pressure lowering effects of nitrite were lower compared to WT. Increased H2O2 concentrations were detected in WT resistance vessel tissue challenged with nitrite. Consistent with this, increased cysteine and glutathione persulfide levels were detected in these vessels by mass spectrometry, matching the temporal profile of nitrite's effects on H2O2 and blood pressure. Conclusion Under physiological conditions, nitrite induces a delayed and long-lasting blood pressure lowering effect, which is NO-independent and occurs via a new redox mechanism involving H2O2, persulfides, and PKG1 alpha oxidation/activation. Targeting this novel pathway may provide new prospects for anti-hypertensive therapy

Red Blood Cell and Endothelial eNOS Independently Regulate Circulating Nitric Oxide Metabolites and Blood Pressure

Background: Current paradigms suggest that nitric oxide (NO) produced by endothelial cells (ECs) through endothelial nitric oxide synthase (eNOS) in the vessel wall is the primary regulator of blood flow and blood pressure. However, red blood cells (RBCs) also carry a catalytically active eNOS, but its role is controversial and remains undefined. This study aimed to elucidate the functional significance of RBC eNOS compared with EC eNOS for vascular hemodynamics and nitric oxide metabolism. Methods: We generated tissue-specific loss- and gain-of-function models for eNOS by using cell-specific Cre-induced gene inactivation or reactivation. We created 2 founder lines carrying a floxed eNOS (eNOSflox/flox) for Cre-inducible knockout (KO), and gene construct with an inactivated floxed/inverted exon (eNOSinv/inv) for a Cre-inducible knock-in (KI), which respectively allow targeted deletion or reactivation of eNOS in erythroid cells (RBC eNOS KO or RBC eNOS KI mice) or in ECs (EC eNOS KO or EC eNOS KI mice). Vascular function, hemodynamics, and nitric oxide metabolism were compared ex vivo and in vivo. Results: The EC eNOS KOs exhibited significantly impaired aortic dilatory responses to acetylcholine, loss of flow-mediated dilation, and increased systolic and diastolic blood pressure. RBC eNOS KO mice showed no alterations in acetylcholine-mediated dilation or flow-mediated dilation but were hypertensive. Treatment with the nitric oxide synthase inhibitor Nγ-nitro-l-arginine methyl ester further increased blood pressure in RBC eNOS KOs, demonstrating that eNOS in both ECs and RBCs contributes to blood pressure regulation. Although both EC eNOS KOs and RBC eNOS KOs had lower plasma nitrite and nitrate concentrations, the levels of bound NO in RBCs were lower in RBC eNOS KOs than in EC eNOS KOs. Reactivation of eNOS in ECs or RBCs rescues the hypertensive phenotype of the eNOSinv/invmice, whereas the levels of bound NO were restored only in RBC eNOS KI mice. Conclusions: These data reveal that eNOS in ECs and RBCs contribute independently to blood pressure homeostasis