scholarly journals Involvement of a local Fenton reaction in the reciprocal modulation by O2 of the glucagon-dependent activation of the phosphoenolpyruvate carboxykinase gene and the insulin-dependent activation of the glucokinase gene in rat hepatocytes

1998 ◽  
Vol 335 (2) ◽  
pp. 425-432 ◽  
Author(s):  
Thomas KIETZMANN ◽  
Torsten PORWOL ◽  
Karl ZIEROLD ◽  
Kurt JUNGERMANN ◽  
Helmut ACKER
Keyword(s):  
Hydroxyl Radical ◽  
Hydroxyl Radicals ◽  
Fenton Reaction ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Gene Activation ◽  
Three Dimensional ◽  
Rat Hepatocytes ◽  
Confocal Laser ◽  
Radical Scavenger ◽  
Insulin Dependent

H2O2 mimicked the action of periportal pO2 in the modulation by O2 of the glucagon-dependent activation of the phosphoenolpyruvate carboxykinase (PCK) gene and the insulin-dependent activation of the glucokinase (GK) gene. H2O2 can be converted in the presence of Fe2+ in a Fenton reaction into hydroxyl anions and hydroxyl radicals (•OH). The hydroxyl radicals are highly reactive and might interfere locally with transcription factors. It was the aim of the present study to investigate the role of and to localize such a Fenton reaction. Hepatocytes cultured for 24 h were treated under conditions mimicking periportal or perivenous pO2 with glucagon or insulin plus the iron chelator desferrioxamine (DSF) or the hydroxyl radical scavenger dimethylthiourea (DMTU) to inhibit the Fenton reaction. PCK mRNA was induced by glucagon maximally under conditions of periportal pO2 and half-maximally under venous pO2. GK mRNA was induced by insulin with reciprocal modulation by O2. DSF and DMTU reduced the induction of PCK mRNA to about half-maximal and increased the induction of GK mRNA to maximal under both O2 tensions. Hydroxyl radical formation was maximal under arterial pO2. Perivenous pO2, DSF and DMTU each decreased the formation of •OH to about 70% of control. The Fenton reaction could be localized in a perinuclear space by confocal laser microscopy and three-dimensional reconstruction techniques. In the same compartment, iron could be detected by electron-probe X-ray microanalysis. Thus a local Fenton reaction is involved in the O2 signalling, which modulated the glucagon- and insulin-dependent PCK gene and GK gene activation.

scholarly journals Production of hydroxyl radicals from the simultaneous generation of superoxide and nitric oxide

1992 ◽  
Vol 281 (2) ◽  
pp. 419-424 ◽  
Author(s):  
N Hogg ◽  
V M Darley-Usmar ◽  
M T Wilson ◽  
S Moncada
Keyword(s):  
Nitric Oxide ◽  
Hydroxyl Radical ◽  
Hydroxyl Radicals ◽  
Sodium Benzoate ◽  
Fenton Reaction ◽  
Fluorescence Emission ◽  
Emission Spectra ◽  
Radical Scavenger ◽  
Fluorescence Emission Spectra ◽  
Simultaneous Generation

Both nitric oxide (NO) and superoxide are generated by macrophages, neutrophils and endothelial cells. It has been postulated that the generation of these two radicals under physiological conditions can lead to the formation of peroxynitrite and (as a result of the homolytic lysis of this molecule) the production of hydroxyl radicals. We have used 3-morpholinosydnonimine N-ethylcarbamide (SIN-1), a sydnonimine capable of generating both NO and superoxide simultaneously, to test this hypothesis. SIN-1 (1 mM) generated superoxide and NO at rates of 7.02 microM/min and 3.68 microM/min respectively in phosphate-buffered saline, pH 7.2, at 37 degrees C. Incubation of SIN-1 with both deoxyribose and sodium benzoate resulted in the formation of malondialdehyde (MDA). In addition, the incubation of SIN-1 with sodium benzoate resulted in the production of compounds with fluorescence emission spectra characteristic of hydroxylated products. Both the production of MDA and the generation of fluorescent compounds were inhibited by the hydroxyl radical scavenger mannitol. In all the above respects, SIN-1 mimicked the production of hydroxyl radicals from the ascorbate-driven Fenton reaction. Catalase had no effect on the SIN-1-dependent generation of MDA, and superoxide dismutase was partially inhibitory. SIN-1 produces an oxidant with the properties of the hydroxyl radical by a mechanism clearly different to that of the Fenton reaction. We conclude that the simultaneous production of NO and superoxide from SIN-1 results in the formation of hydroxyl radicals.

scholarly journals Contribution of Oxidative Damage to Antimicrobial Lethality

2009 ◽  
Vol 53 (4) ◽  
pp. 1395-1402 ◽  
Author(s):  
Xiuhong Wang ◽  
Xilin Zhao
Keyword(s):  
Oxidative Stress ◽  
Escherichia Coli ◽  
Hydroxyl Radical ◽  
Hydroxyl Radicals ◽  
Fenton Reaction ◽  
Iron Chelator ◽  
Radical Scavenger ◽  
Alkyl Hydroperoxide ◽  
Hydroxyl Radical Scavenger ◽  
Rapid Conversion

ABSTRACT A potential pathway linking hydroxyl radicals to antimicrobial lethality was examined by using mutational and chemical perturbations of Escherichia coli. Deficiencies of sodA or sodB had no effect on norfloxacin lethality; however, the absence of both genes together reduced lethal activity, consistent with rapid conversion of excessive superoxide to hydrogen peroxide contributing to quinolone lethality. Norfloxacin was more lethal with a mutant deficient in katG than with its isogenic parent, suggesting that detoxification of peroxide to water normally reduces quinolone lethality. An iron chelator (bipyridyl) and a hydroxyl radical scavenger (thiourea) reduced the lethal activity of norfloxacin, indicating that norfloxacin-stimulated accumulation of peroxide affects lethal activity via hydroxyl radicals generated through the Fenton reaction. Ampicillin and kanamycin, antibacterials unrelated to fluoroquinolones, displayed behavior similar to that of norfloxacin except that these two agents showed hyperlethality with an ahpC (alkyl hydroperoxide reductase) mutant rather than with a katG mutant. Collectively, these data are consistent with antimicrobial stress increasing the production of superoxide, which then undergoes dismutation to peroxide, from which a highly toxic hydroxyl radical is generated. Hydroxyl radicals then enhance antimicrobial lethality, as suggested by earlier work. Such findings indicate that oxidative stress networks may provide targets for antimicrobial potentiation.

scholarly journals On the Nature of Hydrodynamic Cavitation Process and Its Application for the Removal of Water Pollutants

2019 ◽  
Vol 12 ◽  
pp. 117862211988048 ◽  
Author(s):  
Erick R Bandala ◽  
Oscar M Rodriguez-Narvaez
Keyword(s):  
Hydroxyl Radical ◽  
Hydroxyl Radicals ◽  
High Energy ◽  
Optimal Operation ◽  
Hydrodynamic Cavitation ◽  
Radical Scavenger ◽  
Process Conditions ◽  
Cavitation Process ◽  
Operation Conditions ◽  
Hydroxyl Radical Scavenger

Cavitation is considered a high energy demanding process for water treatment. For this study, we used a simple experimental setup to generate cavitation at a low pressure (low energy) and test it for hydroxyl radical production using a well-known chemical probe as a hydroxyl radical scavenger. The conditions for generating the cavitation process (eg, pressure, flow velocity, temperature, and other significant variables) were used to degrade model contaminants, an azo dye and an antibiotic. The amount of hydroxyl radicals generated by the system was estimated using N,N-dimethyl-p-nitrosoaniline (pNDA) as hydroxyl radical scavenger. The capability of hydrodynamic cavitation (HC) to degrade contaminants was assessed using Congo red (CR) and sulfamethoxazole (SMX) as model contaminants. Different chemical models were analyzed using UV-visible spectrophotometry (for pNDA and CR) and high-performance liquid chromatography (HPLC) (for SMX) after HC treatment under different process conditions (ie, pressure of 13.7 and 10.3 kPa, and flow rates of 0.14 to 3.6 × 10−4 m3/s). No pNDA bleaching was observed for any of the reaction conditions tested after 60 minutes of treatment, which suggests that there was no hydroxyl radical generation during the process. However, 50% degradation of CR and 25% degradation of SMX were observed under the same process conditions, comparable with previously reported results. These results suggest that the process is most likely thermally based rather than radically based, and therefore, it can degrade organic pollutants even if no hydroxyl radicals are produced. Hydrodynamic cavitation, either alone or coupled with other advanced water technologies, has been identified as a promising technology for removing organic contaminants entering the water cycle; however, more research is still needed to determine the specific mechanisms involved in the process and the optimal operation conditions for the system.

scholarly journals Towards reliable quantification of hydroxyl radicals in the Fenton reaction using chemical probes

RSC Advances ◽  
2018 ◽  
Vol 8 (10) ◽  
pp. 5321-5330 ◽  
Author(s):  
Burgos Castillo Rutely C. ◽  
Fontmorin Jean-M. ◽  
Tang Walter Z. ◽  
Dominguez-Benetton Xochitl ◽  
Sillanpää Mika
Keyword(s):  
Hydroxyl Radical ◽  
Hydroxyl Radicals ◽  
Fenton Reaction ◽  
Chemical Probes ◽  
Radical Concentration ◽  
Reliable Quantification

Quantification of hydroxyl radical concentration using two chemical probes was assessed through the Fenton reaction.

scholarly journals Superoxide dismutase enhances the formation of hydroxyl radicals in the reaction of 3-hydroxyanthranilic acid with molecular oxygen

1988 ◽  
Vol 251 (3) ◽  
pp. 893-899 ◽  
Author(s):  
H Iwahashi ◽  
T Ishii ◽  
R Sugata ◽  
R Kido
Keyword(s):  
Hydrogen Peroxide ◽  
Superoxide Dismutase ◽  
Hydroxyl Radical ◽  
Molecular Oxygen ◽  
Hydroxyl Radicals ◽  
Fenton Reaction ◽  
Potassium Phosphate ◽  
Radical Formation ◽  
Superoxide Anions ◽  
Hydroxyanthranilic Acid

Superoxide dismutase (SOD) enhanced the formation of hydroxyl radicals, which were detected by using the e.s.r. spin-trapping technique, in a reaction mixture containing 3-hydroxyanthranilic acid (or p-aminophenol), Fe3+ ions, EDTA and potassium phosphate buffer, pH 7.4. The hydroxyl-radical formation enhanced by SOD was inhibited by catalase and desferrioxamine, and stimulated by EDTA and diethylenetriaminepenta-acetic acid, suggesting that both hydrogen peroxide and iron ions participate in the reaction. The hydroxyl-radical formation enhanced by SOD may be considered to proceed via the following steps. First, 3-hydroxyanthranilic acid is spontaneously auto-oxidized in a process that requires molecular oxygen and yields superoxide anions and anthranilyl radicals. This reaction seems to be reversible. Secondly, the superoxide anions formed in the first step are dismuted by SOD to generate hydrogen peroxide and molecular oxygen, and hence the equilibrium in the first step is displaced in favour of the formation of superoxide anions. Thirdly, hydroxyl radicals are generated from hydrogen peroxide through the Fenton reaction. In this Fenton reaction Fe2+ ions are available since Fe3+ ions are readily reduced by 3-hydroxyanthranilic acid. The superoxide anions do not seem to participate in the reduction of Fe3+ ions, since superoxide anions are rapidly dismuted by SOD present in the reaction mixture.

scholarly journals Copper-ion-dependent damage to the bases in DNA in the presence of hydrogen peroxide

1991 ◽  
Vol 273 (3) ◽  
pp. 601-604 ◽  
Author(s):  
O I Aruoma ◽  
B Halliwell ◽  
E Gajewski ◽  
M Dizdaroglu
Keyword(s):  
Ascorbic Acid ◽  
Superoxide Dismutase ◽  
Hydroxyl Radical ◽  
Hydroxyl Radicals ◽  
Copper Ion ◽  
Nitrilotriacetic Acid ◽  
Dna Bases ◽  
Radical Scavenger ◽  
Base Damage

Mixtures of Cu2+ and H2O2 at pH 7.4 caused damage to the bases in DNA greater than that caused by mixtures of Fe3+ and H2O2. Addition of ascorbic acid to the Cu2+/H2O2 system caused a very large increase in base damage, much greater than that produced by the Fe3+/H2O2/ascorbic acid system. The products of base damage in the presence of Cu2+ were typical products that have been shown to result from attack of hydroxyl radicals upon the DNA bases. Cytosine glycol, thymine glycol, 8-hydroxyadenine and especially 8-hydroxyguanine were the major products in both the Cu2+/H2O2 and the Cu2+/H2O2/ascorbic acid systems. Base damage in DNA by these systems was inhibited by the chelating agents EDTA and nitrilotriacetic acid and by catalase, but not by superoxide dismutase, nor by the hydroxyl-radical scavenger mannitol. It is proposed that Cu2+ ions bound to the DNA react with H2O2 and ascorbic acid to generate hydroxyl radicals, which then immediately attack the DNA bases in a site-specific manner. A hypoxanthine/xanthine oxidase system also caused damage to the DNA bases in the presence of Cu2+ ions. This was inhibited by superoxide dismutase and catalase. The high activity of Cu2+ ions, when compared with Fe3- ions, in causing hydroxyl-radical-dependent damage to DNA and to other biomolecules, means that the availability of Cu2+ ions in vivo must be carefully controlled.

scholarly journals Hydroxyl Radical (ºOH) Scavenger Power of Tris (hydroxymethyl) Compared to Phosphate Buffer

2016 ◽  
Vol 6 (1) ◽  
pp. 52 ◽  
Author(s):  
Meysam Khosravifarsani ◽  
Ali Shabestani-Monfared ◽  
Mahdi Pouramir ◽  
Ebrahim Zabihi
Keyword(s):  
Hydroxyl Radicals ◽  
Phosphate Buffer ◽  
Ferrous Sulfate ◽  
Fenton Reaction ◽  
Experimental Investigations ◽  
Radical Scavenger ◽  
Protective Effects ◽  
Two Samples ◽  
Gamma Radiolysis

Tris and phosphate buffer are regularly used in experimental investigations. These buffers might have radical scavenger properties toward different kinds of Reactive Oxygen Species (ROS) produced in solutions during chemical reactions like Fenton reaction and gamma radiolysis. Hydroxyl radicals (ºOH) are the most reactive and oxidizing agents having a great potential in oxidization of macromolecules like DNA and proteins. This <em>in vitro</em> study was aimed to evaluate radio-protective effects of Tris and phosphate buffer toward hydroxyl radicals generated by Fenton reaction. Hence, ºOH radicals were produced using a mixture of Hydrogen Peroxideand Ferrous Sulfate, called Fenton system. Human serum albumin (500µM) was prepared in Tris (10mM) and phosphate buffer (10mM), separately. These two samples were incubated with Fenton reaction (Ferrous Sulfate + Hydrogen peroxide) (10 mM) for 30 minutes and carbonyl groups were quantified by spectrophotometric carbonylation assay. The results of this study revealed the values of 1.04 ± 0.02 and 1.73 ± 0.03 for Tris and phosphate buffer treated samples, respectively. In conclusion, these findings confirmed that Tris buffer is a stronger radical scavenger toward ºOH radicals<strong> </strong>than phosphate buffer.<strong> </strong>

scholarly journals 2′,7′-dichlorofluorescin-based analysis of Fenton chemistry reveals auto-amplification of probe fluorescence and albumin as catalyst for the detection of hydrogen peroxide

2020 ◽  
Vol 477 (24) ◽  
pp. 4689-4710
Author(s):  
Teresa Gonzalez ◽  
Franck Peiretti ◽  
Catherine Defoort ◽  
Patrick Borel ◽  
Roland Govers
Keyword(s):  
Hydrogen Peroxide ◽  
Hydroxyl Radical ◽  
Hydroxyl Radicals ◽  
Ferrous Iron ◽  
Fenton Reaction ◽  
Iron Chelation ◽  
Intact Cells ◽  
Free System ◽  
Fenton Chemistry ◽  
In Cells

Fluorophore 2′,7′-dichlorofluorescin (DCF) is the most frequently used probe for measuring oxidative stress in cells, but many aspects of DCF remain to be revealed. Here, DCF was used to study the Fenton reaction in detail, which confirmed that in a cell-free system, the hydroxyl radical was easily measured by DCF, accompanied by the consumption of H2O2 and the conversion of ferrous iron into ferric iron. DCF fluorescence was more specific for hydroxyl radicals than the measurement of thiobarbituric acid (TBA)-reactive 2-deoxy-D-ribose degradation products, which also detected H2O2. As expected, hydroxyl radical-induced DCF fluorescence was inhibited by iron chelation, anti-oxidants, and hydroxyl radical scavengers and enhanced by low concentrations of ascorbate. Remarkably, due to DCF fluorescence auto-amplification, Fenton reaction-induced DCF fluorescence steadily increased in time even when all ferrous iron was oxidized. Surprisingly, the addition of bovine serum albumin rendered DCF sensitive to H2O2 as well. Within cells, DCF appeared not to react directly with H2O2 but indirect via the formation of hydroxyl radicals, since H2O2-induced cellular DCF fluorescence was fully abolished by iron chelation and hydroxyl radical scavenging. Iron chelation in H2O2-stimulated cells in which DCF fluorescence was already increasing did not abrogate further increases in fluorescence, suggesting DCF fluorescence auto-amplification in cells. Collectively, these data demonstrate that DCF is a very useful probe to detect hydroxyl radicals and hydrogen peroxide and to study Fenton chemistry, both in test tubes as well as in intact cells, and that fluorescence auto-amplification is an intrinsic property of DCF.

scholarly journals Identification of an oxygen-responsive element in the 5′-flanking sequence of the rat cytosolic phosphoenolpyruvate carboxykinase-1 gene, modulating its glucagon-dependent activation

1999 ◽  
Vol 339 (3) ◽  
pp. 563-569 ◽  
Author(s):  
Jutta BRATKE ◽  
Thomas KIETZMANN ◽  
Kurt JUNGERMANN
Keyword(s):  
Reporter Gene ◽  
Phosphoenolpyruvate Carboxykinase ◽  
Gene Activation ◽  
Simian Virus 40 ◽  
Rat Hepatocytes ◽  
Simian Virus ◽  
Flanking Sequence ◽  
Sv40 Promoter ◽  
Flanking Region ◽  
Luc Reporter Gene

The glucagon-stimulated transcription of the cytosolic phosphoenolpyruvate carboxykinase-1 (PCK1) gene is mediated by cAMP and positively modulated by oxygen in primary hepatocytes. Rat hepatocytes were transfected with constructs containing the first 2500, 493 or 281 bp of the PCK1 5ʹ-flanking region in front of the chloramphenicol acetyltransferase (CAT) reporter gene. With all three constructs glucagon induced CAT activity with decreasing efficiency maximally under arterial pO2 and to about 65% under venous pO2. Rat hepatocytes were then transfected with constructs containing the first 493 bp of the PCK1 5ʹ-flanking region in front of the luciferase (LUC) reporter gene, which were block-mutated at the CRE1 (cAMP-response element-1; -93/-86), putative CRE2 (-146/-139), promoter element (P) 1 (-118/-104), P2 (-193/-181) or P4 (-291/-273) sites. Glucagon induced LUC activity strongly when the P1 and P2 sites were mutated and weakly when the P4 site was mutated; induction of the P1, P2 and P4 mutants was positively modulated by the pO2. Glucagon also induced LUC activity strongly when the putative CRE2 site was altered; however, induction of the CRE2 mutant was not modulated by the pO2. Glucagon did not induce LUC activity when the CRE1 site was modified. These experiments suggested that the CRE1 but not the putative CRE2 was an essential site necessary for the cAMP-mediated PCK1 gene activation by glucagon and that the putative CRE2 site was involved in the oxygen-dependent modulation of PCK1 gene activation. To confirm these conclusions rat hepatocytes were transfected with simian virus 40 (SV40)-promoter-driven LUC-gene constructs containing three CRE1 sequences (-95/-84), three CRE2 sequences (-148/-137) or three CRE1 sequences plus two CRE2 sequences of the PCK1 gene in front of the SV40 promoter. Glucagon induced LUC activity markedly when the CRE1, but not when the CRE2, sites were in front of the SV40-LUC gene; however, induction of the (CRE1)3SV40-LUC constructs was not modulated by the pO2. Glucagon also induced LUC activity very strongly when the CRE1 and CRE2 sites were combined; induction of the (CRE1)3(CRE2)2SV40-LUC constructs was positively modulated by the pO2. These findings corroborated that sequences of the putative CRE2 site were responsible for the modulation by oxygen of the CRE1-dependent induction by glucagon of PCK1 gene transcription.

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