Involvement of glutathione peroxidase 1 in growth and peroxisome formation in Saccharomyces cerevisiae in oleic acid medium

Saccharomyces cerevisiae is able to use some fatty acids, such as oleic acid, as a sole source of carbon. β-oxidation, which occurs in a single membrane-enveloped organelle or peroxisome, is responsible for the assimilation of fatty acids. In S. cerevisiae, β-oxidation occurs only in peroxisomes, and H₂O₂ is generated during this fatty acid-metabolizing pathway. S. cerevisiae has three GPX genes (GPX1, GPX2, and GPX3) encoding atypical 2-Cys peroxiredoxins. Here we show that expression of GPX1 was induced in medium containing oleic acid as a carbon source in an Msn2/Msn4-dependent manner. We found that Gpx1 was located in the peroxisomal matrix. The peroxisomal Gpx1 showed peroxidase activity using thioredoxin or glutathione as a reducing power. Peroxisome biogenesis was induced when cells were cultured with oleic acid. Peroxisome biogenesis was impaired in gpx1? cells, and subsequently, the growth of gpx1? cells was lowered in oleic acid-containing medium. Gpx1 contains six cysteine residues. Of the cysteine-substituted mutants of Gpx1, Gpx1^C36S was not able to restore growth and peroxisome formation in oleic acid-containing medium, therefore, redox regulation of Gpx1 seems to be involved in the mechanism of peroxisome formation.     

Termination of a toxic Alexandrium bloom with hydrogen peroxide

The dinoflagellate Alexandrium ostenfeldii is a well-known harmful algal species that can potentially cause paralytic shellfish poisoning (PSP). Usually A. ostenfeldii occurs in low background concentrations only, but in August of 2012 an exceptionally dense bloom of more than 1 million cells L⁻¹ occurred in the brackish Ouwerkerkse Kreek in The Netherlands. The A. ostenfeldii bloom produced both saxitoxins and spirolides, and is held responsible for the death of a dog with a high saxitoxin stomach content. The Ouwerkerkse Kreek routinely discharges its water into the adjacent Oosterschelde estuary, and an immediate reduction of the bloom was required to avoid contamination of extensive shellfish grounds. Previously, treatment of infected waters with hydrogen peroxide (H₂O₂) successfully suppressed cyanobacterial blooms in lakes. Therefore, we adapted this treatment to eradicate the Alexandrium bloom using a three-step approach. First, we investigated the required H₂O₂ dosage in laboratory experiments with A. ostenfeldii. Second, we tested the method in a small, isolated canal adjacent to the Ouwerkerkse Kreek. Finally, we brought 50 mg L⁻¹ of H₂O₂ into the entire creek system with a special device, called a water harrow, for optimal dispersal of the added H₂O₂. Concentrations of both vegetative cells and pellicle cysts declined by 99.8% within 48 h, and PSP toxin concentrations in the water were reduced below local regulatory levels of 15 μg L⁻¹. Zooplankton were strongly affected by the H₂O₂ treatment, but impacts on macroinvertebrates and fish were minimal. A key advantage of this method is that the added H₂O₂ decays to water and oxygen within a few days, which enables rapid recovery of the system after the treatment. This is the first successful field application of H₂O₂ to suppress a marine harmful algal bloom, although Alexandrium spp. reoccurred at lower concentrations in the following year. The results show that H₂O₂ treatment provides an effective emergency management option to mitigate toxic Alexandrium blooms, especially when immediate action is required.     

Physiological parameters correlated with Tomato Mosaic Virus inducing defensive response in Datura metel

Programed cell death resembles a real nature active defense in Datura metel against TMV after three days of virus infection. This adaptive plant immune response was quantitatively assessed against Tomato Mosaic Virus infection by the following physiological markers; Chlorophyll-a (mg/g), Chlorophyll-b (mg/g), total protein (mg/g), hydrogen peroxide H₂O₂ (μmol/100 mg), DNA (μg/100 mg), RNA (μg/100 mg), Salicylic acid (μg/g), and Comet Assays. Parameters were assessed for asymptomatic healthy and symptomatic infected detached leaves. The results indicated H₂O₂ and Chlorophyll-a as the most potential parameters. Chlorophyll-a was considered the only significant predictor variant for the H₂O₂ dependent variant with a P value of 0.001 and R-square of 0.900. The plant immune response was measured within three days of virus infection using the cutoff value of H₂O₂ (⩽1.095 μmol/100 mg) and (⩽3.201 units) for the tail moment in the Comet Assay. Their percentage changes were 255.12% and 522.40% respectively which reflects the stress of virus infection in the plant. Moreover, H₂O₂ showed 100% specificity and sensitivity in the symptomatic infected group using the receiver-operating characteristic (ROC). All tested parameters in the symptomatic infected group had significant correlations with twenty-five positive and thirty-one negative correlations where the P value was <0.05 and 0.01. Chlorophyll-a parameter had a crucial role of highly significant correlation between total protein and salicylic acid. Contrarily, this correlation with tail moment unit was (r = −0.930, P < 0.01) where the P value was <0.01. The strongest significant negative correlation was between Chlorophyll-a and H₂O₂ at P < 0.01, while moderate negative significant correlation was seen for Chlorophyll-b where the P value < 0.05. The present study discloses the secret of the three days of rapid transient production of activated oxygen species (AOS) that was enough for having potential quantitative physiological parameters for defensive plant response toward the virus.     

Induction of heme oxygenase-1 by chamomile protects murine macrophages against oxidative stress

AimsProtection of cells from oxidative insult may be possible through direct scavenging of reactive oxygen species, or through stimulation of intracellular antioxidant defense mechanisms by induction of antioxidant gene expression. In this study we investigated the cytoprotective effect of chamomile and elucidated the underlying mechanisms.Main methodsThe cytoprotective effect of chamomile was examined on H₂O₂-induced cellular stress in RAW 264.7 murine macrophages.Key findingsRAW 264.7 murine macrophages treated with chamomile were protected from cell death caused by H₂O₂. Treatment with 50 μM H₂O₂ for 6 h caused significant increase in cellular stress accompanied by cell death in RAW 264.7 macrophages. Pretreatment with chamomile at 10–20 μg/mL for 16 h followed by H₂O₂ treatment protected the macrophages against cell death. Chamomile exposure significantly increased the expression of antioxidant enzymes viz. heme oxygenase-1 (HO-1), peroxiredoxin-1 (Prx-1), and thioredoxin-1 (Trx-1) in a dose-dependent manner, compared with their respective controls. Chamomile increased nuclear translocation of Nrf2 with increased phosphorylated Nrf2 levels, and binding to the antioxidant response element in the nucleus.SignificanceThese molecular findings for the first time provide insights into the mechanisms underlying the induction of phase 2 enzymes through the Keap1-Nrf2 signaling pathway by chamomile, and provide evidence that chamomile possesses antioxidant and cytoprotective properties.     

Antioxidant activity and protection of human umbilical vein endothelial cells from hydrogen peroxide-induced injury by DMC,a chalcone from buds of Cleistocalyx operculatus

The aim of this work was to study the antioxidant activity and the protective effect of 2′,4′-dihydroxy-6′-methoxy-3′,5′-dimethylchalcone (DMC), the main compound from the buds of Cleistocalyx operculatus, on human umbilical vein endothelial cells against cytotoxicity induced by H₂O₂. The antioxidant activities of DMC were measured by ABTS assay, ferric reducing antioxidant power (FRAP) and hydroxyl radical scavenging activity, and protective effects of DMC on human umbilical vein endothelial cells against cytotoxicity induced by H₂O₂ were tested. DMC was found to have high ABTS radical scavenging activity (176.5 ± 5.2 μmol trolox equivalents/500 μmol DMC) and strong ferric reducing antioxidant power (213.3 ± 5.8 μmol trolox equivalents/500 μmol DMC). In addition, DMC scavenged the hydroxyl radicals, with IC₅₀ values of 243.7 ± 6.3 μM, slightly lower than the reference antioxidant ascorbic acid (ASC). Moreover, DMC could protect the human umbilical vein endothelial cells against H₂O₂-induced cytotoxicity by decrease intracellular and extracellular ROS levels, reduction in catalase (CAT) activity and increment in malondialdehyde (MDA) level. These results suggested that DMC has the potential to be used in the therapy of oxidative damage.     

Phytochemical screening of flavonoids with their antioxidant activities from rapeseed (Brassica napus L.)

Ten flavone compounds, including three new flavonoid glycosides, were isolated from defatted rapeseed, and their protective antioxidant effect on H₂O₂-induced oxidative damage in human umbilical vein endothelial cells (ECV-304) was investigated. Three new flavonoid glycosides were identified as kaempferol-3-O-[(6-O-sinapoyl)-β-d-glucopyranosyl-(1 → 2)-β-d-glucopyranoside]-7-O-β-d-glucopyranoside (8), kaempferol-3,7-di-O-β-d-glucopyranoside-4'-O-(6-O-sinapoyl)-β-d-glucopyranoside (9), and kaempferol-3-O-[(3-O-sinapoyl)-β-d-glucopyranosyl-(1 → 2)-β-d-glucopyranoside]-7-O-β-d-glucopyranoside (10). The protective effects of all of the isolated compounds on H₂O₂-induced oxidative damage were assessed, and the activities of superoxide dismutase (SOD) and lactate dehydrogenase (LDH) were measured. All of compounds had a protective effect on H₂O₂-induced oxidative damage in ECV-304 cells and the presence of a substituted sinapoyl group and its position in the structures were used to elucidate the activity differences.     

Protective effects of purified safflower extract on myocardial ischemia in vivo and in vitro

Carthamus tinctorius L. (safflower) is one of the most commonly used Chinese herbal medicines to prevent and treat cardiac disease in clinical practice. However, the mechanisms responsible for such protective effects remain largely unknown. In this study, we investigated the anti-myocardial ischemia effects of a purified extract of C. tinctorius (ECT) both in vivo and in vitro. An animal model of myocardial ischemia injury was induced by left anterior descending coronary artery occlusion in adult rats. Pretreatment with ECT (100, 200, 400, 600 mg/kg body wt.) could protect the heart from ischemia injury by limiting infarct size and improving cardiac function. In the in vitro experiment, neonatal rat ventricular myocytes were incubated to test the direct cytoprotective effect of ECT against H₂O₂ exposure. Pretreatment with 100–400 μg/ml ECT prior to H₂O₂ exposure significantly increased cell viability as revealed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. ECT also markedly attenuated H₂O₂-induced cardiomyocyte apoptosis, as detected by Annexin V and PI double labeling with flow cytometry. The intracellular level of reactive oxygen species (ROS) was shown by 2′,7′-dichlorofluorescin diacetate (DCFH-DA), and ECT pretreatment significantly inhibited H₂O₂-induced ROS increase. We made a preliminary examination of the signaling cascade involved in ECT mediated anti-apoptotic effects. Phosphatidylinositol 3 kinase (PI3K) inhibitor (LY294002) blocked the cytoprotective effect conferred by ECT. Taken together, our findings provide the first evidence that the cardioprotective effects of ECT in myocardial ischemia operate partially through reducing oxidative stress induced damage and apoptosis. The protection is achieved by scavenging of ROS and mediating the PI3K signaling pathway.     

Impact of Moringa oleifera leaf extract in reducing the effect of lead acetate toxicity in mice

This study aimed to assess the impact of Moringa oleifera (M. oleifera) leaf extract against the poisoning of lead acetate; therefore, sixty mice were allocated into 4 groups with 15 in each, as G1) blank control, G2) supplied with 300 mg/kg body weight (BWT). M. oleifera extract, G3) supplied with 60 mg/kg BWT of lead acetate [Pb(C₂H₃O₂)₂], and G4) supplied with extract of M. oleifera + lead acetate. The liver enzymes were elevated post-treatment with Pb(C₂H₃O₂)₂, which then lowered to almost the normal level when M. oleifera was supplied to mice previously treated with Pb(C₂H₃O₂)₂. The values in (G3) decreased when compared with G1 (92.33 ± 12.99, 21.67 ± 2.91 and 98.00 ± 13.20 U/L, respectively. Also, the cholesterol and low-density lipoprotein levels were elevated post-supplementation with M. oleifera and Pb(C₂H₃O₂)₂. Pb(C₂H₃O₂)₂ improves the lipid profile, whereas M. oleifera pretreatment reduced cholesterol (CHOL), high density low cholesterol (HDL-c), and low-density low cholesterol (LDL-c) levels in animals fed Pb(C₂H₃O₂)₂. Pb(C₂H₃O₂)₂ elevates the total protein but lowers the total bilirubin and triglycerides post M. oleifera treatment and Pb(C₂H₃O₂)₂ when contrasted with G1. The protective effect of M. oleifera was caused by the fact that it lowered triglycerides (TG) and total bilirubin (TBIL) and raised total protein (TP). After administration of Pb(C₂H₃O₂)₂, the histological examination revealed alterations in the hepatocytes and kidneys of G3. Also, the liver and kidney cells in mice supplied with M. oleifera after Pb(C₂H₃O₂)₂ poisoning recovered. In conclusion, Pb is toxic, and the usage of M. oleifera partially enhances the negative impacts induced by Pb(C₂H₃O₂)₂.     

White shrimp Litopenaeus vannamei catalase: Gene structure,expression and activity under hypoxia and reoxygenation

Catalase (EC 1.11.1.6) is an antioxidant enzyme involved in redox equilibrium, regulating hydrogen peroxide (H₂O₂) concentration, a harmful reactive oxygen species (ROS) that is produced during hypoxia. Hypoxia occurs commonly in aquatic environments and in shrimp farms. We studied the catalase gene of the shrimp Litopenaeus vannamei and tested its expression and enzyme activity during hypoxia (1.5 mg/L O₂; 6 and 24 h) and reoxygenation (1 h after hypoxia). The complete gene is 2974 bp long and has four introns of 821, 223, 114 and 298 bp, respectively. The first intron has tree microsatellites, with GT and (T)AT(GT) repeated sequences. L. vannamei catalase is part of an invertebrate clade including crustaceans and rotifers. Catalase expression and activity is different in gills and hepatopancreas. Expression in gills increased 3.2 and 3-fold in response to hypoxia and reoxygenation (6 and 24 h hypoxia, followed by 1 h reoxygenation) compared to normoxia, while no differences were detected in the expression and activity in hepatopancreas. Catalase activity in gills had a contrary response to expression in hypoxia and reoxygenation.     

Synthesis,crystal structure and fluorescent characterization of a novel lanthanide tetraphosphonate with a layered structure

Hydrothermal reactions of Gd(III) nitrate with N,N,N′,N′-tetramethylenephosphono-1,4-diaminobutane, (H₂O₃PCH₂)₂N–(CH₂)₄–N(CH₂PO₃H₂)₂ (H₈L), afforded a novel Gd(III) phosphonate, namely, Gd[(O₃PCH₂)(HO₃PCH₂)N(H)(CH₂)₄N(H)(CH₂PO₃H)₂] · 2H₂O, [Gd(H₅L)] · 2H₂O. Its structure was established by a single-crystal X-ray diffraction study. In this compound, the Gd(III) ion is coordinated by eight phoshonate oxygen atoms from five different phosphonate groups, which belong to five different phosphonic ligands. Each Gd atom is connected with its neighboring Gd atoms by two phosphonate oxygens, forming a gadolinium phosphonate slab along the a-axis. Such slabs are bridged by tetraphosphonate H₅L anions, resulting in a 〈0 1 1〉 layer with the butane groups toward the interlayer space. These layers are further interlinked by strong hydrogen bonds formed by uncoordinated phosphonate oxygens into a 3D supermolecular structure. Luminescent studies indicate that this compound exhibits a broad blue fluorescent emission band at 441 nm.     

A dissection of the effects of ethylene,H2O2 and high irradiance on antioxidants and several genes associated with stress and senescence in tobacco leaves

Ethylene and hydrogen peroxide are involved in the modulation of stress responses in plants, but their interrelation is not well understood. This work was designed to find differences between the actions of ethylene and H₂O₂ on antioxidants and senescence markers. Leaves of Nicotiana tabacum were sprayed with H₂O₂ or with ethephon (precursor of ethylene). To find the possible modulation of responses to acute abiotic stress, ethephon- and H₂O₂-sprayed leaves were further subjected to high irradiance (HL). The application of H₂O₂ strongly stimulated ethylene synthesis (ACC). Ethylene and H₂O₂, as single factors, stimulated the trolox equivalent antioxidant capacity (TEAC) and the activity of catalase (CAT), in contrast to HL alone (stimulation of nonspecific peroxidases and the total glutathione pool). However, after combined treatments (ethylene + HL and H₂O₂ + HL), the stimulatory action of H₂O₂ was related to TEAC and CAT activity, while the application of ethylene stimulated the total glutathione pool. Hydrogen peroxide enhanced the expression of the three CAT genes (Cat1, Cat2 and Cat3), in contrast to ethylene (Cat2 and Cat3) and HL (Cat1). In regard to the markers of senescence and pathogenesis the most pronounced difference between the actions of ethylene and H₂O₂, as single factors, was related to NPR1, whereas when leaf spraying was combined with HL, differences were found at WRKY53 and PR1a. HL reversed the stimulatory effects of H₂O₂/ethylene-driven enhancements of the expression of several genes (Cat1, Cat2, NPR1, WRKY53). These results show that multiple stressors, as usually encountered by plants in nature, may largely change those expression patterns of genes determined in a single factor analysis. Moreover, the actions of HL (often considered the internal H₂O₂ trigger) and of exogenous H₂O₂ on gene expression are clearly different.     

Hydrothermal synthesis and structural characterization of an organic–inorganic hybrid material: (H2tptz)2[δ-Mo8O26]·2H2O (tptz=2,4,6-tripyridyltriazine)

The reaction of MoO₃ and 2,4,6-tripyridyltriazine (tptz) in water at 180°C for 48 h and pH 5.5 produces (H₂tptz)₂[Mo₈O₂₆]·2H₂O in 70% yield. The structure is constructed from δ-Mo₈O₂₆ ⁴⁻ clusters, H₂tptz²⁺ and H₃O⁺ cations linked through hydrogen bonding into a network. Crystal data: C₁₈H₁₆Mo₄N₆O₁₄; monoclinic P2₁/n; a=10.2225(5) Å, b=14.0072(6) Å, c=18.1154(8) Å, β=93.896(1)°, V=2587.9(2) Å³, Z=4, D_calc=2.372 g cm⁻³; R₁=0.0271 based on 3212 reflections.     

Enhanced exopolysaccharide production and biological activity of Lactobacillus rhamnosus ZY with calcium and hydrogen peroxide

Exopolysaccharides (EPS) are important food and drug additives with beneficial antioxidant, anticancer, and immune-related effects on human health. However, the EPS is limited by low yields and the need for complex culture conditions in fermentation. Here, we report that hydrogen peroxide and calcium stimulated probiotic activity and production of crude exopolysaccharide (c-EPS) by Lactobacillus rhamnosus ZY. Accordingly, supplementation with 3 mM H₂O₂ allowed c-EPS biosynthesis to reach 567 mg/L after 24 h. Addition of both CaCl₂ and H₂O₂ resulted in a c-EPS yield of 2498 mg/L after 12 h, over 9-fold higher than that of an anaerobic culture. We observed that exposure to calcium and hydrogen peroxide made the cells more hydrophobic and led to the over-expression of GroEL, NADH peroxidase, and glyceraldehyde 3-phosphate dehydrogenase, thus increasing energy storage and EPS production. Chromatographic analysis revealed c-EPS was composed mainly of mannose (5.1%), galactose (15.3%), glucose (20–30%), and rhamnose (50–60%). Preliminary in vitro tests revealed that H₂O₂ and CaCl₂ enhanced the 2,2-diphenyl-1-picrylhydrazyl and hydroxyl radical scavenging capacities, resulting in a notable protective effect against oxidative damage in NIH/3T3 cells. Our study provides a simple and cost-effective approach for achieving high yields of good quality EPS using Lactobacillus rhamnosus.     

Concerted involvement of cooperative proton–electron linkage and water production in the proton pump of cytochrome c oxidase

In cytochrome c oxidase, oxido-reductions of heme a/Cu_A and heme a₃/Cu_B are cooperatively linked to proton transfer at acid/base groups in the enzyme. H⁺/e⁻ cooperative linkage at Fe_a3/Cu_B is envisaged to be involved in proton pump mechanisms confined to the binuclear center. Models have also been proposed which involve a role in proton pumping of cooperative H⁺/e⁻ linkage at heme a (and Cu_A). Observations will be presented on: (i) proton consumption in the reduction of molecular oxygen to H₂O in soluble bovine heart cytochrome c oxidase; (ii) proton release/uptake associated with anaerobic oxidation/reduction of heme a/Cu_A and heme a₃/Cu_B in the soluble oxidase; (iii) H⁺ release in the external phase (i.e. H⁺ pumping) associated with the oxidative (R → O transition), reductive (O → R transition) and a full catalytic cycle (R → O → R transition) of membrane-reconstituted cytochrome c oxidase. A model is presented in which cooperative H⁺/e⁻ linkage at heme a/Cu_A and heme a₃/Cu_B with acid/base clusters, C₁ and C₂ respectively, and protonmotive steps of the reduction of O₂ to water are involved in proton pumping.     

Effect of acetone extract from stem bark of Acacia species (A. dealbata,A. ferruginea and A. leucophloea) on antioxidant enzymes status in hydrogen peroxide-induced HepG2 cells

Acacia species are multipurpose trees, widely used in the traditional systems of medicine to treat various ailments. The major objective of the present study was to determine the gene expression of enzymatic antioxidants by acetone extract from the stem bark of three Acacia species (Acacia dealbata, Acacia ferruginea and Acacia leucophloea) in hydrogen peroxide (H₂O₂)-induced human hepatoma (HepG2) cells. The expression of antioxidant enzymes such as superoxide dismutase containing copper–zinc (CuZnSOD)/manganese (MnSOD), catalase (CAT) and glutathione peroxidase (GPx) in HepG2 cells was evaluated by real-time PCR. The results of antioxidant enzyme expression in real-time PCR study revealed that the H₂O₂ (200 μM) challenged HepG2 cells reduced the expression of enzymes such as SOD, GPx and CAT. However, the cells pre-treated with acetone extracts of all the three Acacia species significantly (P > 0.05) up-regulated the expression of antioxidant enzymes in a concentration dependent manner (25, 50 and 75 μg/mL). In conclusion, the findings of our study demonstrated that the acetone extract of Acacia species effectively inhibited H₂O₂ mediated oxidative stress and may be useful as a therapeutic agent in preventing oxidative stress mediated diseases.     

Glucose-containing flavones—their synthesis and antioxidant and neuroprotective activities

Due to high reactivity, reactive oxygen species can attack biological molecules leading to cell or tissue injury. In this study, glucose moiety was attached at the C-7 position of quercetin 3-O-methyl ether (1) and luteolin (2) through glycosidic bond or ether linkage. The glucose-containing compounds showed potent DPPH and superoxide anion radical scavenging and lipid peroxidation inhibition activities and nearly equivalent protective actions to the parent aglycons against the H₂O₂-induced oxidative neuronal damage in primary cultured rat cortical cells. Among the compounds tested, 3b and 3c were the most potent (IC₅₀ values = 7.33 and 5.34 μM, respectively), exhibiting nearly equivalent actions to the parent compounds 1 and 2 (IC₅₀ = 3.50 and 3.75 μM, respectively).