Glutathione Reductase Gene Expression Research Articles

Montmorillonite-based edible coating enhances the postharvest quality of sweetpotato by regulating ROS and membrane lipid metabolism

The quality deterioration of sweetpotatoes easily during postharvest storage owing to their high respiration rate and accumulation of reactive oxygen species (ROS). In this study, an edible coating (MCW) assembled using montmorillonite, chitosan, and whey protein isolate was fabricated to improve the postharvest quality of sweetpotatoes, and the regulatory mechanism of the edible coating treatment on sweetpotato quality has investigated. The results confirmed that edible coating treatment had a positive effect on maintaining the appearance, firmness, colour, and sensory quality of sweetpotatoes, especially in maintaining free water, starch and soluble protein content, inhibiting respiration rate and delaying weight loss of sweetpotatoes stored at 25 °C for 30 d. Meanwhile, this study illuminated the MCW coating regulates ROS metabolism by upregulating superoxide dismutase, ascorbate peroxidase, and glutathione reductase gene expression, resulting in lower O2−. content in postharvest sweetpotato. Furthermore, the relative expression of the genes encoding lipoxygenase, phospholipase C, and phospholipase D was downregulated by the edible coating treatment and contributed to lower enzyme activity, electrolyte leakage and malondialdehyde content, which further reduced the degree of membrane lipid peroxidation and effectively maintained the structure of plasma membrane and then protected the cell integrity of sweetpotato. Collectively, this study provides useful information at the transcriptional level that MCW coating can enhance the post-harvest quality of sweetpotatoes by regulating ROS and membrane lipid metabolism.

Primer Design and Optimization of Annealing Temperature for Analysis of Glutathione Reductase Gene Expression in Rice (Oryza sativa L.)

Glutathione Reductase (GR) belongs to the NADPH-dependent flavoprotein oxidoreductase family and is found in both prokaryotes and eukaryotes. The GR gene is considered to play a key role in the elimination of oxidative reaction products by looking at the level of gene expression of GR rice in dealing with drought stress using qPCR. One of the important steps to develop a specific, effective and efficient qPCR is the primer design. Several studies analyzing GR gene expression in rice have also designed primers. However, the primer still lacks an ideal characteristic of primer, as it still has a secondary structure. This studies aims to design rice GR specific primers and optimize the annealing temperature for GR gene expression analysis on rice. Primers were designed using the Primer3 and Geneious Prime and checked for specificity using the Primer-BLAST tool. The selected primer pairs were then optimized for annealing temperature using gradient PCR. The best primer design results were GR-Forward 5’-ACGATTGCAGCCAGTGAAGA-3’ and GR-Reverse 5’-TGCGGCAATACTATCAACATCC-3’, with an amplicon length of 204 bp, primer base lengths of 20 and 22 nucleotides, Tm values of 60°C and 58.9°C, %GC of 50% and 45.5%, respectively. This primer pair had no secondary structure, both hairpin and self dimer. Gradient PCR showed the optimum annealing temperature for this primer pair was 52.2oC so that the primer can be used as a specific primer to analyze the GR gene expression in rice using qPCR.

Electroacupuncture promotes expression of glutathione related regulatory enzymes in ovary tissue of rats with diminished ovarian reserve

To observe the effect of electroacupuncture (EA) on ovarian function and expression of glutathione (GSH) related regulatory enzymes γ-glutamylcysteine synthetase (γ-GCS), glutathione reductase (GR) protein and gene in rats with diminished ovarian reserve (DOR), so as to explore its mechanisms underlying up-regulation of antioxidant stress ability. A total of 30 female SD rats with normal estrous cycle were randomly divided into blank control, model and EA groups, with 10 rats in each group. The DOR model was established by gavage of tripterygium wilfordii polyglycoside suspension (50 mg·kg-1·d-1) for 14 consecutive days, while the rats in the blank group were given equal volume of 0.9% sodium chloride solution. One hour after daily gavage, EA (1.0 mA, 100 Hz) was applied alternately to bilateral "Shenshu"(BL23), and "Zhongwan"(CV12)+"Guanyuan"(CV4) for 10 min, for 14 consecutive days. Estrous cycles of rats in each group were observed and recorded daily during intervention.After the intervention, H.E.staining was used to observe histopathological changes of the ovarian tissue. The contents of serum sex hormones ［follicle stimulating hormone (FSH), anti-mullerian hormone (AMH), estradiol (E2)］ and oxidative damage markers ［8-hydroxydeoxyguanosine (8-OHDG) and nitrotyrosine (NTY)］ were determined by ELISA. The contents of GSH and oxidized glutathione (GSSG) in the liver tissue were determined by colorimetry, and their ratios were calculated. Immunohistochemistry and real-time fluorescence quantitative PCR were used to detect the immunoactivity and gene expression levels of γ-GCS and GR in the ovarian tissues, respectively. Compared with the blank group, the model group had a marked increase in the disorder rate of estrous cycle, serum FSH, 8-OHDG and NTY contents (P<0.01) and a considerable decrease in the levels of serum AMH and E2, liver GSH and GSSG contents and GSH/GSSG ratio, ovarian optical density and cell number as well as the expression of γ-GCS and GR mRNAs (P<0.05, P<0.01). After EA intervention, the increase of the disorder rate of estrous cycle, serum FSH, 8-OHDG and NTY contents and the decrease of serum AMH and E2, liver GSH and GSSG contents and GSH/GSSG ratio, ovarian optical density and cell number of γ-GCS and GR as well as the expression of γ-GCS genes were all reversed (P<0.01, P<0.05). H.E. staining showed degenerative changes of the ovarian tissue, fewer follicles at every level and increase of atretic follicles, disarrangement and layer number decrease of granulosa cells, and atrophy of corpus luteum in the model group, which were relatively milder in the EA group. EA can improve ovarian function, and reduce oxidative stress damage in DOR rats, which may be associated with its functions in up-regulating the expression of γ-GCS and GR protein and gene in the ovarian tissue.

Anti-oxidant effect of nitrite in the pancreatic islets of type 2 diabetic male rats.

Nitrite, a nitric oxide (NO) donor, increases insulin secretion from pancreatic islets and has positive metabolic effects in type 2 diabetes (T2D). Here, we test the hypothesis of whether nitrite-induced insulin secretion is due to blunting of diabetes-induced oxidative stress in the islets. T2D was created in male rats using a combination of streptozotocin at 25 mg/kg and a high-fat diet. Wistar rats were assigned to 3 groups (n=6 in each group), including control, T2D, and T2D+nitrite; the latter group consumed drinking water containing sodium nitrite (50 mg/l) for eight weeks. At the end of the study, mRNA levels of NADPH oxidase (Nox1, 2, 3, and 4), superoxide dismutase (SOD1, 2, and 3), glutathione peroxides (GPX1 and 7), glutathione reductase (GR), catalase, thioredoxin (TXN1 and 2), and thioredoxin reductase (TXNRD1) were measured in the isolated pancreatic islets. In the islets of diabetic rats, mRNA expressions of Nox1, 2, and 4 were higher, whereas expressions of SOD1, 2, catalase, GPX1, 7, GR, and TXN1 were lower than controls. Nitrite significantly (all P-values<0.05) decreased gene expression of Nox1 (0.39-fold) and Nox4 (0.23-fold) and increased gene expression of SOD1 (2.2-fold), SOD2 (2.8-fold), catalase (2.7-fold), GPX1 (2.2-fold), GPX7 (6.0-fold), GR (3.0-fold), TXN1 (2.1-fold), and TXNRD1 (2.3-fold) in diabetic rats. Nitrite decreased oxidative stress in isolated pancreatic islets of rats with T2D by suppressing oxidants and augmenting anti-oxidants. These findings favor the notion that nitrite-induced insulin secretion is partially due to decreased oxidative stress.

Cornelian cherry (Cornus mas L.) fruit extract improves growth performance, disease resistance, and serum immune-and antioxidant-related gene expression of common carp (Cyprinus carpio)

This study evaluated the use of cornelian cherry (Cornus mas L.) fruit extract on the growth performance, immune responses, and disease resistance in common carp (Cyprinus carpio). A total of 192 fish (initial body weight of 18.34 ± 1.8 g) were randomly distributed in 12 tanks with 16 fish per tank (4 treatments with 3 replications) and fed with diets containing cornelian cherry extract (CCE11CCE (cornelian cherry extract)) at the rate of 0, 0.25, 0.5 and 1%. The final weight of common carp fed CCE at 0.5% was significantly higher than fish fed 0 and 0.25% doses (p < 0.05). In addition, weight gain and specific growth rate were increased, and feed conversion ratio decreased in fish-fed diets containing 0.5 and 1% CCE compared to other groups. The challenge test with A. hydrophila revealed that common carp-fed CCE had a lower mortality rate than the control. The lysozyme activity was markedly higher in common carp fed 0.5% of cornelian cherry extract than in control. The total immunoglobulin and total protein showed higher values in fish fed 0.5 and 1% than the control. The catalase gene showed higher mRNA levels in fish fed 0.5 and 1% than the control. Glutathione reductase gene expression was markedly upregulated in common carp fed 1% of CCE than the control and 0.25% groups. The glutathione S-transferase gene showed higher mRNA levels in fish fed 0.5 and 1% than the control and 0.25% groups. The superoxide dismutase gene expression was markedly higher in common carp fed 0.5% of CCE than in control. The CC-chemokine and interleukin-10 showed higher mRNA levels in fish fed 0.5% than the remaining groups. Hypoxia-inducible factor 1-alpha gene expression was markedly upregulated in common carp fed 1% of CCE than the control and 0.25% groups. The heat shock 70 kDa protein and interleukin-1 beta genes showed higher mRNA levels in fish fed 0.25, 0.5, and 1% than the control. In conclusion, the supplementation of CCE at 0.5–1% is recommended to enhance the growth performance and relieve the impacts of A. hydrophila infection in common carp.

Role of Alpha Lipoic Acid in Oxidant /Antioxidant Status and Gene Expression of Glutathione Reductase in Hydrogen Peroxide Exposed Rats: (Part -2)

This study was designated to evaluate the protective role of alpha lipoic acid against oxidative stress resulted by hydrogen peroxide on some oxidants/ antioxidants parameters and gene expression of glutathione peroxidase in adult Wistar rats. Forty adult male rats were randomly divided into four equal groups (10 rats /group) and were handled daily as follows for 56 days : Control group were intubated distal water and received ordinary tap water ; group T1 were intubated 60mg/kg B.W of alpha lipoic acid and received ordinary tap water ; group T2 were received hydrogen peroxide in tap water at concentration of 0.5% , while group T3 were intubated 60mg/kg B.W of alpha lipoic acid and received ordinary tap water containing 0.5% hydrogen peroxide. Fasting blood samples were collected at 0, 28 and 56 days of experimental periods for measurement of serum peroxynitrite and malondialdehyde concentrations, as well catalase activity. Furthermore, gene expression of glutathione reductase in liver was investigated. Administration of 0.5% hydrogen peroxide in drinking water (group T2) manifested a significant elevation in serum peroxynitrite and malondialdehyde with significant decrease in catalase and Glutathione, concentrations. Also, a significant decrease in gene expression of glutathione reductase was observed as compared to other treated groups. Nevertheless, rats in group T3 shows a significantly improvement in oxidant /antioxidant status with increase in folds changes of gene expression of glutathione reductase as compared to control and T2. In conclusion, supplementation of alpha lipoic acid to rats significantly reduced oxidative stress –induced by hydrogen peroxide and caused improvement of gene expression of glutathione reductase in liver via its antioxidant properties.

Spermidine application alleviates salinity damage to antioxidant enzyme activity and gene expression in alfalfa.

We investigated whether spermidine (Spd) application alleviates salinity-induced damage in alfalfa (Medicago sativa L), and explored defence mechanisms associated with stress-related ion balance, antioxidant metabolism, and gene expression. We examined the response of 30-day-old alfalfa maintained in hydroponic culture tests for 7 days and subjected to one of six treatments: half-strength Hoagland solution (control); 1% NaCl; 10 μM Spd + 1% NaCl; 20 μM Spd + 1% NaCl; 40 μM Spd + 1% NaCl; and 60 μM Spd + 1% NaCl. In salinity-stressed plants, chlorophyll b, chlorophyll a + b, and total protein showed significant decreases, while marked increases were detected in relative electrolyte leakage, H2O2 content, glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), glutathione reductase (GR) activity, the Na+/K+ ratio, and APX1, APX2, GR, and SOD gene expression levels. Chlorophyll a and total protein content markedly increased under exogenous application of 20 μM Spd, while H2O2 content, GSH, SOD, CAT, POD, GR activity, the Na+/K+ ratio, and APX2, GR, and SOD expression levels all decreased. These results indicated that exogenous application of 20 μM spermidine effectively alleviates salinity-induced damage in alfalfa. These findings could benefit alfalfa cultivation and promote the development and utilization of saline-alkali soil.

Castasterone confers copper stress tolerance by regulating antioxidant enzyme responses, antioxidants, and amino acid balance in B. juncea seedlings

The aim of the present study was to explore the effect of exogenous application of castasterone (CS) on physiologic and biochemical responses in Brassica juncea seedlings under copper (Cu) stress. Seeds were pre-soaked in different concentrations of CS and grown for 7 days under various levels of Cu. The exposure of B. juncea to higher levels of Cu led to decrease of morphologic parameters, with partial recovery of length and fresh weight in the CS pre-treated seedlings. Metal content was high in both roots and shoots under Cu exposure while the CS pre-treatment reduced the metal uptake. Accumulation of hydrogen peroxide (H2O2) and superoxide anion radical (O2-) were chosen as stress biomarker and higher levels of H2O2 (88.89%) and O2- (62.11%) showed the oxidative stress in metal treated B. juncea seedlings, however, CS pre-treatment reduced ROS accumulation in Cu-exposed seedlings. The Cu exposures lead to enhance the plant's enzymatic and non-enzymatic antioxidant system. It was observed that enzymatic activities of ascorbate peroxidase (APOX), dehydroascorbate reductase (DHAR), and glutathione reductase (GR), glutathione perxoidase (GPOX) and gultrathione-s-transferase increased while activity of monodehydroascorbate reductase (MDHAR) decreased under Cu stress. The pre-treatment with CS positively affected the activities of enzymes. RT-PCR analysis showed that mRNA transcript levels were correlated with total enzymatic activity of DHAR, GR, GST and GSH. Increase in the gene expression of DHAR (1.85 folds), GR (3.24 folds), GST-1 (2.00 folds) and GSH-S (3.18 folds) was noticed with CS pre-treatment. Overall, the present study shows that Cu exposure induced severe oxidative stress in B. juncea plants and exogenous application of CS improved antioxidative defense system by modulating the ascorbate-glutathione cycle and amino acid metabolism.

GR1-like gene expression in Lycium chinense was regulated by cadmium-induced endogenous jasmonic acids accumulation.

The G1-like gene from the Lycium chinensewas cloned and transferred into N. tabacum. Evidence showed that endogenous JA accumulation was crucial to LcGRgene expression in cadmium-stressed L. chinense. Glutathione reductase (GR) plays a vital role in glutathione-ascorbate metabolism and is a key enzyme in maintaining the redox state in plants. Jasmonic acids (JA) are important hormones regulating protective responses against bacteria and mechanic damage in plants. At present, the relationship between the endogenous JA accumulation, the glutathione (GSH) content and GR gene expression in plants under cadmium (Cd) stress has not been elucidated. This study primarily aims to explore their interconnected relations. First, we isolated theGR1-like gene fromLycium chinense (LcGR). Real-time PCR showed that gene LcGR and allene oxide cyclase (LcAOC) (a JA synthesis gene) expression in L. chinense plants was significantly enhanced by CdCl2 and reduced by CdCl2 cotreatment with 12,13-epoxy-octadecenoic acid (EOA), a JA synthesis inhibitor. Meanwhile, the JA content in plants strongly increased under Cd stress and decreased under Cd+EOA treatment, which was in accordance with expression pattern of LcAOC. The function of gene LcGR was confirmed in vitro with E. coli expression system. The subcellular localization in chloroplasts of LcGR gene was proved in Nicotiana tabacum leaves with transient transfection system of Agrobacterium tumefaciens. Furthermore, the overexpression of gene LcGR in the transgenic tabacum led to great Cd-tolerance and higher GSH accumulation. Overall, the results showed that the endogenous JA accumulation in Cd-stressed plants affects the GR expression which is crucial to the GSH accumulation and GSH-dependent tolerance to cadmium in LcGR transformants.

Beneficial Rhizobacterium Bacillus amyloliquefaciens SQR9 Induces Plant Salt Tolerance through Spermidine Production.

The inoculation of plants with plant-growth-promoting rhizobacterium has been an effective strategy for enhancing plant salt tolerance to diminish the loss of agricultural productivity caused by salt stress; however, the signal transmitted from bacteria to the plant under salt stress is poorly understood. In this study, the salt tolerance of Arabidopsis thaliana and Zea mays was enhanced by inoculation with Bacillus amyloliquefaciens SQR9. Using dialysis bags with different molecular weight cutoffs, we sorted through the molecules secreted by SQR9 and found that spermidine is responsible for enhancing plant salt tolerance. An SQR9 ΔspeB mutant deficient in spermidine production failed to induce plant salt tolerance. However, the induction of plant salt tolerance was disrupted by mutating genes involved in reduced glutathione (GSH) biosynthesis and the salt overly sensitive pathway in Arabidopsis. Using quantitative real-time polymerase chain reaction, this study demonstrated that spermidine produced by SQR9 leads to increased glutamine synthetase and glutathione reductase gene expression, leading to increased levels of GSH, which is critical for scavenging reactive oxygen species. SQR9-derived spermidine also upregulates the expression of NHX1 and NHX7, which sequesters Na+ into vacuoles and expels Na+ from the cell, thereby reducing ion toxicity.

Transcriptomic responses of selected genes against chromium stress in Arundo donax L.

ABSTRACTRecently, giant reed (Arundo donax L.) has emerged as a useful bioresource capable of coping under harsh abiotic stress conditions produced by exposure to heavy metals, drought and salinity as evidenced by significant biomass production. The current study was undertaken to determine the expression of critical genes involved in resisting oxidative stress and maintaining normal cellular functions. The critical genes examined included glutathione reductase (GR) that prevents oxidative damage of membranes, carotenoid hydroxylase (CR), transcription factor (bHLH) and amidase involved in biosynthesis of indole-3-acetic acid following exposure to chromium (Cr) stress at concentrations of 0, 33.5, 67, 133 or 268 mg/L in giant reed. The S19 gene was used as internal control. It was not possible to extract RNA from severely stressed giant plant leaves at 133 and 268 mg/L Cr. Therefore, this investigation focused on gene expression in the presence of at 0, 33.5 or 67 mg/L Cr. Following Cr-mediated stress, gene expression of GR and CR was increased at 33.5 mg/L but reduced at 67 mg/L. The transcription factor, bHLH, gene expression remained unchanged. Amidase gene expression was elevated at 67 mg/L Cr. Data suggested that over expression of amidase and GR genes at high Cr exposure might increase growth and biomass of giant reed as well as enhance tolerance. These results indicated that giant reed tolerated the presence of Cr stress due to over expression of amidase and GR genes. It would appear that giant reed may serve a beneficial role for phytoremediation of Cr contaminated wastewater and soils at concentrations below 100 mg/kg.

Arginase activity in Arabidopsis thaliana infected with Heterodera schachtii

The activity of arginase (ARGAH), which results in ornithine and urea production, is important for nitrogen metabolism in all organisms as well as for defence responses. The second‐stage juveniles of the cyst‐forming nematode Heterodera schachtii penetrate roots and induce the formation of a permanent feeding site. To determine whether infection with H. schachtii causes the induction of arginase, the expression was studied in Arabidopsis roots and shoots at the day of inoculation and at 3, 7 and 15 days post‐inoculation (dpi). Parasite infection caused a strong decrease of root arginase activity and ARGAH1 gene expression, which persisted over the entire examination period. In shoots, the mRNA expression of ARGAH2 increased at 3 and 7 dpi, but the enzymatic activity was significantly enhanced only at 3 dpi. Thus, while arginase down‐regulation occurs in roots, which is apparently due to the presence of nematode effectors, in shoots the activity is only transiently up‐regulated despite persistently high gene expression. As oxidative stress is possible during nematode infection, the activity and gene expression of glutathione reductase, a marker of the redox equilibrium, were estimated and found to be significantly enhanced at 7 dpi in shoots of infected plants. The level of proline, an amino acid known for its ability to scavenge free radicals, was increased 60‐fold. The results suggest that the disruption of redox homeostasis, as reflected by increased proline level and glutathione reductase expression and activity, accompanies changes in arginine metabolism in the shoots, indicating systemic changes induced by nematode infection.

Effect of Zinc nanoparticles on oxidative stress-related genes and antioxidant enzymes activity in the brain of Oreochromis niloticus and Tilapia zillii

This study was carried out to determine the median lethal concentrations (LC50) of Zinc nanoparticles (ZnNPs) on Oreochromis niloticus and Tilapia zillii. The biochemical and molecular potential effects of ZnNPs (500 and 2000 μg L−1) on the antioxidant system in the brain tissue of O. niloticus and T. zillii were investigated. Four hundred fish were used for acute and sub-acute studies. ZnNP LC50 concentrations were investigated in O. niloticus and T. zillii. The effect of 500 and 2000 μg L−1 ZnNPs on brain antioxidants of O. niloticus and T. zillii was investigated. The result indicated that 69 h LC50 was 5.5 ± 0.6 and 5.6 ± 0.4 for O. nilotica and T. zillii, respectively. Fish exposed to 500 μg L−1 ZnNPs showed a significant increase in reduced glutathione (GSH), total glutathione (tGSH) levels, superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx) and glutathione-S-transferase (GST) activity and gene expression. On the contrary, malondialdehyde (MDA) levels significantly decreased. Meanwhile, fish exposed to 2000 μg L−1 ZnNPs showed a significant decrease of GSH, tGSH levels, SOD, CAT, GR, GPx and GST activity and gene expression. On the contrary, MDA levels significantly increased. It was concluded that, the 96 h LC50 of ZnNPs was 5.5 ± 0.6 and 5.6 ± 0.4 for O. nilotica and T. zillii, respectively. ZnNPs in exposure concentrations of 2000 μg/L induced a deleterious effect on the brain antioxidant system of O. nilotica and T. zillii. In contrast, ZnNPs in exposure concentrations of 500 μg L−1 produced an inductive effect on the brain antioxidant system of O. nilotica and T. zillii.

Amelioration of postharvest chilling injury in sweet pepper by glycine betaine

The reduction in the quality of harvested green peppers due to physiological disorders resulting from chilling injury (CI) results in significant economic losses. In the current study, the ability of glycine betaine (GB, a stress-ameliorating compound) to reduce CI in sweet pepper was investigated. GB at 1mmolL−1 significantly reduced CI in sweet pepper fruit during a sixteen-day period of storage at 3°C followed by an additional 3 days at 20°C. CI index, membrane permeability, chlorophyll content, ascorbic acid (ASA), and malondialdehyde (MDA) were assessed. The activities of a variety of antioxidant enzymes, including peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX), glutathione reductase (GR), and lipoxygenase (LOX) were determined along with transcript levels. Results indicated that amelioration of CI in sweet pepper by GB was associated with a reduction in cellular leakage, MDA content, and lipid peroxidation in sweet pepper. Both gene expression and enzyme activity of POD, CAT, APX, and GR in GB-treated fruit were higher than levels in control fruit. These data suggest that GB increases chilling tolerance in pepper fruit by inducing antioxidant gene expression and enzyme activity, thus alleviating the potential injury resulting from CI.

Processed Panax ginseng, Sun Ginseng, Decreases Oxidative Damage Induced by tert-butyl Hydroperoxide via Regulation of Antioxidant Enzyme and Anti-apoptotic Molecules in HepG2 Cells.

Potential antioxidant effect of processed ginseng (sun ginseng, SG) on oxidative stress generated by tert-butyl hydroperoxide (t-BHP) was investigated in HepG2 cells. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and lactate dehydrogenase (LDH) leakage test demonstrated that SG dose-dependently prevents a loss of cell viability against t-BHP-induced oxidative stress. Also, SG treatment dose-dependently relieved the increment of activities of hepatic enzymes, such as aspartate aminotrasferase and alanine aminotransferase, and lipid peroxidation mediated by t-BHP treatment in HepG2 cells. SG increased the gene expression of antioxidant enzymes such as superoxide dismutase, catalase, and glutathione peroxidase. However, high dose of SG treatment caused decrease in mRNA level of glutathione peroxidase as compared to low dosage of SG-treated cells. The gene expression of glutathione reductase was found to be slightly increased by SG treatment. In addition, SG extract attributed its hepaprotective effect by inducing the mRNA level of bcl-2 and bcl-xL but reducing that of bax. But, the gene expression of bad showed no significant change in SG-treated HepG2 cells. These findings suggest that SG has hepatoprotective effect by showing reduction of LDH release, activities of hepatic enzymes and lipid peroxidation and regulating the gene expression of antioxidant enzymes and apoptosis-related molecules against oxdative stress caused by t-BHP in HepG2 cells.

Effects of 5-aminolevulinic acid on the H&lt;sub&gt;2&lt;/sub&gt;O&lt;sub&gt;2&lt;/sub&gt;-content and antioxidative enzyme gene expression in NaCl-treated cucumber seedlings

The potential of 5-aminolevulenic acid (ALA) to enhance the salt tolerance of cucumber (Cucumis sativus L.) seedlings was investigated. ALA was applied at various concentrations (0, 1, 10, 25, 50, and 100 mg dm-3) as foliar spray or root watering. Then the seedlings were exposed to 0 or 75 mM NaCl for 5 d. NaCl stress reduced the root and leaf dry masses, leaf area, and the leaf net CO2 assimilation rate. These reductions were counteracted by exogenous ALA, and the most efficient was 50 mg dm-3 concentration via foliar spray. ALA decreased the H2O2 contents and increased the activities of ascorbate peroxidase (APX) and glutathione reductase (GR) in NaCl-treated cucumber roots and leaves and the activity of catalase (CAT) in leaves. The ALA application also up-regulated the expressions of CAT and cAPX genes in roots and leaves and the expression of GR gene in roots of the NaCl treated cucumber plants.

Protective Role of Melatonin on Oxidative Stress Status and RNA Expression in Cerebral Cortex and Cerebellum of AβPP Transgenic Mice After Chronic Exposure to Aluminum

Aluminum (Al) has been associated with pro-oxidant effects, as well as with various serious neurodegenerative diseases such as Alzheimer's disease (AD). On the other hand, melatonin (Mel) is a known antioxidant, which can directly act as free radical scavenger, or indirectly by inducing the expression of some genes linked to the antioxidant defense. In this study, 5-month-old AssPP female transgenic (Tg2576) (Tg) and wild-type mice were fed with Al lactate supplemented in the diet (1 mg Al/g diet). Concurrently, animals received oral Mel (10 mg/kg) until the end of the study at 11 months of age. Four treatment groups were included for both Tg and wild-type mice: control, Al only, Mel only, and Al + Mel. At the end of the treatment period, cortex and cerebellum were removed and processed to examine the following oxidative stress markers: reduced glutathione, oxidized glutathione, cytosolic Cu-Zn superoxide dismutase (SOD1), glutathione reductase (GR), glutathione peroxidase, catalase (CAT), and thiobarbituric acid reactive substances. Moreover, the gene expression of SOD1, GR, and CAT was evaluated by real-time RT-PCR. The biochemical changes observed in cortex and cerebellum suggest that Al acted as a pro-oxidant agent. Melatonin exerted an antioxidant action by increasing the mRNA levels of the enzymes SOD1, CAT, and GR evaluated in presence of Al and Mel, independently on the animal model.

Oxidative stress status and RNA expression in hippocampus of an animal model of Alzheimer's disease after chronic exposure to aluminum

It is well established that aluminum (Al) is a neurotoxic agent that induces the production of free radicals in brain. Accumulation of free radicals may cause degenerative events of aging such as Alzheimer's disease. On the other hand, melatonin (Mel) is a known antioxidant, which can directly act as free radical scavenger, or indirectly by inducing the expression of some genes linked to the antioxidant defense. In this study, AbetaPP female transgenic (Tg2576) (Tg) and wild-type mice (5 months of age) were fed with Al lactate supplemented in the diet (1 mg Al/g diet). Simultaneously, animals received oral Mel (10 mg/kg) dissolved in tap water until the end of the study at 11 months of age. Four treatment groups were included for both Tg and wild-type mice: control, Al only, Mel only, and Al+Mel. At the end of the period of treatment, hippocampus was removed and processed to examine the following oxidative stress markers: reduced glutathione (GSH), oxidized glutathione (GSSG), superoxide dismutase (SOD), glutathione reductase (GR), glutathione peroxidase (GPx), catalase (CAT), and thiobarbituric acid reactive substances (TBARS). Moreover, the gene expression of Cu-ZnSOD, GR, and CAT was evaluated by real-time RT-PCR. Aluminum concentration in hippocampus was also determined. The biochemical changes observed in this tissue suggest that Al acts as a pro-oxidant agent. Melatonin exerts an antioxidant action by increasing the mRNA levels of the antioxidant enzymes SOD, CAT, and GR evaluated in presence of Al and Mel, with independence of the animal model.

(h)GR, beans and drought stress

Reactive oxygen species (ROS) are commonly found in plants as natural by-products of the metabolism but their production is greatly enhanced under abiotic stresses. Particular metabolites and enzymes belonging to the ascorbate-glutathione cycle are able to scavenge these deleterious molecules and modulate the cellular redox-status. In the March issue of Journal of Plant Physiology, we have shown that drought stress induces a raise in glutathione reductase (GR) activity and gene expression that could be related to the intensity of the drought treatment and the drought susceptibility of the bean cultivar (cowpea and/or common bean). In the present addendum we show new data on GR specific activity during progressive drought stress and recovery of the drought-susceptible bean cultivar which can be related to the previously found dual-targeted GR gene expression. Furthermore, since in leguminous plants homoglutathione (hGSH) is generally the most abundant low molecular weight thiol form, we discuss on the occurrence of a (homo)glutathione reductase activity in beans.Addendum to: Torres-Franklin ML, Contour-Ansel D, Zuily-Fodil Y, Pham-Thi AT. Molecular cloning of glutathione reductase cDNAs and analysis of GR gene expression in cowpea and common bean leaves during recovery from moderate drought stress. J Plant Physiol 2008; 165:514-21.

Glutathione Reductase in Leaves of Cowpea: Cloning of Two cDNAs, Expression and Enzymatic Activity under Progressive Drought Stress, Desiccation and Abscisic Acid Treatment

Reactive oxygen species are frequently produced when plants are exposed to abiotic stresses. Among the detoxication systems, two enzymes, ascorbate peroxidase and glutathione reductase (GR) play key roles. GR has also a central role in keeping the reduced glutathione pool during stress thus allowing the adjustments on the cellular redox reactions. The aim of this work was to study the variations in cytosolic and dual-targeted GR gene expression in the leaves of cowpea plants submitted to progressive drought, rapid desiccation and application of exogenous abscisic acid (ABA). Two cowpea (Vigna unguiculata) cultivars, one drought-resistant ('EPACE-1'), the other drought-sensitive ('1183') were submitted to progressive drought stress by withholding irrigation. Cut-off leaves were air-dried or treated with exogenous ABA. Two GR cDNAs, one cytosolic, the other dual-targeted to chloroplasts and mitochondria were isolated by PCR and cloned in plasmid vectors. Reverse-transcription PCR was used to study the variations in GR gene expression. Two new cDNAs encoding a putative dual-targeted and a cytosolic GR were cloned and sequenced from leaves of V. unguiculata. Drought stress induced an up-regulation of the expression of the cytosolic GR gene directly related to the intensity of the stress in both cultivars. The expression of dual-targeted GR was up-regulated by the drought treatment in the susceptible cultivar only. Under a fast desiccation, the '1183' cultivar responded later than the 'EPACE-1', although in 'EPACE-1' it was the cytosolic isoform which responded and in '1183' the dual-targeted one. Exogenous ABA enhanced significantly the activity and expression levels of GR in both cultivars after treatment for 24 h. These results demonstrate a noticeable activation in both cultivars of the antioxidant metabolism under a progressive water stress, which involves both GR genes in the case of the susceptible cultivar. Under a fast desiccation, the susceptible cultivar responded later than the resistant one, suggesting a weaker capacity of response versus the resistant one. Exogenous ABA probably acts on GR gene expression via a mediated signal transduction pathway.