γ-aminobutyric acid (GABA) confers chromium stress tolerance in <Emphasis Type="Italic">Brassica juncea</Emphasis> L. by modulating the antioxidant defense and glyoxalase systems

Chromium (Cr) toxicity is hazardous to the seed germination, growth, and development of plants. γ-aminobutyric acid (GABA) is a non-protein amino acid and is involved in stress tolerance in plants. To investigate the effects of GABA in alleviating Cr toxicity, we treated eight-d-old mustard (Brassica juncea L.) seedlings with Cr (0.15 and 0.3?mM K₂CrO₄, 5 days) alone and in combination with GABA (125?µM) in a semi-hydroponic medium. The roots and shoots of the seedlings accumulated Cr in a dose-dependent manner, which led to an increase in oxidative damage [lipid peroxidation; hydrogen peroxide (H₂O₂) content; superoxide (O₂ ^??) generation; lipoxygenase (LOX) activity], methylglyoxal (MG) content, and disrupted antioxidant defense and glyoxalase systems. Chromium stress also reduced growth, leaf relative water content (RWC), and chlorophyll (chl) content but increased phytochelatin (PC) and proline (Pro) content. Furthermore, supplementing the Cr-treated seedlings with GABA reduced Cr uptake and upregulated the non-enzymatic antioxidants (ascorbate, AsA; glutathione, GSH) and the activities of the enzymatic antioxidants including ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), glutathione reductase (GR), glutathione peroxidase (GPX), superoxide dismutase (SOD), catalase (CAT), glyoxalase I (Gly I), and glyoxalase II (Gly II), and finally reduced oxidative damage. Adding GABA also increased leaf RWC and chl content, decreased Pro and PC content, and restored plant growth. These findings shed light on the effect of GABA in improving the physiological mechanisms of mustard seedlings in response to Cr stress.     

Polyamines-induced aluminum tolerance in mung bean: A study on antioxidant defense and methylglyoxal detoxification systems

We investigated the roles of exogenously applied Spd (0.3?mM spermidine) in alleviating Al (AlCl₃, 0.5?mM, 48 and 72?h)- induced injury in mung bean seedlings (Vigna radiata L. cv. BARI Mung-2). Aluminum toxicity induced oxidative damage overproducing reactive oxygen species (ROS; H₂O₂ and O₂ ^??), increasing lipoxygenase activity and membrane lipid peroxidation. The toxic compound methylglyoxal (MG) also overproduced under Al stress. In order to circumvent Al-induced oxidative stress, enzymatic and non-enzymatic antioxidant defense were activated by the application of exogenous Spd. Exogenous Spd increased ascorbate (AsA) and glutathione (GSH) content, AsA/dehydroascorbate (DHA) ratio, GSH/ glutathione disulfide (GSSG) ratio, activity of ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR), glutathione reductase (GR) and catalase (CAT) which reduced ROS production and oxidative stress under Al stress. Spd-induced improvement of GSH pool and Gly II activity alleviated injurious effects of MG. Exogenous Spd positively modulated the endogenous PAs level. Regulating the osmoprotectant molecule (proline), Spd improved plant water status under Al stress. Exogenous Spd was potent to prevent breakdown of Al-induced photosynthetic pigment and to improve growth performances under Al stress. The mechanism by which Spd enhances antioxidant and glyoxalase components might be studied extensively. Spermidine-induced protection of photosynthetic pigment from damages and growth enhancement were remarkable and recommended for further detailed study to understand the mechanism.     

Salicylic acid alleviates copper toxicity in rice (Oryza sativa L.) seedlings by up-regulating antioxidative and glyoxalase systems

The present study investigated the effect of salicylic acid (SA) on toxic symptoms, lipid peroxidation, reactive oxygen species generation and responses of antioxidative and glyoxalase systems in rice seedlings grown hydroponically under copper (Cu) stress for 48 h. Exposures of 75 and 150 μM Cu²⁺ caused toxicity symptoms (chlorosis, necrosis and rolling in leaves), sharp increases in malondialdehyde (MDA), hydrogen peroxide (H₂O₂) contents and lipoxygenase (LOX) activity with concomitant reductions of chlorophyll (Chl) and relative water content (RWC). Both levels of Cu decreased ascorbic acid (AsA), glutathione (GSH), non-protein thiol (NPT) and proline contents in roots but rather increased in leaves except that AsA decreased in leaves too. These results together with overaccumulation of superoxide (O ₂ ^?? ) and H₂O₂ in leaves revealed that Cu exposures induced oxidative stress. Contrary, SA-pretreatment (100 μM for 24 h) reduced toxicity symptoms and diminished Cu-induced increases in LOX activity, H₂O₂, MDA and proline contents while the levels of RWC, Chl, AsA and redox ratios were elevated. Higher levels of GSH and NPT were also observed in roots of SA-pretreated Cu-exposed seedlings. SA-pretreatment also exerted its beneficial role by inhibiting the Cu upward process. Studies on antioxidant enzymes showed that SA further enhanced the activities of superoxide dismutase, ascorbate peroxidase, glutathione reductase and glutathione peroxidase, and also elevated the depressed activities of catalase, dehydroascorbate reductase and glutathione S-transferase particularly at 150 μM Cu²⁺ stress. In addition, the activity of glyoxalase system (glyoxalase I and II) was further elevated by SA pretreatment in the Cu-exposed seedlings. These results concluded that SA-mediated retention of Cu in roots and enhanced capacity of both antioxidative and glyoxalase systems might be associated with the alleviation of Cu-toxicity in rice seedlings.     

DRDE-07 and its analogues as promising cytoprotectants to nitrogen mustard (HN-2)—An alkylating anticancer and chemical warfare agent

Nitrogen mustard (HN-2), also known as mechlorethamine, is an alkylating anticancer agent as well as blister inducing chemical warfare agent. We evaluated the cytoprotective efficacy of amifostine, DRDE-07 and their analogues, and other antidotes of mustard agents against HN-2. Administration of 1 LD₅₀ of HN-2 (20 mg/kg) percutaneously, decreased WBC count from 24 h onwards. Liver glutathione (GSH) level decreased prominently and the maximum depletion was observed on 7th day post-HN-2 administration. Oxidised glutathione (GSSG) level increased significantly at 24 h post-administration and subsequently showed a progressive decrease. Hepatic malondialdehyde (MDA) level and percent DNA damage increased progressively following HN-2 administration. The spleen weight decreased progressively and reached a minimum on 3–4 days with subsequent increase. The antidotes were administered repeatedly for 4 and 8 days after percutaneous administration of single sublethal dose (0.5 and 0.25 LD₅₀) of HN-2. Treatment with DRDE-07, DRDE-30 and DRDE-35 significantly protected the changes in spleen weight, WBC count, GSH, GSSG, MDA and DNA damage following HN-2 administration (0.5 and 0.25 LD₅₀). There was no alteration in the transaminases (AST and ALT), and alkaline phosphatase (ALP) activities, neither with HN-2 nor with antidotes. The present study shows that HN-2 is highly toxic by percutaneous route and DRDE-07, DRDE-30 and DRDE-35 can partially protect it.     

Taurodeoxycholate-induced intestinal injury is modulated by oxidative stress-dependent pre-conditioning like mechanisms

Mechanisms by which hydrophobic bile salts cause tissue changes below their critical micellar concentration (CMC, 1-2mM) and above (4-8mM) remain poorly understood. In this study, rat colonic mucosa was exposed to different concentrations of taurodeoxycholate (TDC), t-butyl-hydroperoxide (t-BH) or glutathione ester with or without pre-incubation with 2mM TDC. Exposure to 2mM TDC was associated with 10% higher tissue levels of total glutathione (GSH, basal values: 33.7+/-3.3 nmol/mg prot). With TDC 8mM, GSH decreased to 16.4+/-2.3 nmol/mg prot (P<0.05), oxidized glutathione (GSSG) increased by 60% (P<0.05), glutathione peroxidase (GSH-Px) and reductase activities were threefold increased, protein carbonyls fourfold increased, protein sulfhydrils decreased by 78%, lactate dehydrogenase (LDH) and GSSG release in the incubation medium were sixfold higher. In 2mM TDC pre-treated tissues, the subsequent incubation with 8mM TDC induced a lower loss of tissue GSH, and a lower release of LDH and GSSG. Pre-incubation with 2mM TDC partly protected against t-BH toxicity, while glutathione ester protected against 8mM TDC toxicity. In conclusion, TDC exposure causes opposite effects depending on CMC: induction of antioxidant protective systems including glutathione system (pre-conditioning effect) was observed with TDC below CMC, oxidative damages pointing to decreased mucosal detoxification potential with above CMC.     

2-Amino-5-chlorophenol toxicity in renal cortical slices from Fischer 344 rats: effect of antioxidants and sulfhydryl agents

A rapid inhibition of protein synthesis is observed when isolated rat hepatocytes are incubated in the presence of 0.25-0.5 mM of tert-butyl hydroperoxide (tBOOH). Such an inhibition occurs in the absence of a cytolytic effect by tBOOH. Iron chelators (o-phenanthroline and desferrioxiamine), protected against oxidative cell death, but they did not modify the inhibition of protein synthesis caused by tBOOH (0.5 mM), suggesting that free radicals are less implicated in such an impairment. Electron micrographs of hepatocytes under oxidative stress show disaggregation of polyribosomes but not oxidative alterations, such as blebs or mitochondrial swelling. Protein synthesis inhibition is accompanied by a decrease in reduced glutathione (GSH) and an increase in glutathione disulfide (GSSG) and the level of protein S-thiolation (protein mixed disulfides formation). Such an increase of GSSG appears as a critical event since diethylmaleate (DEM) at 0.2 mM reduced GSH content by more than 50% but did not affect either GSSG content or protein synthesis. The addition of exogenous GSH and N-acetylcysteine (NAC) to tBOOH-treated hepatocytes significantly reduced the formation of protein mixed disulfides and restored the depressed protein synthesis either completely or partially. We suggest that S-thiolation of some key proteins may be involved in protein synthesis inhibition by tBOOH.     

Differential Effects of Cisplatin on the Production of NADH-Dependent Superoxide and the Activity of Antioxidant Enzymes in Rat Renal Cortical Slices in vitro

Abstract: Cisplatin-induced cytotoxicity has been investigated with rat renal cortical slices in vitro. Incubation of renal slices with cisplatin caused an unexpected decrease in NADH-dependent, lucigenin-enhanced chemiluminescence in the homogenate of the slices in a time- and concentration-dependent fashion. Cisplatin caused a concentration-related (0.2–1 mM) decrease in the formation of NADH-dependent superoxide anion. Cisplatin (2 mM) significantly suppressed the chemiluminescence to 70% of control as early as 15 min. and to 15% after 90 min. of incubation. A decrease was also observed 90 min. after incubation of slices with 0.25 mM cisplatin. In addition, the activities of superoxide dismutase (SOD) and catalase in the slices were significantly increased after 60 min. of exposure to cisplatin (2 mM) while decreases in the activities of glutathione (GSH) peroxidase and GSSG reductase became significant at 90 and 120 min. of incubation respectively. SOD and catalase activities were increased by 1.6 and 1.5 fold respectively after 90 min. of incubation and the activities of GSH peroxidase and GSSG reductase were decreased to 82% and 72% of control, respectively at 120 min. Both dithiothreitol (2 mM), a sulphydryl agent and diphenylphenylenediamine (5 μM), an antioxidant, protected against cisplatin-induced leakage of lactate dehydrogenase, lipid peroxidation and decreases of GSH peroxidase and GSSG reductase but had no effect on the decrease of chemiluminescence caused by cisplatin. The results suggest that neither an increase in the production of NADH-dependent superoxide anion nor a decrease in activity of several antioxidant enzymes were directly responsible for cisplatin-induced lipid peroxidation.     

Occupational airborne contamination in South Brazil: 2. Oxidative stress detected in the blood of workers of incineration of hospital residues

One of the most useful methods for elimination of solid residues of health services (SRHS) is incineration. However, it also provokes the emission of several hazardous air pollutants such as heavy metals, furans and dioxins, which produce reactive oxygen species and oxidative stress. The present study, which is parallel to an accompanied paper (Ávila Jr. et al., this issue), investigated several enzymatic and non-enzymatic biomarkers of oxidative stress in the blood (contents of vitamin E, lipoperoxidation = TBARS, reduced glutathione = GSH, oxidized glutathione = GSSG, and activities of glutathione S-transferase = GST, glutathione reductase = GR, glutathione peroxidase = GPx, catalase = CAT and superoxide dismutase = SOD), in three different groups (n = 20 each) exposed to airborne contamination associated with incineration of SRHS: workers directly (ca. 100 m from the incinerator) and indirectly exposed (residents living ca. 5 km the incineration site), and controls (non-exposed subjects). TBARS and GSSG levels were increased whilst GSH, TG and α-tocopherol contents were decreased in workers and residents compared to controls. Increased GST and CAT activities and decreased GPx activities were detected in exposed subjects compared to controls, while GR did not show any difference among the groups. In conclusion, subjects directly or indirectly exposed to SRHS are facing an oxidative insult and health risk regarding fly ashes contamination from SRHS incineration.     

Inorganic mercury exposure: toxicological effects,oxidative stress biomarkers and bioaccumulation in the tropical freshwater fish matrinxã, <Emphasis Type="Italic">Brycon amazonicus</Emphasis> (Spix and Agassiz, 1829)

Alterations in the antioxidant cellular system have often been proposed as biomarkers of pollutant-mediated toxicity. This study evaluated the effects of mercury on oxidative stress biomarkers and bioaccumulation in the liver, gills, white muscle and heart of the freshwater fish matrinxã, Brycon amazonicus, exposed to a nominal and sub-lethal concentration (~20% of 96 h-LC₅₀) of 0.15 mg L^?1 of mercury chloride (HgCl₂) for 96 h in a static system. Increases in superoxide dismutase, catalase, glutathione peroxidase (GPx), glutathione S-transferase (GST) and glutathione reductase (GR) were observed in all tissues after HgCl₂ exposure, except for white muscle GR activity and hepatic GPx. In the liver and gills, the exposure to HgCl₂ also induced significant increases in reduced glutathione (GSH). Conversely, exposure to HgCl₂ caused a significant decrease in the GSH levels and an increase in the oxidized glutathione (GSSG) content in the white muscle, while both GSH and GSSG levels increased significantly in the heart muscle. Metallothionein concentrations were significantly high after HgCl₂ exposure in the liver, gills and heart, but remained at control values in the white muscle. HgCl₂ exposure induced oxidative damage, increasing the lipid peroxidation and protein carbonyl content in all tissues. Mercury accumulated significantly in all the fish tissue. The pattern of accumulation follows the order gills > liver ? heart > white muscle. In conclusion, these data suggest that oxidative stress in response to inorganic mercury exposure could be the main pathway of toxicity induced by this metal in fish.     

Time-dependent effect of p-aminophenol (PAP) toxicity in renal slices and development of oxidative stress

p-Aminophenol (PAP), a metabolite of acetaminophen, is nephrotoxic. This study investigated PAP-mediated changes as a function of time that occur prior to loss of membrane integrity. Experiments further evaluated the development of oxidative stress by PAP. Renal slices from male Fischer 344 (F344) rats (N = 4-6) were exposed to 0.1, 0.25, and 0.5 mM PAP for 15-120 min under oxygen and constant shaking at 37 degrees C. Pyruvate-stimulated gluconeogenesis, adenine nucleotide levels, and total glutathione (GSH) levels were diminished in a concentration- and time-dependent manner prior to detection of a rise in lactate dehydrogenase (LDH) leakage. Glutathione disulfide (GSSG) levels were increased by PAP suggesting the induction of oxidative stress. Western blot analysis confirmed a rise in 4-hydroxynonenal (4-HNE)-adducted proteins in tissues exposed to 0.1 and 0.25 mM PAP for 90 min. The appearance of 4-HNE-adducted proteins at the 0.1 mM concentration of PAP occurred prior to development of increased LDH leakage. Pretreatment with 1 mM glutathione (GSH) for 30 min only partially reduced PAP toxicity as LDH values were less severely depleted relative to tissues not pretreated with GSH. In contrast, pretreatment for 15 min with 2 mM ascorbic acid completely protected against PAP toxicity. Further studies showed that ascorbic acid pretreatment prevented PAP-mediated depletion of GSH. In summary, PAP rapidly depletes GSH and adenine nucleotides and inhibits gluconeogenesis prior to a rise in LDH leakage. PAP induces oxidative stress as indicated by an increase in GSSG and 4-HNE-adducted proteins. Ascorbic acid pretreatment prevents PAP toxicity by maintaining GSH status.     

Dietary intake of inorganic mercury: bioaccumulation and oxidative stress parameters in the neotropical fish Hoplias malabaricus

This study evaluated the effects of trophic and subchronic exposure to inorganic mercury (Hg) on the oxidative stress biomarkers and its bioaccumulation potential in the liver, gills, white muscle and heart of the freshwater top predator fish, Hoplias malabaricus, fed with contaminated live juveniles of matrinxã, Brycon amazonicus, as prey vehicle. Inorganic mercury increased superoxide dismutase, catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase, and glutathione reductase (GR) activities in the liver, white muscle and heart. Gills CAT activity remained unchanged while GPx and GR values showed a significant decrease. In the liver and gills, Hg induced significant increase in the reduced (GSH) and oxidized (GSSG) glutathione content, concomitantly with a significant decrease in [GSH]/[GSSG] ratio. Differently, in cardiac tissue, the Hg caused an increase in GSH level and increase in [GSH]/[GSSG] ratio. Lipid and protein oxidation and metallothionein levels were significantly higher after Hg trophic exposure in the liver, gills and heart, but remained at control values in the white muscle. Tissue-specific responses against oxidative stress were observed, and the liver and gills were the most sensitive organs, showing signs of redox homeostasis failure. At the end of the experiment, dietary inorganic mercury accumulated through food chain levels. In order, Hg bioaccumulation was: gills > liver ? white muscle = heart. These results pointed out the potential of inorganic Hg to bioaccumulate in aquatic systems. Taken together, our findings suggest that Hg, even in the inorganic form and sublethal amounts, is a risk factor for aquatic biota.     

Interactive effects of Salicylic acid and nitric oxide in alleviating zinc toxicity of Safflower (<Emphasis Type="Italic">Carthamus tinctorius</Emphasis> L.)

The purpose of this study was to assess the possible protective role of exogenous salicylic acid (SA), sodium nitroprusside (SNP), a donor of nitric oxide, and their combination on 21-day-old safflower (Carthamus tinctorius L.) seedlings grown under zinc (Zn) stress. The results revealed that exposure to 500?µM ZnSO₄.7H₂O for 10 days markedly reduced the root and shoot dry weights in Zn-treated plants, while the application of SA, SNP and specially SA?+?SNP significantly increased the root and shoot dry weights in seedlings subjected to Zn stress. Addition of SA, SNP and SA?+?SNP interestingly reduced root-to-shoot translocation of zinc and increased significantly the level of glutathione (GSH) and ascorbate (ASC) in leaves of Zn-stressed plants. The Zn-treated plants supplemented with SA and SNP revealed an improved activity of ascorbate-glutathione cycle enzymes and those enzymes which are involved in glyoxalase system as compared to the plants treated with Zn only. However, no significant relationship was found between SA or SNP supplementation and glutathione S-transferase activity in Zn-stressed plants. These findings demonstrate that exogenous application of SA or SNP could ameliorate the negative effects of Zn on safflower plants probably by stimulation of antioxidant defense and glyoxalase systems.     

Oxidative stress-related markers and langerhans cells in a hairless rat model exposed to UV radiation

Biomarkers related to the oxidative stress in blood and epidermis and the number of Langerhans cells were determined in hairless rats after acute irradiation with 1.54, 1.93, or 2.41 J/cm2 of ultraviolet (UV) light and chronic exposure to 13 suberythemal UV doses of 1.1 J/cm2 for 2 mo. After acute UV irradiation, in epidermis, the thiobarbituric acid-reactive substances (TBARS) content increased at the highest UV dose, whereas the activities of glutathione S-transferase and catalase rose and the oxidized glutathione (GSSG) content diminished at all UV doses. In erythrocytes, glutathione S-transferase activity increased at the two lowest UV doses, glutathione peroxidase activity rose at all UV doses, and catalase activity increased after the highest UV dose. In plasma, the TBARS content and the reduced glutathione (GSH)/GSSG ratio increased at the highest UV dose; the number of Langerhans cells decreased at all UV doses. Linear Pearson correlation analysis revealed many relationships between different biomarkers, and multiple linear regression analysis indicated that the number of Langerhans cells was predicted by epidermal GSSG and catalase (R2 = .64) and by erythrocytic glutathione peroxidase and GSSG (R2 = .72). After suberythemal UV radiation, in epidermis, the GST activity and the content of GSH and GSSG increased; in erythrocytes, the GST activity decreased and the GSH/GSSG ratio increased. Thus, the hairless rat appears to be a useful model for studying the oxidative stress-related mechanisms after UV radiation, which are involved in the loss of the immune capacity mediated by Langerhans cells, even at suberythemal doses.     

葛根素对青光眼视网膜氧化应激损伤的保护作用

【摘要】目的研究葛根素对青光眼视网膜氧化应激损伤的保护作用。方法24只健康雄性成年家兔随机分为对照组、模型组及治疗组,每组8只。将2.5%羟丙基甲基纤维素溶液0.2mL注入家兔前房内,制作模型组及治疗组兔青光眼模型,治疗组家兔腹腔注射葛根素10mL/kg,每日1次,共21d。模型建立后第4周,测定视网膜抗氧化酶超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）和过氧化氢酶（CAT）活性,检测抗氧化物质谷胱甘肽含量及氧化还原状态和丙二醛（MDA）、一氧化氮（NO）含量。结果模型组视网膜抗氧化酶SOD、GSH-Px和CAT活性、还原型谷胱甘肽（GSH）和GSH/氧化型谷胱甘肽（GSSG）均低于对照组,GSSG、MDA和NO含量高于对照组;治疗组视网膜上述抗氧化酶活性、GSH含量、GSH/GSSG均高于模型组,但低于对照组;GSSG、MDA和NO含量均低于模型组,GSSG含量与对照组差异无统计学意义。结论葛根素可以在一定程度上增强抗氧化酶活性及维持谷胱甘肽氧化还原状态,从而减轻青光眼视网膜的氧化应激损伤。      

Effect of selenium on glutathione metabolism. Induction of gamma-glutamylcysteine synthetase and glutathione reductase in the rat liver

The effect of sodium selenite (Na₂SeO₃, Se) on cellular glutathione metabolism was examined, particularly with respect to its ability to alter the activities of γ-glutamylcysteine synthetase and glutathione disulfide (GSSG) reductase. The treatment of rats with Se (5, 10 and 20 μmoles/kg) caused time- and dose-dependent increases in the activities of the synthetase and the reductase in the liver. The activity of γ-glutamylcysteine synthetase, the rate-limiting enzyme of the glutathione (GSH) biosynthesis, was particularly susceptible to Se treatment. The Se-mediated increases in the activities of the above enzymes were inhibited by puromycin and the increases could not be elicited in vitro. Selenium treatment caused time-dependent perturbations in the levels and ratio of GSSG and GSH in the liver. When compared to the control animals, rats treated for 3 hr with 10 and 20 μmoles Se/kg showed increased cellular levels of GSSG; in contrast, 24 hr after Se treatment the concentration of GSH was increased significantly. The activity of γ-glutamyl transpeptidase, which catalyzes the initial reaction in GSH breakdown, was unaltered by Se treatment. Repeated administration of low doses of Se (7.0 μmoles/kg, three times) also increased the activities of the reductase and the synthetase as well as the cellular levels of hepatic GSH and GSSG. It is suggested that the Se-mediated increases in the activities of γ-glutamylcysteine synthetase and GSSG-reductase represent cellular responses to Se-mediated perturbations in the levels and ratio of GSH and GSSG.     

Effects of glycerol-induced acute renal failure on tissue glutathione and glutathione-dependent enzymes in the rat

The activities of tissue glutathione (reduced and oxidized) and glutathione-dependent enzymes such as glutathione S-transferase (GSH S-transferase), glutathione reductase (GSSG reductase) and glutathione peroxidase (GSH-Px) were determined for control and uremic rats. Acute renal failure (ARF) was produced by glycerol-water injection. Cytosolic and microsomal GSH S-transferase activity in the kidney was decreased by 38% and 15%, respectively. Hepatic microsomal GSH S-transferase was also decreased by 40% in uremic rats. GSH-Px activity was decreased by 51% in the cytosolic fraction and 33% in the microsomal fraction in the kidney, but was not affected in the liver and whole blood. GSSG reductase activity was also decreased by 48% in the cytosolic fraction in the kidney of uremic rats. In whole blood, however, GSSG reductase activity was increased by 12-fold (0.66 +/- 0.12 mumol NADPH oxidized/min/ml blood in the control; 8.03 +/- 3.29 mumol NADPH oxidized/min/ml blood in uremia). Although the total glutathione concentrations were not significantly affected, the GSSG/GSH ratio, which is an indication of oxidative stress, was significantly increased in the liver and whole blood of uremic rats. In addition to the decreases in hepatic and renal GSH S-transferase activities, which is important in drug disposition, ARF caused decreases in GSSG reductase and GSH-Px activity, which are essential for the protection against lipid peroxidation.     

Glutathione redox status of control and cadmium oxide-exposed rat lungs during oxidant stress

Activities of enzymes responsible for the maintenance of reduced glutathione (GSH) levels have been shown in a previous study to be increased in rat lungs following a 3-h exposure to cadmium oxide aerosols at 5.0 mg/m3. In this study, the ability of the lung to maintain levels of GSH during challenge with tert-butyl hydroperoxide (tBuOOH) was evaluated in isolated perfused lungs from control and cadmium oxide-exposed rats. Changes in glutathione redox status were indicated by measurements of nonprotein sulfhydryls (NPSH), total glutathione (1/2 GSH + GSSG), and glutathione disulfide (GSSG) in liquid nitrogen freeze-clamped lungs after 3-min infusions with 0-0.6 mM tBuOOH. In control and cadmium oxide-exposed lungs, levels of 1/2 GSH + GSSG remained constant over the range of 0-0.6 mM tBuOOH, indicating that no loss of glutathione from the system had occurred. In experiments with control lungs, levels of NPSH fell from 8.04 +/- 0.22 to 3.09 +/- 0.40 mumol/g dry weight when tBuOOH concentrations were increased from 0 to 0.6 mM (n = 20-23). In cadmium oxide-exposed lungs, NPSH levels also decreased proportionally to increases in GSSG. However, at concentrations of 0.075 and 0.15 mM tBuOOH, significantly smaller decreases in NPSH levels were observed in cadmium oxide-exposed lungs compared with controls. This protection against the GSH-depleting effects of tBuOOH might be explained by increased tissue levels of GSH-related enzymes.     

Pyruvate reduces 4-aminophenol in vitro toxicity

Pyruvate has been observed to reduce the nephrotoxicity of some agents by maintaining glutathione status and preventing lipid peroxidation. This study examined the mechanism for pyruvate protection of p-aminophenol (PAP) nephrotoxicity. Renal cortical slices from male Fischer 344 rats were incubated for 30-120 min with 0, 0.1, 0.25 or 0.5 mM PAP in oxygenated Krebs buffer containing 0 or 10 mM pyruvate or glucose (1.28 or 5.5 mM). LDH leakage was increased above control by 0.25 and 0.5 mM PAP beginning at 60 min and by 0.1 mM PAP at 120 min. Pyruvate prevented an increase in LDH leakage at 60- and 120-min exposure to 0.1 and 0.25 mM PAP. Pyruvate also prevented a decline in ATP levels. Glucose (1.28 and 5.5 mM) provided less protection than pyruvate from PAP toxicity. Total glutathione levels were diminished by 0.1 and 0.25 mM PAP within 60 and 30 min, respectively. Pyruvate prevented the decline in glutathione by 0.1 mM PAP at both time periods and at 30 min for 0.25 mM PAP. Pyruvate reduced the magnitude of glutathione depletion by 0.25 mM PAP following a 60-min incubation. Glutathione disulfide (GSSG) levels in renal slices were increased at 60 min by exposure to 0.25 mM PAP, while pyruvate prevented increased GSSG levels by PAP. Pyruvate also reduced the extent of 4-hydroxynonenal (4-HNE)-adducted proteins present after a 90-min incubation with PAP. These results indicate that pyruvate provided protection for PAP toxicity by providing an energy substrate and reducing oxidative stress.     

The toxicity of N-methyl-alpha-methyldopamine to freshly isolated rat hepatocytes is prevented by ascorbic acid and N-acetylcysteine

In the past decade, clinical evidence has increasingly shown that the liver is a target organ for 3,4-methylenedioxymethamphetamine (MDMA, "ecstasy") toxicity. The aims of the present in vitro study were: (1) to evaluate and compare the hepatotoxic effects of MDMA and one of its main metabolites, N-methyl-alpha-methyldopamine (N-Me-alpha-MeDA) and (2) to investigate the ability of antioxidants, namely ascorbic acid and N-acetyl-L-cysteine (NAC), to prevent N-Me-alpha-MeDA-induced toxic injury, using freshly isolated rat hepatocytes. Cell suspensions were incubated with MDMA or N-Me-alpha-MeDA in the final concentrations of 0.1, 0.2, 0.4, 0.8, and 1.6 mM for 3 h. To evaluate the potential protective effects of antioxidants, cells were preincubated with ascorbic acid in the final concentrations of 0.1 and 0.5 mM, or NAC in the final concentrations of 0.1 and 1 mM for 15 min before treatment with 1.6 mM N-Me-alpha-MeDA for 3 h (throughout this incubation period the cells were exposed to both compounds). The toxic effects were evaluated by measuring the cell viability, glutathione (GSH) and glutathione disulfide (GSSG), ATP, and the cellular activities of GSH peroxidase (GPX), GSSG reductase (GR), and GSH S-transferase (GST). MDMA induced a concentration- and time-dependent GSH depletion, but had a negligible effect on cell viability, ATP levels, or on the activities of GR, GPX, and GST. In contrast, N-Me-alpha-MeDA was shown to induce not only a concentration- and time-dependent depletion of GSH, but also a depletion of ATP levels accompanied by a loss in cell viability, and decreases in the antioxidant enzyme activities. For both compounds, GSH depletion was not accompanied by increases in GSSG levels, which seems to indicate GSH depletion by adduct formation. Importantly, the presence of ascorbic acid (0.5 mM) or NAC (1 mM) prevented cell death and GSH depletion induced by N-Me-alpha-MeDA. The results provide evidence that MDMA and its metabolite N-Me-alpha-MeDA induce toxicity to freshly isolated rat hepatocytes. Oxidative stress may play a major role in N-Me-alpha-MeDA-induced hepatic toxicity since antioxidant defense systems are impaired and administration of antioxidants prevented N-Me-alpha-MeDA toxicity.     

Prevention of cadmium-induced effects on regional glutathione status of rat brain by vitamin E

The effect of vitamin E on the cadmium-induced changes of glutathione metabolism was investigated in different brain regions. Daily intraperitoneal injection of cadmium (0.4 mg/kg) for 30 days significantly decreased the concentration of reduced glutathione (GSH), and the activities of glutathione reductase (GR) and glucose-6-phosphate dehydrogenase (GPDH) in the cerebellum, cerebral hemispheres and brain stem of rats. Cadmium elevated the levels of oxidized glutathione (GSSG) in cerebellum and cerebral hemisphere regions only, while the GSH/GSSG ratio decreased in all three brain regions. The only effect of intramuscular injections of vitamin E (5 mg/kg) given on alternate days for 30 days was a slight increase in GSH and GR in the cerebral hemispheres. The simultaneous administration of vitamin E and cadmium prevented cadmium-induced changes in GSH and GSSG levels and in the GSH/GSSG ratio, but the cerebellar GSH remained lowered. Furthermore, vitamin E, with the exception of GR in the cerebral hemispheres, did not prevent cadmium-induced changes in enzyme activities. As the simultaneous injections of vitamin E reduced cadmium-induced alterations in glutathione concentration without having any appreciable effect on the activity of related enzymes, it is suggested that the preventive effect of vitamin E is mediated through its antioxidative effect, saving GSH from oxidative destruction in the brain of cadmium-exposed rats.