GSH-related Enzymes Research Articles

Vascular endothelial growth factor is associated with hepatocellular carcinoma recurrence, independent of folate and glutathione-related antioxidant enzymes: A follow-up study

The associations of tumor angiogenesis with folate and antioxidant capacities in patients with hepatocellular carcinoma (HCC) and their effects on HCC recurrence have not yet been investigated. We investigated the changes and relationships of VEGF, folate, GSH, and GSH-related antioxidant enzymes in patients with HCC before tumor resection, as well as 1 month, 1 year, and 3 years after tumor resection, and their effects on HCC recurrence. 95 HCC patients who underwent tumor resection were recruited. Patients were followed up before tumor resection (pre-resection), 1 month after tumor resection (post-resection), 1 year, and 3 years of follow-up. The recurrence and survival status of patients were evaluated. Plasma VEGF concentrations decreased slightly during follow-up. Serum folate and GSH concentrations and plasma GPx and GR activities increased significantly from pre-resection to post-resection and remained stable at follow-up. Pre-resection plasma VEGF was positively correlated with GSH, GPx, and GR, but negatively correlated with folate and GST. The high pre-resection plasma VEGF was a significant predictor of a high HCC rate (hazard ratio = 1.05, p = 0.035), remaining significant after adjustments for folate, GSH, GPx, GR, and GST to diminish their interference with VEGF. Pre-tumor-resection plasma VEGF constitutes a potential independent marker for predicting HCC recurrence. However, the associations of plasma VEGF with folate and GSH-related antioxidant capacities in HCC patients cannot be ignored.

GSH-related enzyme activity and tumor relation: glutathione peroxidase and glutathione reductase status under hypoxia in HepG2 cells

Abstract Objectives In hepatocellular carcinoma (HCC), tumorigenesis, hypoxia and reactive oxygen species (ROS) play a crucial role in altering the tumor microenvironment (TME). Until now, the time-dependent alteration of hypoxia-inducible factor-1α (HIF-1α), the antioxidant enzymes glutathione peroxidase (GPx) and glutathione reductase (GR) under hypoxic conditions in HCC were not clear. Consequently, our main target was to investigate the role of GPx and GR status in HCC cell line (HepG2) under hypoxic conditions. Methods HIF-1α protein levels in cell lysates were determined by ELISA assay and protein expressions were identified using western blot. GPx and GR activity levels of the cell lysates were measured spectrophotometrically. Results HIF-1α protein levels were determined under normoxic and hypoxic conditions (p<0.001). Also, HIF-1α protein levels and expressions were observed under time-dependent hypoxic conditions, the HIF-1α protein level is found to be reached its peak point at 4 h in the HepG2 cell line. We also have detected decreased activity levels of GPx and increased GR activity levels under hypoxia for 4 h (p<0.001). Conclusions More than 4 h of exposure to hypoxic environment reducted the HIF-1α levels in HCC cells. According to the results, we suggest the ideal exposure time to hypoxic conditions as 4 h for the HepG2 cell line. In addition, hypoxia also stimulated the activity levels of GPx and GR. Our results suggest that the activity levels of GPx and/or GR enzymes may be therapeutic targets in the hypoxia-dependent HCC tumorigenesis process.

Time of day differences in the regulation of glutathione levels in the rat lens.

Evidence in non-ocular tissues indicate that the antioxidant glutathione (GSH) may be regulated in a circadian manner leading to the idea that GSH levels in the lens may also be controlled in a circadian manner to anticipate periods of oxidative stress. Male rat Wistar lenses (6 weeks) were collected every 4 hours over a 24-hour period at 6am, 10am, 2pm, 6pm, 10pm and 2am and quantitative-PCR, western blotting and immunohistochemistry performed to examine the expression of core clock genes and proteins (BMAL1, CLOCK, CRY1-2, PER 1-3) and their subcellular localisation over a 24-hour period. Western blotting of lenses was also performed to examine the expression of NRF2, a transcription factor involved in regulating genes involved in GSH homeostasis and GSH related enzymes (GCLC, GS and GR) over the 24-hour period. Finally, HLPC was used to measure GSH levels in the aqueous humour and lenses every 4 hours over a 24-hour period. The rat lens contains the core molecular components of a circadian clock with the expression of core clock proteins, NRF2 and GSH related enzymes fluctuating over a 24-hour period. BMAL1 expression was highest during the day, with BMAL1 localised to the nuclei at 10am. NRF2 expression remained constant over the 24-hour period, although appeared to move in and out of the nuclei every 4 hours. GSH related enzyme expression tended to peak at the start of night which correlated with high levels of GSH in the lens and lower levels of GSH in the aqueous humour. The lens contains the key components of a circadian clock, and time-of-day differences exist in the expression of GSH and GSH related enzymes involved in maintaining GSH homeostasis. GSH levels in the rat lens were highest at the start of night which represents the active phase of the rat when high GSH levels may be required to counteract oxidative stress induced by cellular metabolism. Future work to directly link the clock to regulation of GSH levels in the lens will be important in determining whether the clock can be used to help restore GSH levels in the lens.

Mitochondrial remodeling and energy metabolism adaptations in colonic crypts during spontaneous epithelial repair after colitis induction in mice

Mucosal healing has emerged as a therapeutic goal to achieve lasting clinical remission in ulcerative colitis. Intestinal repair in response to inflammation presumably requires higher energy supplies for the restoration of intestinal barrier and physiological functions. However, epithelial energy metabolism during intestinal mucosal healing has been little studied, whereas inflammation-induced alterations have been reported in the main energy production site, the mitochondria. The aim of the present work was to assess the involvement of mitochondrial activity and the events influencing their function during spontaneous epithelial repair after colitis induction in mouse colonic crypts. The results obtained show adaptations of colonocyte metabolism during colitis to ensure maximal ATP production for supporting energetic demand by both oxidative phosphorylation and glycolysis in a context of decreased mitochondrial biogenesis and through mitochondrial function restoration during colon epithelial repair. In parallel, colitis-induced mitochondrial ROS production in colonic epithelial cells was rapidly associated with transient expression of GSH-related enzymes. Mitochondrial respiration in colonic crypts was markedly increased during both inflammatory and recovery phases despite decreased expression of several mitochondrial respiratory chain complex subunits after colitis induction. Rapid induction of mitochondrial fusion was associated with mitochondrial function restoration. Finally, in contrast with the kinetics expression of genes involved in mitochondrial oxidative metabolism and in glycolysis, the expression of glutaminase was markedly reduced in the colonic crypts both during colitis and repair phases. Overall, our data suggest that the epithelial repair after colitis induction is characterized by a rapid and transient increased capacity for mitochondrial ATP production in a context of apparent restoration of mitochondrial biogenesis and metabolic reorientation of energy production. The potential implication of energy production adaptations within colonic crypts to sustain mucosal healing in a context of altered fuel supply is discussed.

Cysteine Regulates Oxidative Stress and Glutathione-Related Antioxidative Capacity before and after Colorectal Tumor Resection

Cysteine might scavenge free radicals and is a limiting substrate for the cellular synthesis of glutathione (GSH). We investigated the association of cysteine with oxidative stress and GSH-related antioxidant capacity in colorectal cancer (CRC) patients. Plasma samples were drawn from 66 patients 1 day before (pre-resection) and 4 weeks after resection (post-resection). Tumor and adjacent normal tissues were collected. We measured levels of plasma and tissue cysteine, homocysteine, oxidative stress indicators (malondialdehyde, MDA; advanced oxidation protein products, AOPP), GSH, and antioxidant enzyme activities. After tumor resection, patients had significantly higher levels of plasma cysteine, homocysteine, MDA, AOPP, and GSH-related antioxidant enzyme activities when compared with pre-resection. Levels of cysteine, homocysteine, AOPP and all antioxidant capacity indicators in tumor tissue were significantly higher than those levels in the adjacent normal tissue. Plasma cysteine levels measured at pre-resection were positively associated with MDA levels in the tumor and in the adjacent normal tissues. Cysteine levels in tumor and adjacent normal tissues were significantly associated with tissue levels of homocysteine, almost as indicators of oxidative stress and antioxidant capacities. Cysteine in the circulation was likely utilized to mediate GSH-related antioxidant capacity and further cope with increased oxidative stress in tumor and adjacent normal tissues.

Differential Activity of Antioxidants in Testicular Tissues Following Administration of Chlorophytum borivilianum in Gamma-Irradiated Swiss Albino Mice.

Background: Oxidative stress induced by radiation causes variable expression of antioxidant enzymes in a tissue-specific manner. Testicular tissues carry out the complex process of spermatogenesis, and studies indicate that testicular damages due to irradiation require long-term recovery before complete resumption. Ionizing radiation also causes oxidative stress in tissues, leading to testicular damage. Aims and Objectives: This study measured differential expression of antioxidant enzymes following administration of C. borivilianum root extract (CRB) in response to irradiation-induced oxidative stress. The activity of various important endogenous enzymatic defense systems was evaluated and correlated for strength of association. Materials and method: Two forms of C. borivilianum (CB) extracts [CB alone and CB-silver nanoparticles (AgNPs)] were administered at a dose of 50 mg/kg body weight to Swiss albino male mice for 7 consecutive days. After that, they were irradiated with 6 Gy irradiation and further used to study various parameters of antioxidant enzymes. Results: Results indicate a significant increase in the level of glutathione (GSH) and the activity of GSH-related antioxidant enzymes in irradiated mice treated with CRE and CRE-AgNPs (silver nanoparticles biosynthesized using C. borivilianum root extract) in comparison to non-pretreated ones (groups I and II). Reciprocal elevation was observed in related enzymes, that is, glutathione S-transferase activity (GST), glutathione reductase (GR), and glutathione peroxidase activity (GPx). Elevation in the activity of catalase (CAT) and superoxide dismutase (SOD) was also evident in both the irradiated groups pretreated with CRE-AgNPs. However, expression of CAT in the CRE-treated irradiated group was similar to that of the non-treated irradiated group. Higher association among CAT-SOD, CAT-GPx, and GR-GST was observed. Conclusion: Overall, it was observed that testicular cells post-irradiation in all groups go through intense oxidative stress; however, groups pretreated with CRE or CRE-AgNPs indicated better toleration and resumption of antioxidant capacity. CRE or CRE-AgNPs pretreated non-irradiated groups mostly remained within the control range indicating stimulated expression of antioxidants.

Molecular characterization of superoxide dismutase and catalase genes, and the induction of antioxidant genes under the zinc oxide nanoparticle-induced oxidative stress in air-breathing magur catfish (Clarias magur).

The deduced amino acid sequences from the complete cDNA coding sequences of three antioxidant enzyme genes (sod1, sod2, and cat) demonstrated that phylogenetically the magur catfish (Clarias magur) is very much close to other bony fishes with complete conservation of active site residues among piscine, amphibian, and mammalian species. The three-dimensional structures of three antioxidant enzyme proteins are very much similar to mammalian counterparts, thereby suggesting the functional similarities of these enzymes. Exposure to ZnO NPs resulted in an oxidative stress as evidenced by an initial sharp rise of intracellular concentrations of hydrogen peroxide (H2O2) and malondialdehyde (MDA) but decreased gradually at later stages. The level of glutathione (GSH) also increased gradually in all the tissues examined after an initial decrease. Biochemical and gene expression analyses indicated that the magur catfish has the ability to defend the ZnO NP-induced oxidative stress by inducing the SOD/CAT enzyme system and also the GSH-related enzymes that are mediated through the activation of various antioxidant-related genes both at the transcriptional and translational levels in various tissues. Furthermore, it appeared that the stimulation of NO, as a consequence of induction nos2 gene, under NP-induced oxidative stress serves as a modulator to induce the SOD/CAT system in various tissues of magur catfish as an antioxidant strategy. Thus, it can be contemplated that the magur catfish possesses a very efficient antioxidant defensive mechanisms to defend against the oxidative stress and also from related cellular damages during exposure to ZnO NPs into their natural environment.

Oxidative stress activates the Nrf2-mediated antioxidant response and P38 MAPK pathway: A possible apoptotic mechanism induced by BDE-47 in rainbow trout (Oncorhynchus mykiss) gonadal RTG-2 cells

Our previous study showed that 2,2′,4,4′-tetrabromodiphenyl ether (BDE-47), the most biotoxic polybrominated diphenyl ether (PBDE) in the marine environment, induced apoptosis in rainbow trout gonadal RTG-2 cells. This effect occurred via ROS- and Ca2+-mediated apoptotic pathways, but the exact mechanism remains unknown. Therefore, in the present study, the possible mechanism was examined from the perspective of ROS-induced oxidative stress. The results showed that BDE-47 exposure significantly elevated the malondialdehyde (MDA) contents and the intracellular GSH/GSSG ratio, and the GSH-related enzymes were greatly altered, indicating alteration of the redox status and occurrence of oxidative stress. The mRNA levels of nuclear factor E2-related factor 2 (Nrf2) and its downstream genes were simultaneously greatly elevated. The p38 mitogen-activated protein kinase (MAPK) signaling pathway was also found to be induced by BDE-47 exposure. The addition of SB203580, a p38 MAPK inhibitor resulted in decreased apoptosis. In addition, supplementation with Ca2+ inhibitors BAPTA-AM positively affected p38 MAPK activation. Taken together, BDE-47 exposure resulted in the occurrence of oxidative stress and initiated the Nrf2-mediated antioxidant response. Subsequently, the altered redox status induced p38 MAPK activation, which played a pivotal role in the cellular apoptosis of RTG-2 cells.

Effect of phloretin on growth performance, serum biochemical parameters and antioxidant profile in heat-stressed broilers

The objective of this work was to evaluate the effect of phloretin on growth performance, serum biochemical parameters, antioxidant profile, glutathione (GSH)-related enzymes, nuclear factor erythroid 2–related 2 (Nrf2) and heat shock protein 70 (HSP70) in heat-stressed broilers. A total of 240, 22-day-old Arbor Acres broilers were divided into 4 groups. The control group was housed at 23.0 ± 0.61°C and fed with basal diet, while the 3 heat-stressed groups (A, B, and C groups) were housed at 30.5 ± 0.69°C and fed with basal diet containing 0, 100, and 200 mg/kg phloretin, respectively. Serum was taken form 42-day-old broilers. Results showed that heat stress decreased (P < 0.05) the final body weight (FBW), body weight gain (BWG), feed intake (FI), serum total protein (TP), triglyceride (TG), triiodothyronine (T3), thyroxine (T4), GSH, catalase (CAT), and total antioxidant capacity (T-AOC) levels, but increased (P < 0.05) the feed-to-gain ratio (FGR) and serum malondialdehyde (MDA) levels in broilers compared with that in the control group. Among the heat-stressed groups, supplementary 200 mg/kg phloretin increased (P < 0.05) the FBW, BWG, FI, serum TP, TG, T4, GSH, CAT, and T-AOC levels, and decreased (P < 0.05) the FGR and serum MDA in broilers. There were significant decreases (P < 0.05) in the glutathione peroxidase (GSH-Px), γ-glutamylcysteine synthetase (γ-GCS), and Nrf2, but significant increases (P < 0.05) in the HSP70 of the broiler serum after heat stress treatment. Among the heat-stressed groups, supplementary 200 mg/kg phloretin increased (P < 0.05) the GSH-Px, γ-GCS, and Nrf2 levels, but decreased (P < 0.05) the serum HSP70 level in the heat-stressed broilers. Under high temperature condition, FBW, BWG, FI, FGR, serum TP, TG, T4, MDA, GSH, CAT, T-AOC, GSH-Px, γ-GCS, Nrf2 and HSP70 were linearly affected by inclusion of phloretin. These results indicated that phloretin may improve growth performance, serum parameters, and antioxidant profiles through regulated GSH-related enzymes, Nrf2 and HSP70 in heat-stressed broilers.

Evidence for oxidative and not reductive stress in the aged rabbit heart.

Reductive stress, which is due to a paradoxical excess of antioxidants such as reduced glutathione (GSH) and GSH-related enzymes associated with decreased oxidant levels, has emerged as a pathogenetic mechanism of myocardial damage in pathological conditions such as protein aggregation cardiomyopathy. Notably, in the aged heart a cardiomyopathy-like pathology occurs leading to myocardial dysfunction. Whether reductive stress, or instead its counterpart oxidative stress, is operative in the aged mammalian heart needs to be elucidated also for the different therapeutic implications of such redox stress conditions. In the present investigation, we assessed GSH and the specific enzymatic activities of γ-glutamylcysteine synthetase (γ-GCS), glutathione reductase (GSSG-Red) and selenium-dependent glutathione peroxidase (GSH-Px) as endogenous antioxidants, together with oxidized glutathione (GSSG) and the glutathione redox ratio (GSH/GSSG), in the aerobically perfused hearts of aged rabbits (about 4.5years old) and young adult control rabbits (3-4months old). We also assessed in the aged and control hearts H2O2 and catalytically active low molecular weight iron (LMWI) as oxidant forces, as well as fluorescent damage products of lipid peroxidation (FDPL) and protein carbonyls (PC) as biomarkers of lipid and protein oxidation. Moreover, the effects of 4.5mM N-acetylcysteine (NAC) as reducing thiol antioxidant were studied on hemodynamic parameters and lipid peroxidation in the perfused hearts of the aged and control rabbits. The levels of GSH and of the GSH/GSSG ratio were lower, and those of GSSG higher, in the aged than in the control hearts. The aged hearts were also characterized by decreased activities of the antioxidant enzymes γ-GCS, GSSG-Red and GSH-Px, as well as by heightened levels of H2O2, LMWI, FDPL and PC, highlighting the occurrence of aging-dependent oxidative stress. Associated with such biochemical alterations, hemodynamic dysfunction occurred in the aged rabbit hearts, as evidenced by lowered developed pressure (DP) and enhanced end-diastolic pressure (EDP) with decreased coronary flow (CF). Remarkably, NAC administration significantly improved DP and EDP, and lowered lipid peroxidation, electively in the aged hearts. In conclusion, oxidative and not reductive stress is operative in the aged rabbit heart, whose hemodynamic dysfunction is improved by NAC together with reduction in myocardial lipid peroxidation.

Evidence of hippocampal astrogliosis and antioxidant imbalance after L-tyrosine chronic administration in rats.

Tyrosinemia type II is a genetic disorder characterized by elevated blood levels of the amino acid tyrosine caused by the deficiency of tyrosine aminotransferase enzyme, resulting in neurologic and developmental difficulties in the patients. Although neurological sequelae are common in Tyrosinemia type II patients, the mechanisms involved are still poorly understood. The oxidative stress appears to be, at least in part, responsible for neurological complication in this inborn error metabolism. We observed that an acute injection of tyrosine in rats caused a massive oxidative stress in different brain structures. The glutathione system and superoxide dismutase enzyme are relevant antioxidant strategies of the cells and tissues, including in the brain. Other important point is the strong relation between oxidative damage and inflammatory events. Herein, we investigated the effects of chronic administration of tyrosine in the hippocampus of young rats, with emphasis in the activity of GSH related enzymes and superoxide dismutase enzyme, and the astrocytosis. We observed that rats exposed to high levels of tyrosine presented an increased content of tyrosine, which was associated with an increment in the activity of glutathione peroxidase and glutathione reductase as well as with a diminished activity of superoxide dismutase. This antioxidant imbalance was accompanied by enhanced glial fibrillary acidic protein immunoreactivity, a marker of astrocytes, in the brain area studied. In conclusion, hippocampus astrogliosis is also a characteristic of brain alteration in Tyrosinemia. In addition, the chronic exposition to high levels of tyrosine is associated with an alteration in the activity of fundamental antioxidant enzymes.

Activity of Antioxidant Enzymes in the Tumor and Adjacent Noncancerous Tissues of Non-Small-Cell Lung Cancer.

Lung tissue is directly exposed to high oxygen pressure, as well as increased endogenous and exogenous oxidative stress. Reactive oxygen species (ROS) generated in these conditions play an important role in the initiation and promotion of neoplastic growth. In response to oxidative stress, the antioxidant activity increases and minimizes ROS-induced injury in experimental systems. The aim of the present study was to evaluate the activity of antioxidant enzymes, such as superoxide dismutase (SOD; isoforms: Cu/ZnSOD and MnSOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione S-transferase (GST), along with the concentration of malondialdehyde (MDA) in tumor and adjacent noncancerous tissues of two histological types of NSCLC, i.e., adenocarcinoma and squamous cell carcinoma, collected from 53 individuals with surgically resectable NSCLC. MDA concentration was similar in tumors compared with adjacent noncancerous tissues. Tumor cells had low MnSOD activity, usually low Cu/ZnSOD activity, and almost always low catalase activity compared with those of the corresponding tumor-free lung tissues. Activities of GSH-related enzymes were significantly higher in tumor tissues, irrespective of the histological type of cancer. This pattern of antioxidant enzymes activity could possibly be the way by which tumor cells protect themselves against increased oxidative stress.

Protective role of l-glutamic acid and l-cysteine in mitigation the chlorpyrifos-induced oxidative stress in rats

Exposure to the organophosphorus insecticide chlorpyrifos (CPF) can lead to oxidative stress. The aim of this work was to investigate and compare the protective effects of amino acids (l-glutamic acid (l-Glu) and l-cysteine (L-Cys) alone or in combination) for the purpose of suppression and mitigation of CPF-induced oxidative stress in rats. Rats were divided into five groups: CPF, CPF/L-Glu, CPF/L-Glu and l-Cys, CPF/L-Cys, control. The level of GSH and the activities of glutathione-related enzymes were determined. The content of lipid peroxidation products was also monitored. The obtained results suggest that level of GSH and activity of GSH-related enzymes was significantly inhibited by CPF. l-Glu and l-Cys were able to prevent CPF-induced oxidative stress. In rats treated with amino acids, we observed less significant or no changes in studied parameters. It was established that the above-mentioned amino acids, administered alone and in their combination, can mitigate and suppress CPF-induced oxidative stress. The most significant mitigation effect was found in rats treated with l-Glu only.

Protective effects of L-theanine against doxorubicin-induced nephrotoxicity in rats

L-theanine is the unique amino acid found in tea plants, has antioxidant and antiinflammatory activities, and functions in mental concentration and sleep quality. In this study, it is aimed to investigate the effects of L-theanine on doxorubicin (DOX, a chemotherapeutic agent) induced nephrotoxicity in rats, especially via GSH related enzymes. 32 male Sprague Dawley rats weighing 300-400 g were randomly assigned into 4 groups (n = 8) and the substances were given intraperitoneally to them: Control group (saline for 5 days); Theanine group (200 mg/kg/day theanine for 5 days); DOX group (single dose of 20 mg/kg DOX); DOX + Theanine group (20 mg/kg DOX at first day and 200 mg/kg/day theanine for 5 days). Kidney tissues were evaluated by histopathological analysis. Serum levels of blood urea nitrogen (BUN) and creatinine by spectrophotometrically; percentage of apoptosis indexes (AI%) in the tissues by TUNEL method; caspase-3 levels, reduced and oxidized glutathione (GSH and GSSG), gamma-glutamyltransferase 1 (GGT1), glutathione reductase (GR), glutathione peroxidase (GPx), and glutathione S-transferase (GST), nuclear factor kappa B p65 (NF-kB p65) by commercial kits; malondialdehyde (MDA) by spectrophotometrically were determined in plasma and kidney tissues. According to DOX group, the DOX + Theanine group has much lower tissue and plasma GSSG, GGT1, NF-κB p65 levels and tissue AI%, whereas significantly higher GSH levels and GPx, GR, GST activities (P < 0.05). It is suggested that L-theanine may have protective effects by enhancing effects on the antioxidant system of GSH and GSH-related enzymes against DOX-induced nephrotoxicity in rats. But this finding needs to be supported with further studies.

Curcumin attenuates heat-stress-induced oxidant damage by simultaneous activation of GSH-related antioxidant enzymes and Nrf2-mediated phase II detoxifying enzyme systems in broiler chickens

The object of this study was to investigate the effect of curcumin on modulating the glutathione (GSH)-related antioxidant enzymes and antioxidant responses via NF-E2-related factor 2 (Nrf2) signaling pathway in heat-stressed broiler chickens. A total of 400 one-day-old male Arbor Acres broiler chicks was reared in an environmentally controlled room. At 21 d, broiler chicks were divided into 5 treatment groups and were fed one of 4 diets under 2 temperature conditions: 22°C + a basal diet (CON treatment); 34°C for 8h (0900-1700) + a basal diet supplemented with 0, 50, 100, or 200mg/kg curcumin (HS, CMN1, CMN2, and CMN3 treatments, respectively). The heat treatment lasted for 20 consecutive days. The results showed that heat stress significantly increased (P <0.05) the weekly rectal temperature and average head and feet temperature. Compared to the HS treatment, feed conversion was significantly decreased (P <0.05) in CMN1 and CMN2 treatments. CMN1 administration significantly improved (P <0.05) the pH24 of muscle. The abnormal changes of serum malonaldehyde and corticosterone concentrations were prevented (P <0.05) by curcumin. Mitochondrial GSH concentration in the liver was significantly increased (P <0.05) in CMN1 and CMN2 treatments compared with the HS treatment. The CMN1, CMN2, and CMN3 supplementations significantly increased (P <0.05) γ-GCL, GSH-Px, and GST activities. Curcumin significantly increased (P <0.05) the expression of Nrf2, HO-1, and γ-GCLc in the liver as compared to the CON diet. The expression of Cu/ZnSOD and CAT were increased (P <0.05) by feeding CMN2, respectively, as compared to the HS treatment. It was concluded that curcumin supplementation enhanced the resistance of broilers to heat stress, as evidenced by reversing the FC, increasing the GSH content and GSH-related enzyme activities, and inducing the expression of Nrf2 and Nrf2-mediated phase II detoxifying enzyme genes.

C60 Fullerene Prevents Restraint Stress-Induced Oxidative Disorders in Rat Tissues: Possible Involvement of the Nrf2/ARE-Antioxidant Pathway.

The effects of C60FAS (50 and 500 μg/kg) supplementation, in a normal physiological state and after restraint stress exposure, on prooxidant/antioxidant balance in rat tissues were explored and compared with the effects of the known exogenous antioxidant N-acetylcysteine. Oxidative stress biomarkers (ROS, O2·−, H2O2, and lipid peroxidation) and indices of antioxidant status (MnSOD, catalase, GPx, GST, γ-GCL, GR activities, and GSH level) were measured in the brain and the heart. In addition, protein expression of Nrf2 in the nuclear and cytosol fractions as well as the protein level of antiradical enzyme MnSOD and GSH-related enzymes γ-GCLC, GPx, and GSTP as downstream targets of Nrf2 was evaluated by western blot analysis. Under a stress condition, C60FAS attenuates ROS generation and O2·− and H2O2 releases and thus decreases lipid peroxidation as well as increases rat tissue antioxidant capacity. We have shown that C60FAS supplementation has dose-dependent and tissue-specific effects. C60FAS strengthened the antiradical defense through the upregulation of MnSOD in brain cells and maintained MnSOD protein content at the control level in the myocardium. Moreover, C60FAS enhanced the GSH level and the activity/protein expression of GSH-related enzymes. Correlation of these changes with Nrf2 protein content suggests that under stress exposure, along with other mechanisms, the Nrf2/ARE-antioxidant pathway may be involved in regulation of glutathione homeostasis. In our study, in an in vivo model, when C60FAS (50 and 500 μg/kg) was applied alone, no significant changes in Nrf2 protein expression as well as in activity/protein levels of MnSOD and GSH-related enzymes in both tissues types were observed. All these facts allow us to assume that in the in vivo model, C60FAS affects on the brain and heart endogenous antioxidative statuses only during the oxidative stress condition.

Glutathione homeostasis is significantly altered by quercetin via the Keap1/Nrf2 and MAPK signaling pathways in rats

Previously, we showed that 0.5% quercetin simultaneously decreased serum homocysteine and glutathione (GSH) levels in rats. The aim of the present study was to investigate the effects of 0.5% quercetin on GSH metabolism, related enzymes and signal pathways in rats. Rats were fed the control diet and 0.5% quercetin-supplemented diet for 6 weeks. The results showed that quercetin reduced serum and hepatic content of GSH and the ratio of GSH and oxidized glutathione (GSSG), enhanced hepatic activity and mRNA expression of glutathione S-transferase (GST), inhibited hepatic activity and mRNA expression of glutamate cysteine ligase (GCL), and decreased hepatic glutathione reductase (GR) mRNA expression. Levels of phosphorylated p38 and extracellular signal-regulated kinase (ERK) 1/2 mitogen-activated protein kinases (MAPKs) increased, while that of nuclear factor E2-like 2 (Nrf2) protein decreased after quercetin treatment. However, no significant hepatotoxicity was noted. We concluded that quercetin treatment altered hepatic GSH metabolism by modulating GSH metabolic enzyme activities and mRNA expression in rats, and p38, ERK1/2 MAPKs, and Nrf2 were involved in modulating GSH metabolism-related enzymes.

Glutathione reductase mediates drug resistance in glioblastoma cells by regulating redox homeostasis

Glutathione (GSH) and GSH-related enzymes constitute the most important defense system that protects cells from free radical, radiotherapy, and chemotherapy attacks. In this study, we aim to explore the potential role and regulatory mechanism of the GSH redox cycle in drug resistance in glioblastoma multiforme (GBM) cells. We found that temozolomide (TMZ)-resistant glioma cells displayed lower levels of endogenous reactive oxygen species and higher levels of total antioxidant capacity and GSH than sensitive cells. Moreover, the expression of glutathione reductase (GSR), the key enzyme of the GSH redox cycle, was higher in TMZ-resistant cells than in sensitive cells. Furthermore, silencing GSR in drug-resistant cells improved the sensitivity of cells to TMZ or cisplatin. Conversely, the over-expression of GSR in sensitive cells resulted in resistance to chemotherapy. In addition, the GSR enzyme partially prevented the oxidative stress caused by pro-oxidant L-buthionine -sulfoximine. The modulation of redox state by GSH or L-buthionine -sulfoximine regulated GSR-mediated drug resistance, suggesting that the action of GSR in drug resistance is associated with the modulation of redox homeostasis. Intriguingly, a trend toward shorter progress-free survival was observed among GBM patients with high GSR expression. These results indicated that GSR is involved in mediating drug resistance and is a potential target for improving GBM treatment.

The antidepressant- and anxiolytic-like effects of fluoxetine and clozapine in chronically isolated rats involve inhibition of hippocampal TNF-α

Brain oxidative stress and neuroinflammation are implicated in psychiatric disorders. Thus, it is important to investigate the effects of individual psychotropic agents on antioxidative defense and proinflammatory mediators in brain regions associated with these disorders. Psychosocial stress is recognized as a threat to mental health, and the hippocampus is a primary target of stress-related damage. Chronic social isolation (CSIS) is a mild psychosocial stress used to model the pathophysiology of depression. We examined the antioxidative and anti-inflammatory potential of the antidepressant fluoxetine (FLX) and atypical antipsychotic clozapine (CLZ) in the hippocampus in the CSIS model of depression. We measured the effects of FLX and CLZ on depressive- and anxiety-like behaviors in non-stressed rats and rats exposed to 21d of CSIS. We further evaluated the content of reduced glutathione (GSH), the protein expression and activity of the GSH-related enzymes, the subcellular localization of nuclear factor-kappa B (NF-κB) and protein levels of proinflammatory mediators cyclooxygenase-2 (COX-2), interleukin-1beta (IL-1β), and tumor necrosis factor-alpha (TNF-α) in these groups of rats. CSIS resulted in an increase in depressive- and anxiety-like behaviors that corresponded with compromised glutathione peroxidase (GPx)-mediated antioxidative defense and increased TNF-α, but not with changes in NF-κB, IL-1β and COX-2 levels. FLX and CLZ, applied during CSIS, prevented the behavioral changes associated with CSIS, and inhibited the increase in TNF-α, but did not affect GPx-mediated antioxidative defense. Furthermore, both drugs decreased hippocampal GPx activity when applied to non-stressed rats. These results emphasize the significance of hippocampal TNF-α-mediated proinflammmatory signaling in the pathophysiology of depressive symptoms and the importance of the anti-inflammatory action of both FLX and CLZ in the prevention of these symptoms.

Free radical production and antioxidant status in brain cortex non-synaptic mitochondria and synaptosomes at alcohol hangover onset

Alcohol hangover (AH) is the pathophysiological state after a binge-like drinking. We have previously demonstrated that AH induced bioenergetics impairments in a total fresh mitochondrial fraction in brain cortex and cerebellum. The aim of this work was to determine free radical production and antioxidant systems in non-synaptic mitochondria and synaptosomes in control and hangover animals. Superoxide production was not modified in non-synaptic mitochondria while a 17.5% increase was observed in synaptosomes. A similar response was observed for cardiolipin content as no changes were evidenced in non-synaptic mitochondria while a 55% decrease in cardiolipin content was found in synaptosomes. Hydrogen peroxide production was 3-fold increased in non-synaptic mitochondria and 4-fold increased in synaptosomes. In the presence of deprenyl, synaptosomal H2O2 production was 67% decreased in the AH condition. Hydrogen peroxide generation was not affected by deprenyl addition in non-synaptic mitochondria from AH mice. MAO activity was 57% increased in non-synaptic mitochondria and 3-fold increased in synaptosomes. Catalase activity was 40% and 50% decreased in non-synaptic mitochondria and synaptosomes, respectively. Superoxide dismutase was 60% decreased in non-synaptic mitochondria and 80% increased in synaptosomal fractions. On the other hand, GSH (glutathione) content was 43% and 17% decreased in synaptosomes and cytosol. GSH-related enzymes were mostly affected in synaptosomes fractions by AH condition. Acetylcholinesterase activity in synaptosomes was 11% increased due to AH. The present work reveals that AH provokes an imbalance in the cellular redox homeostasis mainly affecting mitochondria present in synaptic terminals.