Showing papers on "Glutathione published in 2000"

Oxidative stress in bacteria and protein damage by reactive oxygen species.

Abstract: The advent of O2 in the atmosphere was among the first major pollution events occurred on earth. The reaction between ferrous iron, very abundant in the reductive early atmosphere, and oxygen results in the formation of harmful superoxide and hydroxyl radicals, which affect all macromolecules (DNA, lipids and proteins). Living organisms have to build up mechanisms to protect themselves against oxidative stress, with enzymes such as catalase and superoxide dismutase, small proteins like thioredoxin and glutaredoxin, and molecules such as glutathione. Bacterial genetic responses to oxidative stress are controlled by two major transcriptional regulators (OxyR and SoxRS). This paper reviews major key points in the generation of reactive oxygen species in bacteria, defense mechanisms and genetic responses to oxidative stress. Special attention is paid to the oxidative damage to proteins.

Phytochelatins and their roles in heavy metal detoxification

Abstract: Plants respond to heavy metal toxicity in a variety of different ways. Such responses include immobilization, exclusion, chelation and compartmentalization of the metal ions, and the expression of more general stress response mechanisms such as ethylene and stress proteins. These mechanisms have

Glutathione, oxidative stress and neurodegeneration.

Abstract: There is significant evidence that the pathogenesis of several neurodegenerative diseases, including Parkinson’s disease, Alzheimer’s disease, Friedreich’s ataxia and amyotrophic lateral sclerosis, may involve the generation of reactive oxygen species and mitochondrial dysfunction. Here, we review the evidence for a disturbance of glutathione homeostasis that may either lead to or result from oxidative stress in neurodegenerative disorders. Glutathione is an important intracellular antioxidant that protects against a variety of different antioxidant species. An important role for glutathione was proposed for the pathogenesis of Parkinson’s disease, because a decrease in total glutathione concentrations in the substantia nigra has been observed in preclinical stages, at a time at which other biochemical changes are not yet detectable. Because glutathione does not cross the blood‐brain barrier other treatment options to increase brain concentrations of glutathione including glutathione analogs, mimetics or precursors are discussed.

Mice Deficient in Cellular Glutathione Peroxidase Show Increased Vulnerability to Malonate, 3-Nitropropionic Acid, and 1-Methyl-4-Phenyl-1,2,5,6-Tetrahydropyridine

Abstract: Glutathione peroxidase (GSHPx) is a critical intracellular enzyme involved in detoxification of hydrogen peroxide (H 2 O 2 ) to water. In the present study we examined the susceptibility of mice with a disruption of the glutathione peroxidase gene to the neurotoxic effects of malonate, 3-nitropropionic acid (3-NP), and 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP). Glutathione peroxidase knock-out mice showed no evidence of neuropathological or behavioral abnormalities at 2–3 months of age. Intrastriatal injections of malonate resulted in a significant twofold increase in lesion volume in homozygote GSHPx knock-out mice as compared to both heterozygote GSHPx knock-out and wild-type control mice. Malonate-induced increases in conversion of salicylate to 2,3- and 2,5-dihydroxybenzoic acid, an index of hydroxyl radical generation, were greater in homozygote GSHPx knock-out mice as compared with both heterozygote GSHPx knock-out and wild-type control mice. Administration of MPTP resulted in significantly greater depletions of dopamine, 3,4-dihydroxybenzoic acid, and homovanillic acid in GSHPx knock-out mice than those seen in wild-type control mice. Striatal 3-nitrotyrosine (3-NT) concentrations after MPTP were significantly increased in GSHPx knock-out mice as compared with wild-type control mice. Systemic 3-NP administration resulted in significantly greater striatal damage and increases in 3-NT in GSHPx knock-out mice as compared to wild-type control mice. The present results indicate that a knock-out of GSHPx may be adequately compensated under nonstressed conditions, but that after administration of mitochondrial toxins GSHPx plays an important role in detoxifying increases in oxygen radicals.

Glutathione S-Transferase Polymorphisms and Their Biological Consequences

Abstract: Two supergene families encode proteins with glutathione S-transferase (GST) activity: the family of soluble enzymes comprises at least 16 genes; the separate family of microsomal enzymes comprises at

Oxidative stress and regulation of glutathione in lung inflammation

Abstract: Inflammatory lung diseases are characterized by chronic inflammation and oxidant/antioxidant imbalance, a major cause of cell damage. The development of an oxidant/antioxidant imbalance in lung inflammation may activate redox-sensitive transcription factors such as nuclear factor-KB, and activator protein-1 (AP-1), which regulate the genes for pro-inflammatory mediators and protective antioxidant genes. Glutathione (GSH), a ubiquitous tripeptide thiol, is a vital intra- and extracellular protective antioxidant against oxidative/nitrosative stresses, which plays a key role in the control of pro-inflammatory processes in the lungs. Recent findings have suggested that GSH is important in immune modulation, remodelling of the extracellular matrix, apoptosis and mitochondrial respiration. The rate-limiting enzyme in GSH synthesis is gamma-glutamylcysteine synthetase (gamma-GCS). The human gamma-GCS heavy and light subunits are regulated by AP-1 and antioxidant response elements and are modulated by oxidants, phenolic antioxidants, growth factors, and inflammatory and anti-inflammatory agents in lung cells. Alterations in alveolar and lung GSH metabolism are widely recognized as a central feature of many inflammatory lung diseases such as idiopathic pulmonary fibrosis, acute respiratory distress syndrome, cystic fibrosis and asthma. The imbalance and/or genetic variation in antioxidant gamma-GCS and pro-inflammatory versus antioxidant genes in response to oxidative stress and inflammation in some individuals may render them more susceptible to lung inflammation. Knowledge of the mechanisms of GSH regulation and balance between the release and expression of pro- and anti-inflammatory mediators could lead to the development of novel therapies based on the pharmacological manipulation of the production as well as gene transfer of this important antioxidant in lung inflammation and injury. This review describes the redox control and involvement of nuclear factor-kappaB and activator protein-1 in the regulation of cellular glutathione and gamma-glutamylcysteine synthetase under conditions of oxidative stress and inflammation, the role of glutathione in oxidant-mediated susceptibility/tolerance, gamma-glutamylcysteine synthetase genetic susceptibility and the potential therapeutic role of glutathione and its precursors in protecting against lung oxidant stress, inflammation and injury.

Tolerance of pea (Pisum sativum L.) to long‐term salt stress is associated with induction of antioxidant defences

Abstract: Using two cultivars of Pisum sativum L. with different sensitivity to NaCl, the effect of long-term (15 d) NaCl (70 m M) treatments on the activity and expression of the foliar ascorbate–glutathione cycle enzymes, superoxide dismutase isozymes and their mRNAs was evaluated and related to their ascorbate and glutathione contents. High-speed supernatant (soluble) fractions, enriched for cytosolic components of the antioxidant system, were used. In this fraction from the NaCl-tolerant variety (cv Granada), the activities of ascorbate peroxidase (APX), glutathione reductase (GR), monodehydroascorbate reductase (MDHAR), Mn-superoxide dismutase (Mn-SOD) and dehydroascorbate reductase (DHAR) increased, while CuZn-SOD activity remained constant. In the NaCl-sensitive plants (cv Challis), salinity did not produce significant changes in APX, MDHAR and GR activities. Only DHAR activity was induced in cv Challis, whereas soluble CuZn-SOD activity decreased by about 35%. Total ascorbate and glutathione contents decreased in both cultivars, but the decline was greater in NaCl-sensitive plants. This difference between the two cultivars was more pronounced when the transcript levels of some these enzymes were examined. Transcript levels for mitochondrial Mn-SOD, chloroplastic CuZn-SOD and phospholipid hydroperoxide glutathione peroxidase (PHGPX), cytosolic GR and APX were strongly induced in the NaCl-tolerant variety but not in the NaCl-sensitive variety. These data strongly suggest that induction of antioxidant defences is at least one component of the tolerance mechanism of peas to long-term salt-stress.

Glutathione metabolism in brain metabolic interaction between astrocytes and neurons in the defense against reactive oxygen species.

Abstract: The cells of the adult human brain consume approximately 20% of the oxygen utilized by the body although the brain comprises only 2% of the body weight. Reactive oxygen species, which are produced continuously during oxidative metabolism, are generated at high rates within the brain. Therefore, the defense against the toxic effects of reactive oxygen species is an essential task within the brain. An important component of the cellular detoxification of reactive oxygen species is the antioxidant glutathione. The main focus of this short review is recent results on glutathione metabolism of brain astrocytes and neurons in culture. These two types of cell prefer different extracellular precursors for glutathione. Glutathione is involved in the disposal of exogenous peroxides by astrocytes and neurons. In coculture astrocytes protect neurons against the toxicity of reactive oxygen species. One mechanism of this interaction is the supply by astrocytes of glutathione precursors to neurons.

Roles of the Glutathione- and Thioredoxin-Dependent Reduction Systems in the Escherichia Coli and Saccharomyces Cerevisiae Responses to Oxidative Stress

Abstract: The glutathione- and thioredoxin-dependent reduction systems are responsible for maintaining the reduced environment of the Escherichia coli and Saccharomyces cerevisiae cytosol. Here we examine the roles of these two cellular reduction systems in the bacterial and yeast defenses against oxidative stress. The transcription of a subset of the genes encoding glutathione biosynthetic enzymes, glutathione reductases, glutaredoxins, thioredoxins, and thioredoxin reductases, as well as glutathione- and thioredoxin-dependent peroxidases is clearly induced by oxidative stress in both organisms. However, only some strains carrying mutations in single genes are hypersensitive to oxidants. This is due, in part, to the redundant effects of the gene products and the overlap between the two reduction systems. The construction of strains carrying mutations in multiple genes is helping to elucidate the different roles of glutathione and thioredoxin, and studies with such strains have recently revealed that these two reduction systems modulate the activities of the E. coli OxyR and SoxR and the S. cerevisiae Yap1p transcriptional regulators of the adaptive responses to oxidative stress.

Levels of antioxidant defenses are decreased in bovine spermatozoa after a cycle of freezing and thawing.

Abstract: Growing evidence suggests that the generation of reactive oxygen species (ROS) and their detoxification by antioxidants plays a very important role in fertility. However, the relationship between the level of antioxidants in spermatozoa and the decreased fecundity following a freeze/thaw cycle remains poorly understood. We assessed the activities of antioxidant enzymes such as catalase, glutathione peroxidase (GPx), superoxide dismutase (SOD), and levels of reduced/oxidized glutathione (GSH/GSSG) in bovine semen. Sperm cells were isolated using a Percoll gradient to avoid contamination from seminal plasma, cellular debris, and other cell types. We found that bovine spermatozoa are poorly adapted to metabolize the toxic hydrogen peroxide (H(2)O(2)). Indeed, very low levels of GPx and an absence of catalase were observed. We also studied the effect of freezing and thawing bovine spermatozoa in a egg yolk-Tris-glycerol extender (EYTG). Cryopreservation significantly reduced sperm GSH levels by 78% and SOD activity by 50%. We also investigated whether the decrease in GSH level could be linked to oxidative metabolism and found that a greater reduction in intracellular GSH level occurred when fresh sperm cells were incubated in EYTG for 6 hr at 38.5 degrees C under aerobic conditions than when incubated under restricted oxygen availability. Our results strongly suggest the involvement of an oxidative stress during a freeze/thaw cycle and are consistent with the hypothesis that ROS generated during such a cycle are detrimental to sperm function.

Schizophrenia: glutathione deficit in cerebrospinal fluid and prefrontal cortex in vivo.

Abstract: Schizophrenia is a major psychiatric disease, which affects the centre of the personality, with severe problems of perception, cognition as well as affective and social behaviour. In cerebrospinal fluid of drug-free schizophrenic patients, a significant decrease in the level of total glutathione (GSH) by 27% (P<0.05) was observed as compared to controls, in keeping with the reported reduced level of its metabolite gamma-glutamylglutamine. With a new non-invasive proton magnetic resonance spectroscopy methodology, GSH level in medial prefrontal cortex of schizophrenic patients was found to be 52% (P = 0.0012) lower than in controls. GSH plays a fundamental role in protecting cells from damage by reactive oxygen species generated among others by the metabolism of dopamine. A deficit in GSH would lead to degenerative processes in the surrounding of dopaminergic terminals resulting in loss of connectivity. GSH also potentiates the N-methyl-D-aspartate (NMDA) receptor response to glutamate, an effect presumably reduced by a GSH deficit, leading to a situation similar to the application of phencyclidine (PCP). Thus, a GSH hypothesis might integrate many established biological aspects of schizophrenia.

The Quantitatively Important Relationship between Homocysteine Metabolism and Glutathione Synthesis by the Transsulfuration Pathway and Its Regulation by Redox Changes

Abstract: Homocysteine is a key junction metabolite in methionine metabolism. It suffers two major metabolic fates: transmethylation catalyzed by methionine synthase or betaine homocysteine methyl transferase and transsulfuration catalyzed by cystathionine beta-synthase leading to cystathionine. The latter is subsequently converted to cysteine, a precursor of glutathione. Studies with purified mammalian methionine synthase and cystathionine beta-synthase have revealed the oxidative sensitivity of both junction enzymes, suggesting the hypothesis that redox regulation of this pathway may be physiologically significant. This hypothesis has been tested in a human hepatoma cell line in culture in which the flux of homocysteine through transsulfuration under normoxic and oxidative conditions has been examined. Addition of 100 microM H(2)O(2) or tertiary butyl hydroperoxide increased cystathionine production 1.6- and 2.1-fold from 82 +/- 7 micromol h(-)(1) (L of cells)(-)(1) to 136 +/- 15 and 172 +/- 23 micromol h(-)(1) (L of cells)(-)(1), respectively. The increase in homocysteine flux through the transsulfuration pathway exhibited a linear dose dependence on the concentrations of both oxidants (50-200 microM H(2)O(2) and 10-200 microM tertiary butyl hydroperoxide). Furthermore, our results reveal that approximately half of the intracellular glutathione pool in human liver cells is derived from homocysteine via the transsulfuration pathway. The redox sensitivity of the transsulfuration pathway can be rationalized as an autocorrective response that leads to an increased level of glutathione synthesis in cells challenged by oxidative stress. In summary, this study demonstrates the importance of the homocysteine-dependent transsulfuration pathway in the maintenance of the intracellular glutathione pool, and the regulation of this pathway under oxidative stress conditions. Aberrations in this pathway could compromise the redox buffering capacity of cells, which may in turn be related to the pathophysiology of the different homocysteine-related diseases.

Stress tolerance in transgenic tobacco seedlings that overexpress glutathione S-transferase/glutathione peroxidase.

Abstract: Overexpression of a tobacco glutathione S-transferase with glutathione peroxidase activity (GST/GPX) in transgenic tobacco (Nicotiana tabacum L.) enhanced seedling growth under a variety of stressful conditions. In addition to increased GST and GPX activity, transgenic GST/GPX-expressing (GST+) seedlings had elevated levels of monodehydroascorbate reductase activity. GST+ seedlings also contained higher levels of glutathione and ascorbate than wild-type seedlings and the glutathione pools were more oxidized. Thermal or salt-stress treatments that inhibited the growth of wild-type seedlings also caused increased levels of lipid peroxidation. These treatments had less effect on the growth of GST+ seedling growth and did not lead to increased lipid peroxidation. Stress-induced damage resulted in reduced metabolic activity in wild-type seedlings while GST+ seedlings maintained metabolic activity levels comparable to seedlings grown under control conditions. These results indicate that overexpression of GST/GPX in transgenic tobacco seedlings provides increased glutathione-dependent peroxide scavenging and alterations in glutathione and ascorbate metabolism that lead to reduced oxidative damage. We conclude that this protective effect is primarily responsible for the ability of GST+ seedlings to maintain growth under stressful conditions.

Energy dissipation and radical scavenging by the plant phenylpropanoid pathway.

Abstract: Environmental stresses such as high light, low temperatures, pathogen infection and nutrient deficiency can lead to increased production of free radicals and other oxidative species in plants. A growing body of evidence suggests that plants respond to these biotic and abiotic stress factors by increasing their capacity to scavenge reactive oxygen species. Efforts to understand this acclimatory process have focused on the components of the 'classical' antioxidant system, i.e. superoxide dismutase, ascorbate peroxidase, catalase, monodehydroascorbate reductase, glutathione reductase and the low molecular weight antioxidants ascorbate and glutathione. However, relatively few studies have explored the role of secondary metabolic pathways in plant response to oxidative stress. A case in point is the phenylpropanoid pathway which is responsible for the synthesis of a diverse array of phenolic metabolites such as flavonoids, tannins, hydroxycinnamate esters and the structural polymer lignin. These compounds are often induced by stress and serve specific roles in plant protection, i.e. pathogen defence, ultraviolet screening, antiherbivory, or structural components of the cell wall. This review will highlight a novel antioxidant function for the taxonomically widespread phenylpropanoid metabolite chlorogenic acid (CGA; 5-O-caffeoylquinic acid) and assess its possible role in abiotic stress tolerance. The relationship between CGA biosynthesis and photosynthetic carbon metabolism will also be discussed. Based on the properties of this model phenolic metabolite, we propose that under stress conditions phenylpropanoid biosynthesis may represent an alternative pathway for photochemical energy dissipation that has the added benefit of enhancing the antioxidant capacity of the cell.

Induction of Oxidative Stress by Glutathione Depletion Causes Severe Hypertension in Normal Rats

Abstract: Several recent studies have shown that certain forms of genetic or acquired hypertension are associated with oxidative stress and that animals with those types of hypertension respond favorably to antioxidant therapy. We hypothesize that oxidative stress may cause hypertension via (among other mechanisms) enhanced oxidation and inactivation of nitric oxide (NO). To test this hypothesis, Sprague-Dawley rats were subjected to oxidative stress by glutathione (GSH) depletion by means of the GSH synthase inhibitor buthionine sulfoximine (BSO, 30 mmol/L in drinking water) for 2 weeks. The control group was given drug-free drinking water. In parallel experiments, subgroups of animals were provided vitamin E-fortified chow and vitamin C-supplemented drinking water. The BSO-treated group showed a 3-fold decrease in tissue GSH content, a marked elevation in blood pressure, and a significant reduction in the urinary excretion of the NO metabolite nitrate plus nitrite, which suggests depressed NO availability. These characteristics were associated with a significant accumulation in various tissues of nitrotyrosine, which is the footprint of NO inactivation by reactive oxygen species. Administration of vitamin E plus vitamin C ameliorated hypertension, improved urinary nitrate-plus-nitrite excretion, and mitigated nitrotyrosine accumulation (despite GSH depletion) in the BSO-treated animals but had no effect in the control group. In conclusion, GSH depletion resulted in perturbation of the NO system and severe hypertension in normal animals. The effects of BSO were mitigated by concomitant antioxidant therapy despite GSH depletion, which supports the notion that oxidative stress was involved in the pathogenesis of hypertension in this model.

Thiol-based antioxidants

Abstract: The thiol redox status of intracellular and extracellular compartments is critical in the determination of protein structure, regulation of enzyme activity, and control of transcription factor activity and binding. Thiol antioxidants act through a variety of mechanisms, including (1) as components of the general thiol/disulfide redox buffer, (2) as metal chelators, (3) as radical quenchers, (4) as substrates for specific redox reactions (GSH), and (5) as specific reductants of individual protein disulfate bonds (thioredoxin). The composition and redox status of the available thiols in a given compartment is highly variable and must play a part in determining the metabolic activity of each compartment. It is generally beneficial to increase the availability of specific antioxidants under conditions of oxidant stress. Cells have devised a number of mechanisms to promote increased intracellular levels of thiols such as GSH and thioredoxin in response to a wide variety of stresses. Exogenous thiols have been used successfully to increase cell and tissue thiol levels in cell cultures, in animal models, and in humans. Increased levels of GSH and other thiols have been associated with increased tolerance to oxidant stresses in all of these systems and in some cases, with disease prevention or treatment in humans. A wide variety of thiol-related compounds have been used for these purposes. These include thiols such as GSH and its derivatives, cysteine and NAC, dithiols such as lipoic acid, which is reduced to the thiol form intracellularly, and "prothiol" compounds such as OTC, which are enzymatically converted to free thiols within the cell. In choosing a thiol for a specific function (e.g., protection of lung from oxidant exposure or protection of organs from ischemia reperfusion injury), the global effects must also be considered. For example, large increases in free thiols in the circulation are associated with toxic effects. These effects may be the result of thiyl radical-mediated reactions but could also be due to destabilizing effects of increases in thiol/disulfide ratios in the plasma, which normally is in a more oxidized state than intracellular compartments. Changes in the thiol redox gradient across cells could also adversely affect any transport or cell signaling processes, which are dependent on formation and rupture of disulfide linkages in membrane proteins. Therapeutic thiol administration has been shown to have great potential, and its efficacy should be increased by selecting compounds and methods of delivery that will minimize perturbations in the thiol status of regions external to the targeted areas.

Chemoprotection against cancer by induction of Phase 2 enzymes

Abstract: Induction of Phase 2 enzymes is an effective and sufficient strategy for achieving protection against the toxic and neoplastic effects of many carcinogens. It is proposed that the concept of Phase 2 enzymes as being responsible only for the conjugation of functionalized xenobiotics with endogenous cellular ligands such as glutathione (glutathione S-transferases) and glucuronic acid (UDP-glucuronosyltransferases) be expanded to include proteins with the following common characteristics: (a) coordinate induction by a broad range of chemical agents that all have the capacity to react with sulfhydryl groups; (b) possible regulation by common promoter elements; and (c) catalysis of reactions that lead to comprehensive protection against electrophile and reactive oxygen toxicities, by a wide variety of mechanisms. These mechanisms include: conjugation with endogenous ligands, chemical modification of reactive features of molecules that can damage DNA and other macromolecules, and generation or augementation of cellular antioxidants. In addition to the above conjugating enzymes, a provisional and partial list of Phase 2 proteins might include: NAD(P)H:quinone reductase, epoxide hydrolase, dihydrodiol dehydrogenase, gamma-glutamylcysteine synthetase, heme oxygenase-1, leukotriene B4 dehydrogenase, aflatoxin B1 dehydrogenase, and ferritin.

AN9, a petunia glutathione S-transferase required for anthocyanin sequestration, is a flavonoid-binding protein.

Abstract: AN9 is a glutathione S-transferase from petunia (Petunia hybrida) required for efficient anthocyanin export from the site of synthesis in the cytoplasm into permanent storage in the vacuole. For many xenobiotics it is well established that a covalent glutathione (GSH) tag mediates recognition of molecules destined for vacuolar sequestration by a tonoplast-localized ATP-binding cassette pump. Here we inquired whether AN9 catalyzes the formation of GSH conjugates with flavonoid substrates. Using high-performance liquid chromatography analysis of reaction mixtures containing enzyme, GSH, and flavonoids, including anthocyanins, we could detect neither conjugates nor a decrease in the free thiol concentration. These results suggest that no conjugate is formed in vitro. However, AN9 was shown to bind flavonoids using three assays: inhibition of the glutathione S-transferase activity of AN9 toward the common substrate 1-chloro 2,4-dinitrobenzene, equilibrium dialysis, and tryptophan quenching. We conclude that AN9 is a flavonoid-binding protein, and propose that in vivo it serves as a cytoplasmic flavonoid carrier protein.

Glutathione: a vital lens antioxidant.

Abstract: The reducing compound glutathione (GSH) exists in an unusually high concentration in the lens where it functions as an essential antioxidant vital for maintenance of the tissue's transparency. In conjunction with an active glutathione redox cycle located in the lens epithelium and superficial cortex, GSH detoxifies potentially damaging oxidants such as H2O2 and dehydroascorbic acid. Recent studies have indicated an important hydroxyl radical-scavenging function for GSH in lens epithelial cells, independent of the cells' ability to detoxify H2O2. Depletion of GSH or inhibition of the redox cycle allows low levels of oxidant to damage lens epithelial targets such as Na/K-ATPase, certain cytoskeletal proteins and proteins associated with normal membrane permeability. The level of GSH in the nucleus of the lens is relatively low, particularly in the aging lens, and exactly how the compound travels from the epithelium to the central region of the organ is not known. Recently, a cortical/nuclear barrie...