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Abstract

The immune system works best if the lymphoid cells have a delicately balanced intermediate level of glutathione. Even moderate changes in the intracellular glutathione level have profound effects on lymphocyte functions. Certain functions, such as the DNA synthetic response, are exquisitely sensitive to reactive oxygen intermediates and, therefore, are favoured by high levels of the antioxidant glutathione. Certain signal pathways, in contrast, are enhanced by oxidative conditions and favoured by low intracellular glutathione levels. The available evidence suggests that the lymphocytes from healthy human subjects have, on average, an optimal glutathione level. There is no indication that immunological functions such as resistance to infection or the response to vaccination may be enhanced in healthy human subjects by administration of glutathione or its precursor amino acid cysteine. However, immunological functions in diseases that are associated with a cysteine and glutathione deficiency may be significantly enhanced and potentially restored by cysteine supplementation. This factor has been studied most extensively in the case of human immunodeficiency virus (HIV)-infected patients who were found to experience, on average, a massive loss of S equivalent to a net loss of approximately 4 g cysteine/d. Two randomized placebo-controlled trials have shown that treatment of HIV-infected patients with N-acetyl-cysteine caused in both cases a significant increase in all immunological functions under test, including an almost complete restoration of natural killer cell activity. It remains to be tested whether cysteine supplementation may be useful also in other diseases and conditions that are associated with a low mean plasma cystine level and impaired immunological functions.
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... Glutathione (GSH) is a major intracellular antioxidant capable of scavenging free radicals and detoxifying electrophiles from endogenous and exogenous sources via the free thiol group. Several studies have investigated GSH efficacy in immune diseases [22][23][24][25][26], but no studies have been published regarding its use in CAD, HLH, and PMR. We tested GSH prophylactic antibiotics, and a prednisone taper for long-term management. ...
... Nevertheless, these levels were lower in severe AD patients in comparison to mAD subjects. The glutathione cycle is one of the main intracellular mechanisms to preserve a competent intracellular redox state (70). The role of this has been extensively studied in AD, in both humans and animal models, where a depletion in GSH contents and an imbalance in GSSG/GSH ratios in favor of GSSG have been noted at central (39) and peripheral levels, such as in plasma, peritoneal leukocytes, and blood cells (47,52,71). ...
Article
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Since aging is considered the most risk factor for sporadic Alzheimer’s Disease (AD), the age-related impairment of the immune system (immunosenescence), based on a chronic oxidative-inflammatory stress situation, could play a key role in the development and progression of AD. Although AD is accompanied by systemic disturbance, reflecting the damage in the brain, the changes in immune response and redox-state in different types of blood cells in AD patients have been scarcely studied. The aim was to analyze the variations in several immune functions and oxidative-inflammatory stress and damage parameters in both isolated peripheral neutrophils and mononuclear blood cells, as well as in whole blood cells, from patients diagnosed with mild (mAD) and severe AD, and of age-matched controls (elderly healthy subjects) as well as of adult controls. The cognitive decline of all subjects was determined by Mini-Mental State Examination (MMSE) test (mAD stage was established at 20 ≤ MMSE ≤ 23 score; AD stage at <18 MMSE; elderly subjects >27 MMSE). The results showed an impairment of the immune functions of human peripheral blood neutrophils and mononuclear cells of mAD and AD patients in relation to healthy elderly subjects, who showed the typical immunosenescence in comparison with the adult individuals. However, several alterations were only observed in severe AD patients (lower chemotaxis, lipopolysaccharide lymphoproliferation, and interleukin (IL)-10 release; higher basal proliferation, tumor necrosis factor (TNF)-α release, and IL-10/TNF-α ratio), others only in mAD subjects (higher adherence), meanwhile others appeared in both mAD and AD patients (lower phytohemaglutinin lymphoproliferation and higher IL-6 release). This impairment of immune functions could be mediated by: (1) the higher oxidative stress and damage also observed in blood cells from mAD and AD patients and in isolated neutrophils [lower glutathione (GSH) levels, high oxidized glutathione (GSSG)/GSH ratio, and GSSG and malondialdehyde contents], and (2) the higher release of basal pro-inflammatory cytokines (IL-6 and TNF-α) found in AD patients. Because the immune system parameters studied are markers of health and rate of aging, our results supported an accelerated immunosenescence in AD patients. We suggest the assessment of oxidative stress and function parameters in peripheral blood cells as well as in isolated neutrophils and mononuclear cells, respectively, as possible markers of AD progression.
... For example, our group has shown that GSH treatment strongly inhibits viral replication by impairing glycoprotein folding (10); on the other hand, we have recently shown that GSH depletion increased influenza virus replication by preventing activation of innate antiviral response (7). Indeed, the role of GSH in modulating immune response is well known (8,(77)(78)(79). For example, in antigen-presenting cells, GSH depletion correlates with defective antigen processing and reduced secretion of T helper 1 (Th1) cytokines, thus favoring polarization from the typical Th1 profile toward a Th2 response (8). ...
Article
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Influenza virus replicates intracellularly exploiting several pathways involved in the regulation of host responses. The outcome and the severity of the infection are thus strongly conditioned by multiple host factors, including age, sex, metabolic, and redox conditions of the target cells. Hormones are also important determinants of host immune responses to influenza and are recently proposed in the prophylaxis and treatment. This study shows that female mice are less susceptible than males to mouse-adapted influenza virus (A/PR8/H1N1). Compared with males, PR8-infected females display higher survival rate (+36%), milder clinical disease, and less weight loss. They also have milder histopathological signs, especially free alveolar area is higher than that in males, even if pro-inflammatory cytokine production shows slight differences between sexes; hormone levels, moreover, do not vary significantly with infection in our model. Importantly, viral loads (both in terms of viral M1 RNA copies and tissue culture infectious dose 50%) are lower in PR8-infected females. An analysis of the mechanisms contributing to sex disparities observed during infection reveals that the female animals have higher total antioxidant power in serum and their lungs are characterized by increase in (i) the content and biosynthesis of glutathione, (ii) the expression and activity of antioxidant enzymes (peroxiredoxin 1, catalase, and glutathione peroxidase), and (iii) the expression of the anti-apoptotic protein Bcl-2. By contrast, infected males are characterized by high expression of NADPH oxidase 4 oxidase and phosphorylation of p38 MAPK, both enzymes promoting viral replication. All these factors are critical for cell homeostasis and susceptibility to infection. Reappraisal of the importance of the host cell redox state and sex-related effects may be useful in the attempt to develop more tailored therapeutic interventions in the fight against influenza.
... Furthermore, GSH affects the production of most inflammatory cytokines and is required to maintain adequate interferon (IFN)-γ production by dendritic cells [82]. Based on these data, Dröge and Breitkreutz [83] claimed that the immune system is highly affected by the level of GSH in the body. However, for many years, it was not known whether disrupted GSH levels alter a complex ...
Article
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Glutathione is one of the most important and potent antioxidants. The development of pharmacological compounds that can either increase or decrease glutathione concentrations has allowed investigation into the role of glutathione in various biological processes, including immune responses. Recent findings have shown that glutathione not only affects certain factors involved in immunological processes but also modifies complex immune reactions such as fever. Until recently, it was not known why some patients do not develop fever during infection. Data suggest that fever induction is associated with oxidative stress; therefore, antioxidants such as glutathione can reduce pyrexia. Surprisingly, new studies have shown that low glutathione levels can also inhibit fever. In this review, we focus on recent advances in this area, with an emphasis on the role of glutathione in immune responses accompanied by fever. We describe evidence showing that disturbed glutathione homeostasis may be responsible for the lack of fever during infections. We also discuss the biological significance of the antipyretic effects produced by pharmacological glutathione modulators.
... Recent studies have found that serum concentrations of GSH are associated with various disease conditions (Droge and Breitkreutz, 2000;Prousky, 2008;Forman et al., 2009;Smeyne and Smeyne, 2013). For example, decreased serum concentration of GSH has been linked to cancer and neurodegenerative disease susceptibility (Smeyne and Smeyne, 2013). ...
Article
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There is an ongoing interest in cellular antioxidants and oxidants as well as cellular mechanisms underlying their effects. Several reports suggest that vitamin C (L-ascorbic acid) functions as a pro-oxidant with selective toxicity against specific types of tumor cells. In addition, reduced glutathione plays an emerging role in reducing oxidative stress due to xenobiotic toxins such as metals and oxidants associated with diseases such as cancer, cardiovascular disease, and stroke. High-dose intravenous vitamin C and intravenous glutathione have been used as complementary, alternative, and adjuvant medicines. Here, we review the molecular mechanisms underlying the regulation of oxidation/reduction systems, focusing on the altered metabolomics profile in cancer cells following treatment with pharmacological vitamin C. This review focuses on the role of vitamin C in energy metabolism in terms of adenosine triphosphate, cysteine, and reduced glutathione levels, affecting cancer cell death.
... It was found that continuous exposure to even low levels of oxidants can eventually cause depletion of reduced glutathione. 22 Upon treatment with an anti-inflammatory agent, infliximab, in RA, a short term decrease in protein carbonyl group levels were reported, and in the long-term treatment the levels were found to be similar to those of controls, suggesting the beneficial effects of infliximab on the protein oxidation process. Similarly, anti-tumor necrosis factor-α therapy induced a significant and sustained increase in thiol levels indicating improvement in antioxidant status, which correlated with a reduction in C-reactive protein concentrations. ...
Article
Background: Oxidative stress (OS) has an important role in the pathogenesis and progression of rheumatoid arthritis (RA). OS causes protein modification, thereby impairing the biological functions of the protein. This study was conducted to assess the oxidatively modified protein as protein carbonyl content and the antioxidant status as protein thiols, and to study the association between protein carbonyls and protein thiols in RA.Methods: Newly diagnosed RA patients who were not taking any disease modifying anti-rheumatic drugs were included into the study group (n=45) along with age and sex matched healthy controls (n=45). Serum protein carbonyl content and protein thiols were estimated.Results: Elevated protein carbonyl content and decreased protein thiol levels (p<0.001) were observed in RA. A significant negative correlation was observed between protein carbonyl content and protein thiol levels (p<0.001).Conclusions: Oxidative stress in RA is evidenced by enhanced protein oxidation and decreased antioxidant protein thiol levels. Decreased protein thiols may also reflect protein modifications leading to compromise in the antioxidant properties. This oxidant and antioxidant imbalance needs to be addressed by therapeutic interventions to prevent disease progression.
... Leukocytes are particularly sensitive to redox changes and changes in GSH levels (147)(148)(149)(150). The immune response starts with the activation of monocytes, macrophages, and antigen-presenting cells (APC). ...
Article
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Glutathione (γ-glutamyl-cysteinyl-glycine) is an intracellular thiol molecule and a potent antioxidant that participates in the toxic metabolism phase II biotransformation of xenobiotics. It can bind to a variety of proteins in a process known as glutathionylation. Protein glutathionylation is now recognised as one of important posttranslational regulatory mechanisms in cell and tissue physiology. Direct and indirect regulatory roles in physiological processes include glutathionylation of major transcriptional factors, eicosanoids, cytokines, and nitric oxide (NO). This review looks into these regulatory mechanisms through examples of glutathione regulation in apoptosis, vascularisation, metabolic processes, mitochondrial integrity, immune system, and neural physiology. The focus is on the physiological roles of glutathione beyond biotransformational metabolism.
... Glutathione (γ-L-glutamyl-L-cysteinyl-glycine, GSH), which is synthesized from glutamic acid (Glu), cysteine (Cys) and glycine (Gly), is the most abundant non-protein thiol compound in almost all organisms. Its unique structure of a γ-carboxyl of glutamate and a free sulfhydryl moiety of the Cys residue give this tripeptide a wide variety of biological activities, such as anti-oxidation 1 , detoxification 2,3 and immune regulation 4 . GSH plays a pivotal role in maintaining an appropriate redox environment for organisms and is used as a supplement in various pharmaceuticals 2 . ...
Article
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Acrolein (Acr) was used as a selection agent to improve the glutathione (GSH) overproduction of the prototrophic strain W303-1b/FGPPT. After two rounds of adaptive laboratory evolution (ALE), an unexpected result was obtained wherein identical GSH production was observed in the selected isolates. Then, a threshold selection mechanism of Acr-stressed adaption was clarified based on the formation of an Acr-GSH adduct, and a diffusion coefficient (0.36 ± 0.02 μmol·min-1·OD600-1) was calculated. Metabolomic analysis was carried out to reveal the molecular bases that triggered GSH overproduction. The results indicated that all three precursors (glutamic acid (Glu), glycine (Gly) and cysteine (Cys)) needed for GSH synthesis were at a relativity higher concentration in the evolved strain and that the accumulation of homocysteine (Hcy) and cystathionine might promote Cys synthesis and then improve GSH production. In addition to GSH and Cys, it was observed that other non-protein thiols and molecules related to ATP generation were at obviously different levels. To divert the accumulated thiols to GSH biosynthesis, combinatorial strategies, including deletion of cystathionine β-lyase (STR3), overexpression of cystathionine γ-lyase (CYS3) and cystathionine β-synthase (CYS4), and reduction of the unfolded protein response (UPR) through up-regulation of protein disulphide isomerase (PDI), were also investigated.
... Besides phase I, phase II mechanisms like GSH conjugation could be impaired due to lower levels of GSH found in people with certain health conditions. A study conducted by Droge and Breitkreutz (2000) showed that the immune system works best when the GSH levels of lymphoid cells are kept within optimal levels. The study showed that some HIV-infected patients lose the GSH precursor cysteine at more levels than a non-infected person. ...
Conference Paper
The Brazilian Unified Public Health System (SUS) shortlisted various plant species of interest (RENISUS) for future clinical use. However, very little is known about their effects on metabolic and transporter proteins, which could potentially lead to herb-drug interactions (HDI). To evaluate this, we conducted in vitro preclinical studies on twenty-four plant extracts to disclose their effects on CYP3A4 mRNA gene expression, intracellular glutathione (GSH) levels, inhibition of γ-glutamyl transferase (GGT) in HepG2 cells and P- glycoprotein (P-gp) activity in vincristine resistant Caco-2 (Caco-2 VCR) cells. We also investigated whether four Brazilian native species were able to activate the human pregnane X receptor (hPXR) in transiently co-transfected HeLa cells. This preclinical research showed that all but two plant extracts were able to modulate at least one of the selected targets. CYP3A4 mRNA gene expression in HepG2 cells was significantly affected by half of the extracts. The antagonistic effect of Solanum paniculatum L. on hPXR could explain its ability to inhibit CYP3A4. GSH levels were affected by 80% of the extracts. There was depletion of intracellular GSH levels by Cordia verbenacea A. DC., Costus spicatus (Jacq.) Sw., Persea americana Mill., Salix alba L., Schinus terebinthifolia Raddi and Syzygium jambolanum (Lam.) DC. accompanied because of the inhibition of GGT activity. P-gp activity was modulated in a significant manner by 17% of the extracts. The approaches used for the conduction of in vitro preclinical studies in herbal medicines revealed a series of challenges faced especially by academics in order to anticipate cases of HDI. Clinicians have also to consider the presence of intrinsic factors such as genetic polymorphisms in each patient. The possible presence of undesirable interactions between RENISUS herbal medicines and essential drugs in SUS need eventually be clinically confirmed to attest our observed in vitro effects.
... Glutathione (GSH) is an important detoxifier in liver and takes care of free oxygen radicals and peroxides [64,65]. GSH is also the way sulphur is stored in liver. ...
Article
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About 42% of Indian soils are deficient in sulphur(s) and good yield responses to S fertilization are reported in cereals, pulses and oilseeds. However, little information is available in the country on the effect of S fertilization on quality of grain, seed or oil. Soil or fertilizer S is the main source of S for the plants, which in part being the main source of S-containing essential amino acid methionine and essential vitamin thiamine for the humans. In addition, plants also synthesize a number of metabolites, such as alliin/ allicin, glucosinolates (GSLs) and methylsulfonylmethane (MSM), which have great medicinal value. Studies on the effect of S fertilization on the content of essential amino acids and vitamins and some of the discussed metabolites in grain or seed of food crops will go a long way in enhancing their commercial value and in improving human health. Effects of S fertilization on the composition of fatty acids are also suggested. Development of such analytical facilities at agricultural institutes/ universities will also help in developing varieties with higher contents of desired S compounds.
... Glutathione is the most abundant thiol-containing tripeptide in all living organisms (Meister and Andersen 1983). Glutathione is widely used in the medical, food, and cosmetic industries (Li et al. 2004;Arjinpathana and Asawanonda 2012) because of its various physiological functions, such as antioxidant, xenobiotic detoxifying, and immune boosting effects (Dröge and Breitkreutz 2000;Singh 2002;Vartanyan et al. 2000;Penninckx 2002;Ray et al. 2002;Rolseth et al. 2002). Thus, the demand for glutathione has increased in recent years. ...
Article
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Glutathione has diverse physiological functions, and therefore, the demand for it has increased recently. Currently, industrial mass production of glutathione is performed from D-glucose via fermentation by the budding yeast Saccharomyces cerevisiae. However, use of D-glucose often competes with demands for various other industries, leading to high production costs. To affordably produce glutathione, we aimed to produce high amounts of glutathione from D-glucose and D-xylose, which are the main constituents of lignocellulosic biomass pre-treated with acids. Genetically engineered S. cerevisiae strains that can produce high amounts of glutathione and assimilate D-xylose were constructed and cultured in media containing D-xylose. Among these recombinant strains, a S. cerevisiae GCI (XR/XDH/XK) strain over-expressing γ-glutamylcysteine synthetase, glutathione synthetase, D-xylose reductase, xylitol dehydrogenase, and xylulokinase genes successfully consumed D-xylose in the medium and produced the highest amount of glutathione. When strains were grown in media containing D-glucose and D-xylose, the GCI (XR/XDH/XK) strain showed 4.6-fold higher volumetric glutathione production (mg/L-broth), 2.2-fold higher glutathione content (%), and 2.1-fold higher cell growth (g-cell/L-broth) than the vector control strain of YPH499 (Vector). Furthermore, when recombinant S. cerevisiae strains were grown in medium containing fermentation inhibitory materials, the GCI (XR/XDH/XK) strain produced 5.8- and higher volumetric glutathione, 2.6-fold higher intracellular glutathione, and 2.9-fold higher cell growth than the vector control YPH499 (Vector) strain. The gradual sugar consumption by recombinant S. cerevisiae strains in medium containing D-glucose and D-xylose leads to high yields of glutathione. These results indicate the potential for glutathione production from lignocellulosic materials.
... Glutathione (GSH) is the most abundant nonprotein thiol in cells and has an array of critical functions, which include detoxifying drugs, protecting macromolecules from oxidative damage and maintaining immune functions. [1][2][3][4][5][6][7] GSH is synthesized from cysteine (Cys), glutamic acid and glycine with Cys most often being the rate-limiting substrate. 8,9 As a result, GSH levels can be depleted when Cys levels are limited such as during periods of fasting. ...
Article
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Background/objectives: Glutathione (GSH) is the most abundant endogenous antioxidant and a critical regulator of oxidative stress. Maintenance of optimal tissues for GSH levels may be an important strategy for the prevention of oxidative stress-related diseases. We investigated if oral administration of liposomal GSH is effective at enhancing GSH levels in vivo. Subjects/methods: A 1-month pilot clinical study of oral liposomal GSH administration at two doses (500 and 1000 mg of GSH per day) was conducted in healthy adults. GSH levels in whole blood, erythrocytes, plasma and peripheral blood mononuclear cells (PBMCs) were assessed in 12 subjects at the baseline and after 1, 2 and 4 weeks of GSH administration. Results: GSH levels were elevated after 1 week with maximum increases of 40% in whole blood, 25% in erythrocytes, 28% in plasma and 100% in PBMCs occurring after 2 weeks (P<0.05). GSH increases were accompanied by reductions in oxidative stress biomarkers, including decreases of 35% in plasma 8-isoprostane and 20% in oxidized:reduced GSH ratios (P<0.05). Enhancements in immune function markers were observed with liposomal GSH administration including Natural killer (NK) cell cytotoxicity, which was elevated by up to 400% by 2 weeks (P<0.05), and lymphocyte proliferation, which was elevated by up to 60% after 2 weeks (P<0.05). Overall, there were no differences observed between dose groups, but statistical power was limited due to the small sample size in this study. Conclusions: Collectively, these preliminary findings support the effectiveness of daily liposomal GSH administration at elevating stores of GSH and impacting the immune function and levels of oxidative stress.European Journal of Clinical Nutrition advance online publication, 30 August 2017; doi:10.1038/ejcn.2017.132.
... The reversible binding of glutathione to protein sulfhydryl groups protects proteins from irreversible oxidative damage (Cotgreave and Gerdes, 1998). Glutathione also performs important physiological functions such as detoxifying xenobiotics (Penninckx, 2002;Rolseth et al., 2002), and is involved in immune functioning (Dröge and Breitkreutz, 2000). Glutathione is widely used as a food additive and as a cosmetic agent (Wei et al., 2003;Yoshida et al., 2002). ...
Article
This work aims to produce glutathione directly from mannan-based bioresources using engineered Saccharomyces cerevisiae. Mannan proved to be a valuable carbon source for glutathione production by this organism. Mannan-hydrolyzing S. cerevisiae was developed by heterologous expression of mannanase/mannosidase on its cell surface. This strain efficiently produced glutathione from mannose polysaccharide, β-1,4-mannan. Furthermore, it produced glutathione from locust bean gum (LBG), a highly dense and inexpensive mannan-based bioresource, as sole carbon source. Glutathione productivity from LBG was enhanced by engineering the glutathione metabolism of mannan-hydrolyzing S. cerevisiae. Expression of extracellular mannanase/mannosidase protein combined with intracellular metabolic engineering is potentially applicable to the efficient, environmentally friendly bioproduction of targeted products from mannan-based bioresources.
... Glutathione is the most abundant thiol-containing tripeptide in all living organisms [1]. Glutathione is widely used in the medical, food, and cosmetic industries [2,3] due to its various physiological functions such as acting as an antioxidant, a detoxifier of xenobiotics, and an immune booster [4][5][6][7][8][9]. Thus the demand for glutathione has increased in recent years. ...
Article
Full-text available
Background Oxidized glutathione (GSSG) is the preferred form for industrial mass production of glutathione due to its high stability compared with reduced glutathione (GSH). In our previous study, over-expression of the mitochondrial thiol oxidase ERV1 gene was the most effective for high GSSG production in Saccharomyces cerevisiae cells among three types of different thiol oxidase genes. ResultsWe improved Erv1 enzyme activity for oxidation of GSH and revealed that S32 and N34 residues are critical for the oxidation. Five engineered Erv1 variant proteins containing S32 and/or N34 replacements exhibited 1.7- to 2.4-fold higher in vitro GSH oxidation activity than that of parental Erv1, whereas the oxidation activities of these variants for γ-glutamylcysteine were comparable. According to three-dimensional structures of Erv1 and protein stability assays, S32 and N34 residues interact with nearby residues through hydrogen bonding and greatly contribute to protein stability. These results suggest that increased flexibility by amino acid replacements around the active center decrease inhibitory effects on GSH oxidation. Over-expressions of mutant genes coding these Erv1 variants also increased GSSG and consequently total glutathione production in S. cerevisiae cells. Over-expression of the ERV1S32A gene was the most effective for GSSG production in S. cerevisiae cells among the parent and other mutant genes, and it increased GSSG production about 1.5-fold compared to that of the parental ERV1 gene. Conclusions This is the first study demonstrating the pivotal effects of S32 and N34 residues to high GSH oxidation activity of Erv1. Furthermore, in vivo validity of Erv1 variants containing these S32 and N34 replacements were also demonstrated. This study indicates potentials of Erv1 for high GSSG production.
... It has been well established that the lymphocytes of HIV/AIDS patients have low GSH content [47,48]. To address this, several human clinical trials have investigated the potential of NAC to replenish GSH. ...
Article
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Objective: To determine if orally dosed γ-glutamylcysteine (γ-GC) can increase cellular glutathione (GSH) levels above homeostasis. Many chronic and age-related disorders are associated with down-regulation, or impairment, of glutamate cysteine ligase (GCL). This suggests that γ-GC supply may become limiting for the maintenance of cellular GSH at the normal levels required to effectively protect against oxidative stress and any resulting physiological damage. Methods: GSH levels were measured in lymphocytes of healthy, non-fasting participants before and after single oral doses (2 and 4g) of γ-GC. Blood samples were immediately processed using high speed fluorescence-activated cell sorting to isolate 10(6) lymphocytes that were then assayed for GSH content. Results: A single 2g dose of γ-GC increased lymphocyte GSH content above basal levels (53±47%, p<0.01, n=14) within 90min of administration. A randomized dosage (2 and 4g γ-GC) crossover design was used to explore the pharmacokinetics of this GSH increase. In general, for both dose levels (n=9), GSH increased from initial basal levels over 3h (tmax) before reaching maximum GSH concentrations (Cmax) that were near two (2g γ-GC) to three (4g γ-GC) fold basal levels (0.4 nmol/10(6) lymphocytes). Beyond tmax, GSH levels progressively declined reaching near basal levels by 5h. The GSH half-life was between 2 and 3h with exposure (AUC) to increased GSH levels of 0.7 (2g γ-GC) and 1.8 (4g γ-GC) nmol.h/10(6) lymphocytes. Conclusions: Oral γ-GC is a non-toxic form of cysteine that can be directly taken up by cells and transiently increase lymphocyte GSH above homeostatic levels. Our findings that γ-GC can increase GSH levels in healthy subjects suggests that it may have potential as an adjunct for treating diseases associated with chronic GSH depletion. This trial was registered at anzctr.org.au as ACTRN12612000952842.
... Animals' immune system has a relatively high sensitivity to oxidative stress. Glutathione (GSH) as the most important intracellular regenerative factor has various functions such as protection against oxidative stress, regulation of gene expression, regulation of apoptosis and activation and proliferation of T lymphocytes; however, it is observed that the low level of GSH is associated with the incidence of functional disorders in lymphocytes (Droge et al., 2000;Townsend et al., 2003). Therefore, strengthening of fishes' immune system by providing dietary supplements can be a suitable solution for preventing their immune system weakness. ...
Article
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The pollutants due to effect on the immune system of fish increase fish sensitivity to pathogens. Diazinon is one of the most used organophosphates pesticide in many agricultural areas. This study aimed to evaluate the effect of diazinon on the immune system of rainbow trout (Oncorhynchus mykiss) and application of Milk thistle plant, Vitis vinifera extract to reduce the adverse effects of this pesticide on its immune system. The reduction in the level of plasma peroxides, IgM, total complement and lysozyme were observed in fish exposed to diazinon showing its effect on the fish’s immune system. No significant difference between control group and fish fed by Milk thistle plant extract and exposed to diazinon can reflect protective impact of Milk thistle plant extract on the immune system of rainbow trout by eliminating the free radicals and boosting the immune system.
... Glutathione (GSH) is an important detoxifier in liver and takes care of free oxygen radicals and peroxides [64,65]. GSH is also the way sulphur is stored in liver. ...
Article
Full-text available
Sulphur is now recognized as the fourth major plant nutrient after N, P and K globally. Sulphur in soils comes from the sulphur containing minerals present in parent materials from which the soils are derived and from the plants and animals residues or from the external addition of elemental S or its minerals. Sulphur enters the biological systems from soil through microbial activities involving mineralization of organic matter, immobilization, oxidation and reduction. Plants take up sulphur only as SO 4 = and reduce it to form S containing amino acids and other compounds. Amino acid cysteine is the source of sulphur for most other S-compounds in plants. Sulphur containing vitamin thiamine (Vitamin B1) is also synthesized only in plants and not in humans or other animals. Plants also produce vitamin biotin and a number of S-containing metabolites including glutathione, glucosinolates and alliin/allicin. Sulphur deficiency in wheat can lead to poor baking quality and in oilseeds it can lead to reduced oil content and yield. Sulphur is taken in as sulphur containing amino acids (SAAs) cysteine and methionine by human beings. The recommended dietary allowance for SAAs for humans is 14 mg kg−1 body weight. Lack of sulphur can lead to arthritis, muscle and joint stiffness, spondylitis, etc. Dietary supplements containing (chondroitin sulphate, glucosamine sulphate, methylsulfonylmethane etc.) can be beneficial in the treatment of joint diseases.
... The present study reported a deficiency in levels of reduced glutathione, which is 3.2 times lower than in healthy controls. These results go hand in hand with a previous study [17], which suggested that glutathione (GSH) is a major interacellular reducing agent, is very sensitive to oxidative pressures and has several important functions such as: protection against oxidative stress, regulation of gene expression, induction of apoptosis, activation, and proliferation in T lymphocytes. ...
... Oxidative and inflammatory processes are closely related [18]. An oxidant/antioxidant imbalance is responsible for changes in the nervous and immune system [18,68,[73][74][75][76][77][78]. Moreover, immune cells are an important source of both oxidant and proinflammatory compounds, such as reactive oxygen and nitrogen species (ROS and RNS) and inflammatory cytokines, which stimulate NF-ĸB activation, leading to production of more oxidants and inflammatory compounds, and thereby establishing a vicious circle [18,78]. ...
Article
The objective of this study is to compare oxidative stress and immune biomarkers between attention-deficit/hyperactivity disorder (ADHD) patients and controls without ADHD. A case–control comparison between 57 paediatric (6–12 years) untreated ADHD patients from the Antwerp University Hospital and 69 controls without ADHD from random schools in Flanders, Belgium, was conducted. Erythrocyte glutathione (GSH) and plasma lipid-soluble antioxidants (retinol, α-tocopherol, γ-tocopherol, retinyl palmitate, β-carotene, and co-enzyme Q10) were determined by HPLC with electrochemical detection, plasma malondialdehyde (MDA) by HPLC with fluorescence detection, plasma cytokines (interleukin (IL)-1β, IL-5, IL-6, IL-8, IL-10, tumour necrosis factor (TNF) and interferon (INF)-γ) and immunoglobulins (IgE, IgG and IgM) by flow cytometry and urinary 8-hydroxy-2′deoxyguanosine (8-OHdG) levels by ELISA assay. Dietary habits were determined by a food frequency questionnaire. Plasma MDA levels were on average 0.031 µM higher in patients than in controls (p < 0.05), and a trend for higher urinary 8-OHdG was observed. Erythrocyte GSH and plasma retinyl palmitate levels, as well as IgG and IgE levels, were higher in patients than in controls as well (on average 93.707 µg/ml, 0.006 µg/ml, 301.555 µg/ml and 125.004 µg/ml, resp., p < 0.05). Finally, a trend for lower plasma IL-5 levels was observed. After Bonferroni correction for multiple testing, the difference in GSH levels remained statistically significant (nominally significant for retinyl palmitate), while significance was lost for MDA, IgG and IgE levels. Dietary habits do not appear to cause the observed differences. These results point at the potential involvement of slight oxidative stress and immune disturbances in ADHD.
... The present study reported a deficiency in levels of reduced glutathione, which is 3.2 times lower than in healthy controls. These results go hand in hand with a previous study [17], which suggested that glutathione (GSH) is a major interacellular reducing agent, is very sensitive to oxidative pressures and has several important functions such as: protection against oxidative stress, regulation of gene expression, induction of apoptosis, activation, and proliferation in T lymphocytes. ...
Article
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The present work is concerned with the study of the effects of antioxidant vitamins on antioxidant status and liver function in homozygous β-thalassemic patients. The patients were treated with vitamins E, C and A for twelve months. With respect to antioxidant vitamins, before treatment there were deficiencies in these vitamins in thalassemic patients as compared with healthy controls. Also, before treatment there was a significant elevation in the malondialdehyde (MDA) concentration, and a significant deficiency in levels of reduced glutathione (GSH). After treatment, patients with β-thalassemia major exhibited significant improvements in the levels of non-enzymatic parameters as compared with the levels of these parameters before treatment. Also, MDA significantly decreased and reduced glutathione was highly increased after treatment. Also, values of Hb and ferritin showed improvements of their values after treatment. The results of enzymes showed that thalassemic major children suffer from high levels of ALT, AST, glutathione peroxidase, and superoxide dismutase enzymes activities before vitamins treatment. The activities of ALT, AST, GPx, and SOD decreased significantly, also the activities of catalase and glutathione reductase significantly increased in β-thalassemic patients after treatment compared with their activities before treatment. Finally, it seems clear that treatment of β-thalassemic patients with antioxidant vitamins serves to improve the healthy status of the patients through an enhancement of the levels of antioxidants and reduction of the oxidative damage, and hence improve the hemoglobin levels and liver function.
... For instance, NAC ingestion is proposed to result in an anti-oxidant effect that minimizes the oxidative stress and inflammatory response imposed from physical activity [151] (see Sect. 4.2). Furthermore, NAC is proposed to enhance fatigue resistance [152] and improve athletic performance [151]; to enhance immune system function [153], hemodynamics and muscle blood flow [154]; and to modulate EPO production and the hypoxic ventilatory response [155]. Intuitively, each of these mechanisms appears likely to support a positive adaptation to hypoxic environments such as altitude exposure. ...
Article
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Training at low to moderate altitudes (~ 1600-2400 m) is a common approach used by endurance athletes to provide a distinctive environmental stressor to augment training stimulus in the anticipation of increasing subsequent altitude- and sea-level-based performance. Despite some scientific progress being made on the impact of various nutrition-related changes in physiology and associated interventions at mountaineering altitudes (> 3000 m), the impact of nutrition and/or supplements on further optimization of these hypoxic adaptations at low-moderate altitudes is only an emerging topic. Within this narrative review we have highlighted six major themes involving nutrition: altered energy availability, iron, carbohydrate, hydration, antioxidant requirements and various performance supplements. Of these issues, emerging data suggest that particular attention be given to the potential risk for poor energy availability and increased iron requirements at the altitudes typical of elite athlete training (~ 1600-2400 m) to interfere with optimal adaptations. Furthermore, the safest way to address the possible increase in oxidative stress associated with altitude exposure is via the consumption of antioxidant-rich foods rather than high-dose antioxidant supplements. Meanwhile, many other important questions regarding nutrition and altitude training remain to be answered. At the elite level of sport where the differences between winning and losing are incredibly small, the strategic use of nutritional interventions to enhance the adaptations to altitude training provides an important consideration in the search for optimal performance.
... For instance, its ability to suppress melanin synthesis through tyrosinase inhibition allows it to be used as a skin whitening agent in cosmeceuticals [5][6][7]. In addition, GSH may be used as an immune booster, an antidote for metal poisoning, and for the treatment of diseases such as fibrosis, glaucoma, and arthritis [8][9][10][11]. Unfortunately, its poor bioavailability and unpleasant odor limit the use of GSH in clinics despite its many therapeutic applications. ...
Article
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Glutathione is a natural anti-aging substance that prevents the oxidation of protein thiols from reactive oxygen species. In the pharmaceutical industry, reduced glutathione (GSH) has been widely used for skin whitening due to its ability to inhibit tyrosinase. However, its poor permeability and foul odor limit its use in skin applications. Herein, we report a GSH-loaded dissolving microneedle (MN) patch prepared with hyaluronic acid (HA) that enables enhanced permeation across the skin and reduces the foul odor of GSH. HA was selected to prepare odorless GSH solutions and used for MN fabrications as a carrier of GSH. GSH-loaded MN (GSH-MN) arrays prepared from MN-forming solution containing up to 10% GSH showed good pattern uniformity and appropriate mechanical properties for insertion into the skin. The GSH-MNs with a loading capacity of 17.4% dissolve within 10 min following insertion into porcine skin and release the loaded GSH without being oxidized. This new approach combines functional biopolymers to reduce the characteristic GSH odor and advanced transdermal delivery based on MN technology to enhance skin permeation without pain. We believe this technique could expand the application of GSH in many cosmeceutical fields.
... Механизм биохимического действия тиолов заключается в способности восстанавливать дисульфидные связи при патологических нарушениях, инактивировать токсические агенты, а также повышать содержание сульфгидрильных групп, обеспечивая антиоксидантный эффект (16,17). Тиольные соединения в первую очередь подвергаются воздействию активных кислородных радикалов, что предохраняет от их влияния функциональные группы биологических молекул и клеточных мембран (18,19). ...
... Механизм биохимического действия тиолов заключается в способности восстанавливать дисульфидные связи при патологических нарушениях, инактивировать токсические агенты, а также повышать содержание сульфгидрильных групп, обеспечивая антиоксидантный эффект (16,17). Тиольные соединения в первую очередь подвергаются воздействию активных кислородных радикалов, что предохраняет от их влияния функциональные группы биологических молекул и клеточных мембран (18,19). ...
... In particular, some lymphocyte functions, such as DNA synthesis, are favored by high levels of GSH, while other redox-sensitive pathways are favored by low intracellular GSH. However, immunological functions in the diseases characterized by oxidative stress can be restored by cysteine or GSH supplementation [27,28]. The brain generates high levels of ROS due to its high oxygen consumption, hence it is more susceptible to the damaging effects of ROS than other tissues [29,30]. ...
Article
Glutathione (GSH) has poor pharmacokinetic properties; thus, several derivatives and biosynthetic precursors have been proposed as GSH-boosting drugs. I-152 is a conjugate of N-acetyl-cysteine (NAC) and S-acetyl-β-mercaptoethylamine (SMEA) designed to release the parent drugs (i.e., NAC and β-mercaptoethylamine or cysteamine, MEA). NAC is a precursor of L-cysteine, while MEA is an aminothiol able to increase GSH content; thus, I-152 represents the very first attempt to combine two pro-GSH molecules. In this review, the in-vitro and in-vivo metabolism, pro-GSH activity and antiviral and immunomodulatory properties of I-152 are discussed. Under physiological GSH conditions, low I-152 doses increase cellular GSH content; by contrast, high doses cause GSH depletion but yield a high content of NAC, MEA and I-152, which can be used to resynthesize GSH. Preliminary in-vivo studies suggest that the molecule reaches mouse organs, including the brain, where its metabolites, NAC and MEA, are detected. In cell cultures, I-152 replenishes experimentally depleted GSH levels. Moreover, administration of I-152 to C57BL/6 mice infected with the retroviral complex LP-BM5 is effective in contrasting virus-induced GSH depletion, exerting at the same time antiviral and immunomodulatory functions. I-152 acts as a pro-GSH agent; however, GSH derivatives and NAC cannot completely replicate its effects. The co-delivery of different thiol species may lead to unpredictable outcomes, which warrant further investigation.
... Dietary supplements containing S (chondroitin sulfate, glucosamine sulfate, methylsulfonylmethane) can be benefi cial in the treatment of joint diseases (Nimini et al., 2006). Glutathione (GSH) takes care of free oxygen radicals and peroxides (Dorge and Bretkreutz, 2000;Townsend et al., 2004). GSH is also the form S is stored in the liver. ...
... Glutathione (GSH) is required by the immune system for two important reasons: it protects host immune cells through its antioxidant mechanism and it provides the optimal functioning of lymphocytes and other cells of the immune system [1]. Conditions under which GSH supplementation may be recommended occur in the elderly who have low levels of GSH and in individuals with chronic inflammation [2]. ...
Article
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Glutathione and aged garlic extract are sulfur-containing products that play important protective and regulatory roles within the immune system and in oxidative processes. Hydrogen sulfide (H2S), an endogenous, gaseous, signaling transmitter, has also been shown to be involved in the regulation of inflammation. Recent studies have shown that sulfur-containing compounds from garlic have beneficial effects in attenuating outcomes associated with cardiovascular disease and inflammation by a mechanism that may be related to the H2S signaling pathway. In this review, we summarize the main functions of glutathione (GSH), garlic derivatives and H2S and their role in the immune response and impact on health and disease.
... Decreased level of CoQ10 is linked with the development of the cardiovascular disease. Glutathione (GSH) is an intracellular antioxidant essential for the immune cells functioning [201]. Low level of GSH is correlated with chronic heart failure and myocardial infarction [189,202,203]. ...
Article
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Inflammation is one of the common events in the majority of acute as well as chronic debilitating diseases and represent a chief cause of morbidity in today’s era of modern lifestyle. If unchecked, inflammation leads to development of rheumatoid arthritis, diabetes, cancer, Alzheimer’s disease, and atherosclerosis along with pulmonary, autoimmune and cardiovascular diseases. Inflammation involves a complex network of many mediators, a variety of cells, and execution of multiple pathways. Current therapy for inflammatory diseases is limited to the steroidal and non-steroidal anti-inflammatory agents. The chronic use of these drugs is reported to cause severe adverse effects like gastrointestinal, cardiovascular, and renal abnormalities. There is a massive need to explore new anti-inflammatory agents with selective action and lesser toxicity. Plants and isolated phytoconstituents are promising and interesting sources of new anti-inflammatories. However, drug development from natural sources has been linked with hurdles like the complex nature of extracts, difficulties in isolation of pure phytoconstituents, and the yield of isolated compounds in minute quantities that is insufficient for subsequent lead development. Although various in-vivo and in-vitro models for anti-inflammatory drug development are available, judicious selection of appropriate animal models is a vital step in the early phase of drug development. Systematic evaluation of phytoconstituents can facilitate the identification and development of potential anti-inflammatory leads from natural sources. The present review describes various techniques of anti-inflammatory drug screening with its advantages and limitations, elaboration on biological targets of phytoconstituents in inflammation and biomarkers for the prediction of adverse effects of anti-inflammatory drugs. The systematic approach proposed through present article for anti-inflammatory drug screening can rationalize the identification of novel phytoconstituents at the initial stage of drug screening programs.
... Thus, it is understandable to think that the higher the GSH concentration and the higher Glutathione Peroxidase activity in immune cells together with the lower one of MDA, the better the redox balance of these cells, and therefore their function. In fact, GSH has been shown to be essential for proper immune cell functioning given that even a moderate depletion of GSH has been shown to impair several leukocyte functions (Dröge and Breitkreutz, 2000). MDA, which is an end-product of reactive oxygen species (ROS)-induced peroxidation and therefore is used as a marker of oxidative stress, has also been shown to play an active role by inducing the cross-links in proteins and forming irreversible advanced glycated end-products (AGEs) (Esterbauer et al., 1990). ...
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The identification of predictive markers of life span would help to unravel the underlying mechanisms influencing ageing and longevity. For this aim, 30 variables including immune functions, inflammatory-oxidative stress state and behavioural characteristics were investigated in ICR-CD1 female mice at the adult age (N = 38). Mice were monitored individually until they died and individual life spans were registered. Multiple linear regression was carried out to construct an Immunity model (adjusted R2 = 75.8%) comprising Macrophage chemotaxis and phagocytosis and Lymphoproliferation capacity, a Redox model (adjusted R2 = 84.4%) involving Reduced Glutathione and Malondialdehyde concentrations and Glutathione Peroxidase activity and a Behavioural model (adjusted R2 = 79.8%) comprising Internal Locomotion and Time spent in open arms indices. In addition, a Combined model (adjusted R2 = 92.4%) and an Immunity-Redox model (adjusted R2 = 88.7%) were also constructed by combining the above-mentioned selected variables. The models were also cross-validated using two different sets of female mice (N = 30; N = 40). Correlation between predicted and observed life span was 0.849 (P < 0.000) for the Immunity model, 0.691 (P < 0.000) for the Redox, 0.662 (P < 0.000) for the Behavioural and 0.840 (P < 0.000) for the Immunity-Redox model. Thus, these results provide a new perspective on the use of immune function, redox and behavioural markers as prognostic tools in ageing research.
... [1][2][3] The imbalance of GSH homeostasis is concerned with various clinical diseases, such as liver or lung damage, cancer, AIDS, Alzheimer's and Parkinson's disease. [4][5][6][7] Therefore, analysis of GSH in biological samples is of continuous interest and great significance in biomedical applications. ...
Article
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A bioinspired fluorometric method has been developed for the detection of glutathione (GSH) in biological fluids. It is based on the use of near-infrared fluorescent semiconducting polymer dots (P-dots) and of the dopamine (DA)-melanin nanosystem. The P-dots were prepared from poly(styrene-co-maleic anhydride), the semiconducting polymer poly[(9,9′-dioctyl-2,7-divinylenefluorenylene)-alt-2-methoxy-5-(2-ethyl-hexyloxy)-1,4-phenylene] and the fluorescent dye tetraphenylporphyrin. They have excitation/emission maxima at 458/656 nm, and this enables measurement to be performed with low autofluorescence and scattering background. DA can self-polymerize on the surface of the P-dots to yield a poly-DA coating. This coating, at weak alkaline pH values, causes the quenching of the fluorescence of the P-dots. However, the polymerization of DA is inhibited by GSH. Hence, quenching of fluorescence is prevented. This effect was used to design a fluorometric assay for GSH that has good selectivity and sensitivity. Under optimal conditions, the method has a linear response in the 0.2 to 20 μM GSH concentration range and a 60 nM detection limit. It was successfully applied to the determination of GSH in HepG2 cells and in spiked human serum. Graphical abstractSchematic representation of using a NIR fluorescent P-dots and dopamine (DA)-melanin nanohybrid as a probe for glutathione (GSH) detection. The P-dots were prepared from poly(styrene-co-maleic anhydride) (PSMA), the semiconducting polymer poly[(9,9′-dioctyl-2,7-divinylenefluorenylene)-alt-2-methoxy-5-(2-ethyl-hexyloxy)-1,4-phenylene] (PEPV) and the fluorescent dye tetraphenylporphyrin (TPP).The GSH can inhibit the dopamine self-polymerization and prevented the formation of PDA and fluorescence quenching of P-dots.
... The glutathione cycle is one of the most important intracellular mechanisms that play a key role in the preservation of an adequate intracellular redox state [59]. In the present study, AD patients showed lower GR activities and GSH concentrations, together with higher GSSG concentrations and GSSG/GSH ratios. ...
Article
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Oxidative stress plays an essential and early role in the pathophysiology of Alzheimer's disease (AD). Alterations in the redox state in AD and in mild cognitive impairment (MCI) patients appear in the brain and at peripheral level. Given that it is easier to study the latter, most of the research has been focused on plasma. However, the analysis of redox parameters in whole blood cells (including erythrocytes and leukocytes) has not really been investigated. Moreover, the association of these parameters with Mini-Mental State Examination (MMSE) clinical scores, has scarcely been studied. Therefore, the aim of the present work was to analyze several redox markers in whole blood cells from male and female MCI and AD patients. Antioxidant (superoxide dismutase, catalase (CAT), glutathione peroxidase (GPx), and reductase (GR) activities, and reduced glutathione (GSH) concentration) together with oxidant parameters (oxidized glutathione (GSSG) and thiobarbituric acid-reactive substances (TBARS)) were investigated using MCI and AD (10 women and 10 men in each group) and their age-matched control groups (15 women and 15 men). The results show an altered redox state in whole blood cells from AD patients (higher CAT, GSSG/GSH, TBARS and lower GPx, GR, GSH). Some of these redox parameters are already affected in MCI patients (higher TBARS and lower GPx and GR activities) in both sexes and, consequently, they could be used as markers of prodromal AD. Since GR, GSH, GSSG, and GSSG/GSH were found to be associated with MMSE scores, they seem to be useful clinically to monitor cognitive decline in AD progression.
... PON1 was depressed in GWV generally (relative to military controls) -and particularly so in those with GWI or more symptoms 60, 61, 151 . Natural killer activity is reduced with OS and increased with antioxidants 135,239,240 . Blunted heart rate variability has been linked to OSMD 242,243 . ...
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Background: Overlapping chronic multisymptom illnesses (CMI) include Chronic Fatigue Syndrome (CFS), fibromyalgia, irritable bowel syndrome, multiple chemical sensitivity, and Gulf War illness (GWI), and subsets of autism spectrum disorder (ASD). GWI entails a more circumscribed set of experiences that may provide insights of relevance to overlapping conditions.Objectives: To consolidate evidence regarding a role for oxidative stress and mitochondrial dysfunction (OSMD), as primary mediators in CMI, using GWI as a departure point.Methods: Exposure relations, character, timecourse and multiplicity of symptoms, and objective correlates of GWI are compared to expectation for OSMD. Objective correlates of OSMD in GWI and overlapping conditions are examined.Discussion: OSMD is an expected consequence of known GWI exposures; is compatible with symptom characteristics observed; and accords with objective markers and health conditions linked to GWI, extending to autoimmune disease and infection. Emergent triangulating evidence directly supports OSMD in multisymptom “overlap” CMI conditions, with similarities to, and diagnosed at elevated rates in, GWI, suggesting a common role in each.Conclusions: GWI is compatible with a paradigm by which uncompensated exposure to oxidative/nitrative stressors accompanies and triggers mitochondrial dysfunction, cell energy compromise, and multiple downstream effects such as vulnerability to autoantibodies. This promotes a profile of protean symptoms with variable latency emphasizing but not confined to energy-demanding post-mitotic tissues, according with (and accounting for) known properties of multisystem overlap conditions. This advances understanding of GWI; health conditions attending GWI at elevated rates; and overlap conditions like CFS and ASD, providing prospects for vulnerability assessment, mitigation of progression, treatment, and future prevention – with implications germane to additive and excessive environmental oxidative stressor exposures in the civilian setting.
... 24 GSH is also crucial for the normal functioning of the immune system 25 and its levels affect many of the intricate steps involved in the development of an immune response, including activation and proliferation of lymphocytes. 26 GSH is synthesised within the cell from its three constituent amino acids, glycine, glutamate and cysteine, the thiol (-SH) group on the cysteinyl residue giving the tripeptide its biological activity. 27 The availability of cysteine limits the ability of the cell to generate GSH. ...
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Objective The prevalence and costs associated with treating pressure ulcers (PU) are at high levels. Frequently, PUs heal slowly or not at all, which may be due to the patient's catabolic state which may include protein energy malnutrition. The objective of this open label clinical trial was to improve healing rates by providing patients with a patented, high-quality protein containing all essential amino acids to ensure positive nitrogen balance. An additional benefit of this protein is the delivery of bioavailable cysteine (cystine) to promote glutathione (GSH) synthesis which supports immune function and heightens antioxidant defences. Methods Patients with category II, III and IV PUs were fed 20g BID whey protein dietary supplement for 16–120 days, without change in ongoing ‘best practice’ PU management and their progress recorded. Results A total of 10 patients were recruited, with an average age of 77 years. Most had shown no improvement in healing for ≥2 months before treatment and usually had other complications including chronic obstructive pulmonary disease (COPD), diabetes and various cardiovascular diseases. There were a total of 23 PUs, with some patients having more than one. Of these, 44% (n=10) showed complete resolution 83% (n=19) had better than 75% resolution over the observation period. Healing rates ranged from 16.9–0.2cm ² /month (healed PUs) and 60.0–1.6cm ² /month for resolving PUs. Conclusion By providing the necessary amino acids to rebuild tissues and bioactive cysteine (cystine) to promote synthesis of intracellular GSH and positive nitrogen balance, improvement in PUs healing was achieved.
... [5] The enhanced by oxidative conditions and favored by low intracellular glutathione levels. [26] GENERAL MULTIPLE FUNCTIONS OF GSH [5,17] Reducing agent, antioxidant ...
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In this review article would selected and talk in some sides related to Glutathione and highlights the some points as general. Glutathione (γ-glutamyl-cysteinylglycine) is an endogenous peptide with antioxidant plays many useful functions in human body and therefore determination of this small molecule is very important for present-day medicine and pharmacy. General multiple functions of GSH A.
... GSH is a key contributor to a widespread biotic action, such as immune regulation, detoxification. GSH has been used in pharmaceuticals, cosmetics, and food industries [2], [3]. ...
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Objective: Glutathione is an antioxidant, which presents in mammalian and known as the most powerful antioxidant. Glutathione is called as "Master Antioxidant" because of its intracellular and possesses the aptitude to exploit the performance of other antioxidants, these include vitamins C & E, CoQ10 (ubiquinone) and alpha-lipoic acid, and rich in fresh vegetables and fruits. Conclusion: The major role of Glutathione is to protecting cells and mitochondria from oxidative and peroxidative damage also needed for cleansing, energy utilization, reduction of aging-associated diseases, elimination of toxins from the cells, and protection from the damaging effects of radiation, chemicals, and environmental pollutants. Our body's ability to produce glutathione decreases as you age. This review suggests that Glutathione can recommend as the best therapeutic agent in pharma industries and skin-lightening agent in cosmetic industries.
... A growing number of families with children diagnosed with developmental disabilities, particularly ASD, are implementing CAM supplements to treat symptoms that they believe are affected by oxidative stress pathways [21,22]. The reported use of CAM in this population ranges from 32 to 87% in the United States [23,24]. ...
Article
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Oxidative stress is understood to be involved in the ontology and maintenance of different developmental disabilities. Some complementary and alternative medicine (CAM) therapies have been proposed to modify this relationship by affecting oxidative stress pathways. However, it is unclear which of these CAM therapies are used among children with different developmental disabilities. This study examines the use of these therapies among 10,218 children between the ages of 4 and 17 using the 2012 Child Complementary and Alternative Medicine (CAM) Supplement of the National Health Interview Survey (NHIS) to highlight a potential avenue for intervention and prevention efforts. The results suggest that children with developmental disabilities are more likely to utilize particular CAM therapies that may alter oxidative stress pathways. Future work is needed to assess the potential moderating effect of these CAM therapies and oxidative stress levels among children with different developmental disabilities.
... Most of the polyphenols act through the Nrf2 pathway [37; 38], which in turn activates a series of downstream molecules such as glutathione and glutathione-S-transferase. The reactions catalyzed by the GSH-dependent enzymes (that have been described in detail earlier paragraphs) set the balance of cellular redox system. In addition to these, GSH activation plays important roles in immune system modulation [39]. GSH is an essential factor for inducible nitric oxide synthase (iNOS) activity [40], where a depletion in the GSH level causes a concomitant reduction in the iNOS activity. ...
Article
Glutathione (GSH or reduced glutathione) is a tripeptide of gamma-Glutamyl-cysteinyl-glycine and the predominant intracellular antioxidant in many organisms including humans. GSH and associated enzymes are controlled by a transcription factor-nuclear factor-2 related erythroid factor-2 (Nrf2). In cellular milieu, GSH protects the cells essentially against a wide variety of free radicals including reactive oxygen species, lipid hydroperoxides, xenobiotic toxicants, and heavy metals. It has two forms, the reduced form or reduced glutathione (GSH) and oxidized form (GSSG), where two GSH moieties combine by sulfhydryl bonds. Glutathione peroxidase (GPx) and glutathione-s-transferase (GST) essentially perform the detoxification reactions using GSH, converting it into GSSG. Glutathione reductase (GR) operates the salvage pathway by converting GSSG to GSH with the expense of NADPH and restores the cellular GSH pool. Hence, GSH and GSH-dependent enzymes are necessary for maintaining the normal redox balance in the body and help in cell survival under stress conditions. In addition, GST removes various carcinogenic compounds offering a chemopreventive property, whereas the GSH system plays a significant role in regulating the cellular survival by offering redox stability in a variety of cancers including prostate, lung, breast, and colon cancer. Studies have also indicated that GSH inhibitors, such as buthionine sulfoximine, improves the chemo-sensitivity in cancer cells. In addition, GSH and dependent enzymes provide survival advantage for cancer cells against chemotherapeutic drugs and radiotherapy.
... GSH protects cells against exogenous and endogenous harmful molecules including reactive oxygen and nitrogen species (ROS/RNS), limiting the damaging effects of oxidative/nitrosative stress (2,3). Beside its function as intracellular redox buffer, GSH exerts a key role in the immune system, in antiviral and inflammatory response (4)(5)(6)(7). Concerning the inflammatory response, it has been demonstrated that, intracellular GSH depletion represents the first event of the signaling process (8)(9)(10). This alteration is accompanied by an increased production of cytokine such as tumor necrosis factor (TNF-α), IL-1β, IL-6, and IL-8 (11,12). ...
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An imbalance in GSH/GSSG ratio represents a triggering event in pro-inflammatory cytokine production and inflammatory response. However, the molecular mechanism(s) through which GSH regulates macrophage and cell autonomous inflammation remains not deeply understood. Here, we investigated the effects of a derivative of GSH, the N-butanoyl glutathione (GSH-C4), a cell permeable compound, on lipopolisaccharide (LPS)-stimulated murine RAW 264.7 macrophages, and human macrophages. LPS alone induces a significant production of pro-inflammatory cytokines, such as IL-1β, IL-6, and TNF-α and a significant decrement of GSH content. Such events were significantly abrogated by treatment with GSH-C4. Moreover, GSH-C4 was highly efficient in buffering cell autonomous inflammatory status of aged C2C12 myotubes and 3T3-L1 adipocytes by suppressing the production of pro-inflammatory cytokines. We found that inflammation was paralleled by a strong induction of the phosphorylated form of NFκB, which translocates into the nucleus; a process that was also efficiently inhibited by the treatment with GSH-C4. Overall, the evidence suggests that GSH decrement is required for efficient activation of an inflammatory condition and, at the same time, GSH-C4 can be envisaged as a good candidate to abrogate such process, expanding the anti-inflammatory role of this molecule in chronic inflammatory states.
Chapter
The principal role of adaptive immunity is to distinguish between “self” and “nonself” and thus provide a highly specific line of immunological defence for the efficient removal of foreign material. It comprises of two arms: the effector B-cell arm and effector T-cell arm which act together to remove nonself. The balance between oxidising and reducing agents within these immune cells governs their redox state. This is important as transient controlled changes in the redox state, such as increased production of reactive oxygen species, are vital for signalling and induction of various biological processes, including cell growth and apoptosis. However, in chronic inflammatory diseases, the prolonged and persistent production of ROS, which overwhelms cellular antioxidant systems leading to oxidative stress, may influence T-cell function. This contributes to a T-cell phenotype which is hyporesponsive to growth and death signals and persists at the site of inflammation, perpetuating the immune response. The regulation of T-cell function by oxidative stress therefore has implications for rheumatoid arthritis.
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Feeding ruminants a high-grain (HG) diet is a widely used strategy to improve milk yield and cost efficiency. However, it may cause certain metabolic disorders. At present, information about the effects of HG diets on the systemic metabolic profile of goats and the correlation of such diets with rumen bacteria is limited. In the present study, goats were randomly divided into two groups: one was fed the hay diet (hay; n = 5), while the other was fed HG diets (HG; n = 5). On day 50, samples of rumen contents, peripheral blood serum and liver tissues were collected to determine the metabolic profiles in the rumen fluid, liver and serum and the microbial composition in rumen. The results revealed that HG diets reduced ( P < 0.05) the community richness and diversity of rumen microbiota, with an increase in the Chao 1 and Shannon index and a decrease in the Simpson index. HG diets also altered the composition of rumen microbiota, with 30 genera affected ( P < 0.05). Data on the metabolome showed that the metabolites in the rumen fluid, liver and serum were affected (variable importance projection > 1, P <0.05) by dietary treatment, with 47, 10 and 27 metabolites identified as differentially metabolites. Pathway analysis showed that the common metabolites in the shared key pathway (aminoacyl-transfer RNA biosynthesis) in the rumen fluid, liver and serum were glycine, lysine and valine. These findings suggested that HG diets changed the composition of the rumen microbiota and metabolites in the rumen fluid, liver and serum, mainly involved in amino acid metabolism. Our findings provide new insights into the understanding of diet-related systemic metabolism and the effects of HG diets on the overall health of goats.
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In this study, the CPsittF(R) and Or1(2) primers, targeting the ompA gene of Chlamydia psittaci, were used for PCR detection of Chlamydia abortus DNA in clinical samples from aborted animals. The sensitivity and specificity of the reaction were compared with those achieved by C. abortus-specific primers CpsiA(B). The comparative analysis of the results showed that all samples that were positive for C. abortus in PCR, performed with primers CpsiA(B) also gave a positive result in the PCR using the CPsittF(R) primers, generating a 1041 bp specific amplification product. PCR amplification using the Or1(2) primers was uncertain and produced an amplification product of 212 bp, which was different from the expected length of 245 bp. In an attempt to improve these primers, their sequence was modified at the 11th and the 21st nucleotide. Although the sensitivity of the reaction performed with the modified primers was improved, it was still lower compared to that achieved with the original C. abortus-specific primers CpsiA(B) and primers CPsittF(R). The results show that the primer pair CPsittF(R), developed for the detection of C. psittaci could be successfully used in the diagnosis of abortions, induced by C. abortus, while the primer set Or1(2) is less effective.
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During influenza pandemics in the last few years many people have died from severe complications associated with this pandemic despite receiving intensive care. This suggests that a definitive medical treatment for severe influenza-associated complications has not yet been established. About the pathogenicity of flu, many studies have shown that superoxide anion produced by macrophages infiltrated into the virus-infected organs is implicated in the development of severe influenza-associated complications. Selected antioxidants, such as alpha-tocopherol, N-acetyl-L-cysteine, glutathione, ascorbic acid, 5,7,4-trihydroxy-8-methoxyflavone, catechins, quercetin 3-rhamnoside, iso-quercetin etc. inhibit the proliferation of influenza virus and scavenge superoxide anions. The combination of antioxidants with antiviral drugs synergistically reduces the lethal effects of influenza virus infections. These results suggest that the combination of an agent with antiviral activities and an antioxidant could be a drug of choice for the treatment of patients with severe influenza-associated complications. Hopefully, this update of knowledge of antioxidant therapy of flu could be used as a potential approach to overcome the influenza-associated complications. Key words: influenza virus infection, oxidative stress, antioxidants, specific inhibitors, combination therapy
Chapter
Small-molecules containing sulfur can ameliorate oxidative damage, acting as effective antioxidants, and playing a crucial role on health. In this chapter, we discuss the antioxidant activities of organosulfur compounds, highlighting the different role of the three main families of natural compounds containing sulfur: glucosinolates, sulfur amino acids and disulfides. Glucosinolates are present in vegetables included in the cruciferous family such as broccoli, Brussels sprouts and cauliflower. Consumption of these vegetables has been associated to lower incidences of many diseases including cancer and cardiovascular disorders. Glucosinolates are transformed into isothiocyanates such as sulforaphane or phenethyl isothiocyanate which inhibit carcinogenesis through unique mechanisms of cancer inhibition. Sulfur amino acids include cysteine and cysteine derivatives such as glutathione, a tripeptide with an important antioxidant role in preventing damage to cellular components caused by reactive oxygen species. NAcetylcysteine is extensively used as a mucolytic agent, and it is a precursor of taurine. Taurine exhibits numerous beneficial actions such as improvement of cardiovascular health and lowering of blood pressure. Vegetables of the genus Allium such as garlic and onions have shown potential chemopreventive properties, which have been attributed to the presence of high levels of organosulfur compounds such as diallyl disulfide. This disulfide has shown anti-proliferative effects against different types of cancer cells. Lipoic acid, a naturally occurring cyclic disulfide derived from octanoic acid, has been related to the prevention of age-related cognitive dysfunction and progression of Alzheimer's disease.
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Glutathione (GSH) deficiency may play a pivotal role in a variety of apparently unrelated clinical conditions and diseases. Orally administered N-acetylcysteine (NAC), which replenishes the cysteine required for GSH synthesis, has been tested in a large number of randomized placebo-controlled trials involving these diseases and conditions. This chapter focused on developing a base of evidence suggesting that NAC administration improves disease by increasing cysteine and/or GSH in a variety of diseases, thereby implying a significant role for GSH deficiency in the clinical basis of many diseases. To develop this base of evidence, we systematically selected studies which considered the hypothesis that the therapeutic efficacy for NAC is an indication that cysteine and/or GSH deficiency is a pathophysiological part of the diseases studied. In this manner we focus this chapter on explaining the biological mechanisms of NAC therapy in a wide variety of disorders and demonstrate its ubiquitous role in improving disease that involves disrupted GSH and/or cysteine metabolism.
Literature Review
Article
In studies on fatty livers due to amino acid imbalances, it was found that addition of 0.3% of methionine and threonine to a protein free diet significantly reduced the loss of body weight and urinary nitrogen excretion of young adult female rats. Addition of methionine alone or of a combination of methionine and an essential amino acid other than threonine had a little effect in reducing urinary nitrogen excretion. Supplemented amount and the ratio of methionine and threonine was also investigated and the nitrogen sparing action of methionine and threonine was observed in fairly wide range of the supplemented amount and the ratio of the two amino acids. Effect of supplementation of methionine and threonine on some amino acid catabolizing enzymes in the livers of rats was also investigated.
Article
Purified CD4+ and CD8+ T cells, prepared from seven aged rhesus monkeys (>20 years), were 40-97% suppressed in their mitogenic responses to Con A or anti-CD3 (with PMA) when compared to T cell responses from four young control animals. The suppressed mitogenic responses were paralleled by diminished synthesis of interleukin-2 (IL-2). The CD4+ and CD8+ T cells from a given aged animal were suppressed approximately the same degree. Addition of NAC (5 mM) enhanced the aged T cell proliferative response and IL-2 production from 4-15 fold; the more strongly suppressed cells generally were more responsive to NAC Control T cell responses were modestly enhanced from 1.5-3 fold. Since the cysteine of NAC readily incorporates into glutathione, the observation that NAC restores aged T cell responses suggests that glutathioneassociated metabolism, an important component for T cell mitogenesis, may be altered in aged rhesus T cells. Supported by NIH grants 1-S06RR08239 and 1G12RR03050.
Article
The nitrogen sparing action of methionine and arginine supplementations to a protein-free diet were studied with colostomized adult cocks.Body weight loss of the cocks fed a protein-free diet was alleviated with methionine or arginine supplementation, but neither effect was significant. Faecal nitrogen showed comparable values and urinary nitrogen of the cocks fed the methionine supplemented diet was the lowest of the 3 groups. The nitrogen balance were, in the order of increasing, methionine supplemented, arginine supplemented and protein-free groups.Each of the major urinary nitrogenous compounds of cocks fed the methionine supplemented diet showed lower values than the corresponding values for the cocks fed a protein-free diet. From the result of faecal nitrogen and faecal amino acid analysis, there was no evidence that the re-utilization of endogenous nitrogen of the birds fed a protein-free diet was enhanced by the addition of methionine or arginine.
Article
To establish whether the low cysteine and glutathione levels in HIV-infected patients and SIV-infected rhesus macaques may be consequences of an abnormal cysteine catabolism, we analyzed sulfate and glutathione levels in macaques. Muscle tissue (m. vastus lateralis and m. gastrocnemius) of SIV-infected macaques (n = 25) had higher sulfate and lower glutathione and glutamate levels than that of uninfected controls (n = 9). Hepatic tissue, in contrast, showed decreased sulfate and glutathione disulfide (GSSG) levels, and increased γ-glutamylcysteine synthetase (γ-GCS) activity. These findings suggest drainage of the cysteine pool by increased cysteine catabolism in skeletal muscle tissue, and by increased hepatic glutathione biosynthesis. Cachectic macaques also showed increased urea levels and decreased glutamine/urea ratios in the liver, which are obviously related to the abnormal urea excretion and negative nitrogen balance commonly observed in cachexia. As urea production and net glutamine synthesis in the liver are strongly influenced by proton-generating processes, the abnormal hepatic urea production may be the direct consequence of the cysteine deficiency and the decreased catabolic conversion of cysteine into sulfate and protons in the liver. To establish whether the low cysteine and glutathione levels in HIV-infected patients and SIV-infected rhesus macaques may be consequences of an abnormal cysteine catabolism, we analyzed sulfate and glutathione levels in macaques. Muscle tissue (m. vastus lateralis and m. gastrocnemius) of SIV-infected macaques (n = 25) had higher sulfate and lower glutathione and glutamate levels than that of uninfected controls (n = 9). Hepatic tissue, in contrast, showed decreased sulfate and glutathione disulfide (GSSG) levels, and increased γ-glutamylcysteine synthetase (γ-GCS) activity. These findings suggest drainage of the cysteine pool by increased cysteine catabolism in skeletal muscle tissue, and by increased hepatic glutathione biosynthesis. Cachectic macaques also showed increased urea levels and decreased glutamine/urea ratios in the liver, which are obviously related to the abnormal urea excretion and negative nitrogen balance commonly observed in cachexia. As urea production and net glutamine synthesis in the liver are strongly influenced by proton-generating processes, the abnormal hepatic urea production may be the direct consequence of the cysteine deficiency and the decreased catabolic conversion of cysteine into sulfate and protons in the liver.
Article
HIV-1 proviral DNA contains two binding sites for the transcription factor NF-x B. HIV-1-infected individuals have, on average, abnormally high levels of tumour necrosis factor [alpha] (TNF[alpha]) and abnormally low plasma cysteine levels. We therefore investigated the effects of cysteine and related thiols on HIV-1 replication and NF-x B expression. The experiments in this report show that cysteine or N-acetylcysteine (NAC) raise the intracellular glutathione (GSH) level and inhibit HIV-1 replication in persistently infected Molt-4 and U937 cells. However, inhibition of HIV-1 replication appears not to be directly correlated with CSH levels. Cysteine and NAC also inhibit NF-x B activity as determined by electrophoretic mobility shift assays and chloramphenicol acetyl-transferase (CAT) gene expression under control of NF-x B binding sites in uninfected cells. This suggests that the cysteine deficiency in HIV-1-infected individuals may cause an over-expression of NF-x B-dependent genes and enhance HIV-1 replication. NAC may be considered for the treatment of HIV-1-infected individuals. (C) Lippincott-Raven Publishers.
Article
Three bioassays were conducted to determine the limiting order of amino acids for endogenous amino acid utilization in chicks fed a protein-free diet. The studies were conducted during the period 10 to 21 d posthatching. Experiment 1 was a deletion assay in which a protein-free basal diet was supplemented with an amino acid mixture containing methionine, cystine, threonine, arginine, phenylalanine and glutamine. Each amino acid, or methionine + cystine together, was then deleted singly from the amino acid mixture. Supplementing the protein-free basal diet with the amino acid mixture reduced weight loss. Deletion of methionine and cystine from the amino acid mixture increased (P < 0.05) weight loss. Deleting threonine from the amino acid mixture also resulted in weight loss that was intermediate between the amino acid-supplemented diet and the protein-free basal diet, indicating it was second limiting after sulfur amino acids. Experiments 2 and 3 were amino acid addition assays. Additions of methionine or cystine to the protein-free basal diet, either singly or in combination, resulted in lower rates of weight loss and protein depletion. Addition of threonine to the diet supplemented with methionine and cystine further reduced weight loss. These studies indicate that sulfur amino acids are the first-limiting amino acids for utilization of endogenous amino acids. However, our results clearly demonstrate that the primary need is for cystine, and not for methionine per se.
Article
Cocks were fed a protein-free diet supplemented with methionine plus arginine or glutamic acid for 25 days to investigate the effect of these amino acids on fecal and urinary nitrogen excretion. Addition of either methionine plus arginine or glutamic acid did not change the fecal nitrogen excretion. Methionine plus arginine supplementation reduced the urniary nitrogen excretion compared to the protein-free diet, whereas glutamic acid supplementation increased it. The reduced urinary nitrogen excretion resulting from supplementation with methionine plus arginine was mostly accounted for by a reduction in uric acid excretion. In the methionine plus arginine group, free amino acid analysis showed that free glutamine and glutamic acid content significantly decreased in liver while no differences were found in plasma. Since glutamine may play an important role in the formation of uric acid for chickens, the reduced amount of free glutamine and glutamic acid in the liver of cocks fed the diet suplemented with methionine plus arginine might account for the reduced excretion of uric acid, and therefore for the nitrogen sparing action of methionine plus arginine in chickens fed a protein-free diet.
Article
It was previously reported that methionine and threonine supplementation of a protein free diet had a greater nitrogen sparing action than methionine supplementation alone. Investigated in this study were (1) effects of depletion of labile body protein on the nitrogen sparing action of methionine and threonine supplementation of a protein free diet, (2) the effects of graded levels of methionine and threonine on urinary nitrogen excretion, (3) the effect of supplementation of other amino acids to the protein free diet plus methionine and threonine on the urinary excretion of nitrogen and (4) sex differences in the nitrogen sparing action of methionine and threonine supplementation of a protein free diet. After 10 days of feeding a protein free diet to deplete the body labile proteins, nitrogen excretion in urine of female rats was significantly reduced within the first 2 days of feeding a protein free diet supplemented with methionine and threonine. After 7 days of feeding a protein free diet supplemented with methionine and threonine, nitrogen excretion was reduced when even as little as 0.0188% of each amino acid was added. The supplementation of all essential amino acids to the protein free diet did not reduce the urinary excretion of nitrogen further than the supplementation of methionine and threonine, but improved the nitrogen balance slightly. The excretion of urinary nitrogen by female rats fed the protein free diet supplemented with small amounts (0.0188%) of methionine and threonine was significantly reduced after 7 to 14 days of feeding. In males, small amounts of methionine and threonine had no significant effect at either 7 or 14 days.
Article
Markedly decreased plasma cystine and cysteine concentrations have been found in HIV-infected patients at all stages of the disease and in SIV-infected rhesus macaques. The elevated glutamate levels found in the same patients aggravate the cysteine deficiency by inhibiting the membrane transport activity for cystine. The intact immune system appears to require a delicate balance between pro-oxidant and antioxidant conditions, maintained by a limited and well-regulated supply of cysteine. This balance is obviously disturbed in HIV infection and may contribute to the pathogenesis of AIDS.
Article
To determine whether a single oral dose of N-acetylcysteine corrects the deficiency of cysteine and glutathione in plasma and mononuclear cells of HIV-infected patients. Pharmacokinetic and pharmacodynamic study. Cysteine and glutathione were measured in plasma and peripheral blood mononuclear cells of patients at different stages of HIV infection before and after a single oral dose of N-acetylcysteine. At baseline, the plasma concentrations of glutathione and cysteine were significantly lower in HIV-infected patients than in healthy controls. The intracellular concentration of glutathione correlated with the absolute CD4 lymphocyte counts: the concentration of glutathione in mononuclear cells was significantly lower in patients with more advanced immunodeficiency. A single oral dose of N-acetylcysteine increased the concentration of cysteine in plasma and mononuclear cells of HIV-infected patients. Four hours after N-acetylcysteine administration, intracellular glutathione concentrations in the patients were moderately higher than at baseline and at 2 h. Oral N-acetylcysteine transiently increases the concentrations of cysteine and glutathione in mononuclear cells of patients with HIV infection. A sustained increase in intracellular cysteine may be necessary to normalize intracellular glutathione. This may be accomplished by repeat administration of N-acetylcysteine.
Article
Even moderate variations of the extracellular cysteine concentration were previously shown to affect T cell functions in vitro despite high concentrations of cystine. We therefore analyzed the membrane transport activities of T cells for cysteine and cystine, and the role of low molecular weight thiol in T cell-mediated host responses against a T cell tumor in vivo. A series of T cell clones and tumors including the highly malignant lymphoma L5178Y ESb and its strongly immunogenic variant ESb-D was found to express extremely weak transport activity for cystine but strong transport activity for cysteine. However, not all cells showed the expected requirement for cysteine (or 2-mercaptoethanol (2-ME)) in the culture medium. One group of clones and tumors including the malignant ESb-lymphoma did not respond to changes of extracellular cystine concentrations and was strongly thiol dependent. This group released only little acid soluble thiol (cysteine) if grown in cystine-containing cultures. The other T cell lines, in contrast, were able to maintain high intracellular GSH levels and DNA synthesis activity in cystine-containing culture medium without cystein or 2-ME and released substantial amounts of thiol. This group included the immunogenic ESb-D line. Additional thiol-releasing ESb variants were obtained by culturing large numbers of L5178Y ESb tumor cells in cultures without cysteine or 2-ME. All of these ESb variants showed a significantly decreased tumorigenicity and some of them induced cytotoxic and protective host responses even against the malignant ESb parent tumor. Taken together, our experiments suggest that the host response against a tumor may be limited in certain cases by the failure of the stimulator (i.e., the tumor) cell to deliver sufficient amounts of cysteine to the responding T cells.
Article
To establish whether the high plasma glutamate and low acid-soluble thiol (mainly cysteine) concentrations previously found in patients with HIV-1 infection are a consequence of the infection or a risk factor for its development, a closely related animal model, rhesus and fascicularis macaques infected with simian immunodeficiency virus (SIVmac251), was studied. The 23 infected macaques had significantly lower mean plasma thiol and higher glutamate concentrations than 18 uninfected controls (p less than 0.001). The changes were apparent by 1 week after infection. Thus, abnormal plasma glutamate and thiol concentrations are, at least in this model, a direct and early consequence of the retroviral infection.
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Maintenance of intracellular glutathione (GSH) levels has been implicated in blocking cytokine-stimulated HIV replication in vitro, in both acute and latent infection models. We demonstrate here that subsets of human peripheral blood mononuclear cells differ substantially in mean GSH levels, as measured on a cell-by-cell basis with the fluorescence-activated cell sorter (FACS): B cells have the lowest GSH levels; T cells are intermediate; and monocytes and macrophages have the highest levels. Furthermore, GSH levels subdivide the CD4 and CD8 T cell subsets into two classes each: high- and low-GSH cells, which cannot be distinguished by cell size or by currently known surface markers. Significantly, the high-GSH T cells are selectively depleted early during the HIV infection, and are effectively missing in all ARC and AIDS patients.
Article
The concentration of L-lactate in the blood plasma of higher vertebrates is about 1 mM but can be as high as 30 mM under certain physiological and pathological conditions or in the vicinity of glycolytically active cells including macrophages. Here we report that high but physiologically relevant concentrations of lactate increase the expression of interleukin 2 (IL 2)-specific mRNA and the production of IL 2 activity in cultures of mitogenically stimulated T cells. Lactate supports IL 2 production most effectively if added 0-8 h after T cell stimulation and only in cultures of CD4+ but not of CD8+ T cells. In contrast to the DNA synthesis activity in these cell cultures, IL 2 production is not augmented but rather inhibited by exogenous glutathione (GSH). Lactate causes a reduction of intracellular GSH levels, and lactate-containing cultures require accordingly higher extracellular cysteine concentrations than control cultures to achieve similar intracellular GSH levels. In view of the strong variations of extracellular lactate concentrations in vivo, our experiments suggest that lactate may be part of a previously unknown mechanism by which the metabolic microenvironment modulates gene expression in T cells.
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HIV-1 proviral DNA contains two binding sites for the transcription factor NF-kappa B. HIV-1-infected individuals have, on average, abnormally high levels of tumour necrosis factor alpha (TNF alpha) and abnormally low plasma cysteine levels. We therefore investigated the effects of cysteine and related thiols on HIV-1 replication and NF-kappa B expression. The experiments in this report show that cysteine or N-acetylcysteine (NAC) raise the intracellular glutathione (GSH) level and inhibit HIV-1 replication in persistently infected Molt-4 and U937 cells. However, inhibition of HIV-1 replication appears not to be directly correlated with GSH levels. Cysteine and NAC also inhibit NF-kappa B activity as determined by electrophoretic mobility shift assays and chloramphenicol acetyl-transferase (CAT) gene expression under control of NF-kappa B binding sites in uninfected cells. This suggests that the cysteine deficiency in HIV-1-infected individuals may cause an over-expression of NF-kappa B-dependent genes and enhance HIV-1 replication. NAC may be considered for the treatment of HIV-1-infected individuals.
Article
Glutathione (GSH) is known to play an important role in various lymphocyte functions. We now report that different T cell subsets express different requirements for intracellular GSH. Depletion of intracellular GSH by buthionine sulfoximine (BSO), a specific inhibitor of GSH biosynthesis, decreases the proportion of CD8+ cells (i.e., increases the CD4+/CD8+ ratio), and inhibits particularly the generation of large blast-like CD8+ cells and cytotoxic T lymphocyte (CTL) activity. CTL activity is restored by administration of exogenous GSH. Differential effects of GSH depletion were also seen at the level of individual T cell clones. The CD4+ helper T cell clone D10.G4.1.HD was found to express a high rate of interleukin 2 (IL-2) dependent DNA synthesis even after severe depletion of intracellular GSH, whereas other T cell clones including the clone 29 were severely inhibited by BSO. The results of these studies suggest that the decreased intracellular GSH levels of HIV-1 seropositive persons are probably not (directly) responsible for the selective depletion of the CD4+ T cell subset but may be responsible for a cellular dysfunction of the CD8+ subset and for the ultimate failure of the CTL to control the viral infection in these patients.
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Hydrogen peroxide and oxygen radicals are agents commonly produced during inflammatory processes. In this study, we show that micromolar concentrations of H2O2 can induce the expression and replication of HIV-1 in a human T cell line. The effect is mediated by the NF-kappa B transcription factor which is potently and rapidly activated by an H2O2 treatment of cells from its inactive cytoplasmic form. N-acetyl-L-cysteine (NAC), a well characterized antioxidant which counteracts the effects of reactive oxygen intermediates (ROI) in living cells, prevented the activation of NF-kappa B by H2O2. NAC and other thiol compounds also blocked the activation of NF-kappa B by cycloheximide, double-stranded RNA, calcium ionophore, TNF-alpha, active phorbol ester, interleukin-1, lipopolysaccharide and lectin. This suggests that diverse agents thought to activate NF-kappa B by distinct intracellular pathways might all act through a common mechanism involving the synthesis of ROI. ROI appear to serve as messengers mediating directly or indirectly the release of the inhibitory subunit I kappa B from NF-kappa B.
Article
The stimulation of DNA synthesis in lymphocyte populations was previously shown to depend strongly on the intracellular glutathione (GSH) level. Since T cell growth is known to depend on interleukin 2 (IL-2), the experiments in this report were designed to determine whether intracellular GSH depletion may inhibit IL-2 production or the IL-2 dependent DNA synthesis. Our experiments revealed that IL-2 production and DNA synthesis of mitogenically stimulated splenic T cells have indeed different requirements for GSH. The addition of relatively high concentrations of GSH (5 mM) to cultures of concanavalin A (Con A)-stimulated splenic T cells was found to augment strongly the DNA synthesis but inhibited the production of IL-2. Moderate intracellular GSH levels, however, are apparently not inhibitory for IL-2 production, since intracellular GSH depletion by cysteine starvation or by graded concentrations of DL-buthionine sulfoximine (BSO) had virtually no effect on IL-2-specific mRNA expression and the production of T cell growth factor (TCGF). The DNA synthesis activity, in contrast, was strongly suppressed after GSH depletion with either method. As in cultures of splenic T cells, GSH depletion had no substantial effect on the induction of IL-2 mRNA and TCGF production in several mitogenically stimulated T cell clones. Taken together, our experiments suggest that complex immune response may operate best at intermediate GSH levels that are not too high to inhibit IL-2 production but sufficient to support DNA synthesis.
Article
Macrophages consume cystine and generate approximately equivalent amounts of acid-soluble thiol. Stimulation of macrophages with bacterial lipopolysaccharide (LPS) or tumor necrosis factor (TNF) strongly augments the amount of thiol released into the culture supernatant. Cysteine constitutes most of the acid-soluble thiol. The intracellular glutathione level and the DNA synthesis activity in mitogenically stimulated lymphocytes are strongly increased by either exogenously added cysteine, or (syngeneic) macrophages. This cysteine dependency is observed even in the presence of relatively high extracellular cystine concentration as they occur in the blood plasma. The extracellular cysteine concentration also has a strong influence on the intracellular glutathione concentration, viability, and DNA synthesis of cycling T cell clones. Moreover, the cysteine concentration in the culture medium on Day 3 and Day 4 of a 5-day allogeneic mixed lymphocyte culture (i.e., in the late phase of incubation) has a strong influence on the generation of cytotoxic T cell activity, indicating that regulatory effects of cysteine are not restricted to the early phase of the blastogenic response. The inhibitory effect of cysteine starvation on the DNA synthesis of the T cell clones and on the activation of cytotoxic T lymphocytes can be explained essentially by the depletion of intracellular glutathione, since similar effects are observed after treatment with buthionine sulfoximine (BSO), a specific inhibitor of the glutathione biosynthesis. BSO has practically no influence, however, on the N alpha-benzyloxycarbonyl Ne-t-butyloxycarbonyl-L-lysine-thiobenzyl-ester (BLT)-esterase activity and hemolytic activity of the cell lysates from cytotoxic T cells against sheep red blood cells (perforin activity). Taken together, our experiments indicate that cysteine has a regulatory role in the immune system analogous to the hormone-like lymphokines and cytokines. It is released by macrophages at a variable and regulated rate and regulates immunologically relevant functions of lymphocytes in the vicinity.
Article
To find out whether systemic glutathione deficiency is associated with human immunodeficiency virus (HIV) infection, thus contributing to the immunodeficiency state, glutathione concentrations in venous plasma and lung epithelial lining fluid (ELF) of symptom-free HIV-seropositive and normal individuals were measured. Total and reduced glutathione concentrations in the plasma of the HIV-infected subjects were about 30% of those in the normal individuals. Concentrations of these substances in the ELF of HIV-infected subjects were about 60% of those in the controls. There was no correlation between ELF and plasma concentrations of total or reduced glutathione. Since glutathione enhances immune function, glutathione deficiency may contribute to the progressive immune dysfunction of HIV infection.
Article
Blood plasma samples from HIV-1-infected persons contain elevated glutamate concentrations up to 6-fold the normal level and relatively low concentrations of acid-soluble thiol (i.e. decreased cysteine concentrations). The intracellular glutathione concentration in peripheral blood-mononuclear cells (PBMC) and monocytes from HIV antibody-positive persons are also significantly decreased. Therapy with azidothymidine (AZT) causes a substantial recovery of the plasma thiol levels; but glutamate levels remain significantly elevated and intracellular glutathione levels remain low. Cell culture experiments with approximately physiological amino-acid concentrations revealed that variations of the extracellular cysteine concentration have a strong influence on the intracellular glutathione level and the rate of DNA synthesis [( 3H]thymidine incorporation) in T cell clones and human and murine lymphocyte preparations even in the presence of several-fold higher cystine and methionine concentrations. Cysteine cannot be replaced by a corresponding increase of the extracellular cystine or methionine concentration. These experiments suggest strongly that the low cysteine concentration in the plasma of HIV-infected persons may play a role in the pathogenetic mechanism of the acquired immunodeficiency syndrome.
Article
The acquired immunodeficiency syndrome (AIDS) is accompanied by a metabolic disturbance. Serum samples from persons with antibodies against the AIDS associated human immunodeficiency virus (HIV/LAV/HTLV III) including persons without overt symptoms, patients with lymphadenopathy syndrome (LAS) and patients with AIDS or AIDS-related complex (ARC) contain on the average significantly elevated concentrations of arginine and glutamate. The serum from patients with overt AIDS contains also, on the average, significantly reduced concentrations of methionine and cystine. In vitro experiments revealed that the [3H]thymidine incorporation by mitogenically stimulated murine lymphocytes and cloned T cells is inhibited by an elevation of the extracellular glutamate concentration and augmented by the addition of cysteine. This suggests the possibility that the abnormal concentrations of glutamate and cystine in the blood of HIV-infected persons may contribute to the defect in the lymphoid system.
Article
Activated macrophages are known to release a variety of immunoregulatory substances including the low-molecular-weight substances hydrogen peroxide and lactate. We report here that lactate but not hydrogen peroxide is capable of supporting a substantial production of T-cell growth factor (TCGF) in cultures of accessory cell-depleted splenic T-cell populations after stimulation with concanavalin A. Hydrogen peroxide and its biosynthetic precursor superoxide anion (O2-) mediate, however, a strong augmentation of the TCGF production by accessory cell-depleted T-cell populations in the presence of lactate. Lactate inhibits the incorporation of [3H]thymidine in short-term cultures (18-26 hr) of accessory cell-depleted T cells. This confirms the rule that (optimal) production of T-cell growth factor requires a growth inhibitory signal. Concentrations of hydrogen peroxide which augment TCGF production most effectively (i.e., 1 X 10(-5) M) do not inhibit the incorporation of [3H]thymidine; and higher concentrations (3 X 10(-5)-1 X 10(-4) M) of hydrogen peroxide inhibit both the production of TCGF and the incorporation of [3H]thymidine. In agreement with the augmenting effect of hydrogen peroxide on TCGF production, it was observed that the proliferative response in mixed lymphocyte cultures is suppressed by catalase and augmented by 1 X 10(-5) M H2O2. Proliferative and cytotoxic responses in mixed lymphocyte cultures with an external source of interleukin 2 (IL-2) in contrast, are not augmented by 1 X 10(-5) M H2O2. The relatively high concentration of 1 X 10(-4) M hydrogen peroxide was found to inhibit the proliferative responses in mixed lymphocyte cultures with or without external IL-2 but not the cytotoxic response in the presence of IL-2. This indicates that CTL precursor cells may be relatively resistant against H2O2.
Article
Cysteine and cystine transport activities of resting and activated mouse spleen lymphocytes were characterized in order to examine the contributions of cysteine and cystine to intracellular glutathione contents. Following stimulation with lipopolysaccharide, the lymphocytes markedly increased their capacity to transport cysteine. The uptake of cysteine was mediated mainly by the ASC system (Na+-dependent neutral amino acid transport system especially reactive with alanine, serine, and cysteine). On the other hand, both the resting and the activated lymphocytes had extremely low cystine transport activities. Because of the instability of cysteine, the culture media usually contained cystine but not cysteine. Therefore, both the resting and the activated lymphocytes rapidly decreased their glutathione contents owing to their poor capacities to take up cystine. The effects of freshly added cysteine on the cellular glutathione contents were examined in the presence of bathocuproinedisulfonate, a nontoxic copper-specific chelator that inhibits autoxidation of cysteine. Cysteine added at 25-400 microM only partially prevented the rapid decrease of the glutathione contents in fresh resting lymphocytes. In the lipopolysaccharide-activated cells, however, cysteine enhanced the cellular glutathione contents in a dose-dependent manner. These results indicate that the enhanced activity of the ASC system increases the level of intracellular glutathione in the presence of cysteine.
Article
The role of glutathione (GSH) in lectin-induced lymphocyte activation can be studied by quantitating lectin-induced nuclear size transformation in the presence of variable degrees of GSH depletion. Buthionine sulfoximine (BSO) inhibits intracellular GSH synthesis by inhibition of the enzyme gamma-glutamyl-cysteine synthetase. By combining endogenous GSH depletion in cell cultures with BSO-induced inhibition of GSH synthesis, lectin-induced lymphocyte activation can be studied at various concentrations of soluble intracellular GSH. With this approach, the percentage of lymphocytes undergoing a nuclear size transformation is minimally affected despite depletion of soluble intracellular GSH to 0.27 nmol/10(7) cells (PBL), which represents approximately 95% depletion of intracellular GSH. When soluble intracellular GSH is depleted to undetectable levels (less than 0.10 nmol/10(7) cells) there is a 10 to 12% reduction in the number of cell nuclei transformed. However, in all BSO-pretreated cultures the lectin-induced nuclear size transformation is intermediate between resting and blast-transformed lymphocytes, suggesting only partial (or aborted) activation. The partial activation response observed in BSO-pretreated cultures may be due to mobilization of the protein-bound pool of GSH, which is relatively resistant to depletion by BSO. That the inhibition of full blast transformation is truly due to GSH depletion was proven by experiments in which GSH was repleted exogenously and a full blast transformation was restored. The results of previous work in our laboratory had shown that the sulfhydryl-reactive agent 2-cyclohexene-1-one (2-CHX) was a potent inhibitor of activation at soluble intracellular GSH concentrations well above 0.27 nmol/10(7) PBL. In the present study, the dose-dependent inhibition of activation by 2-CHX was confirmed, but it was shown that the degree of inhibition caused by 2-CHX could be at least partially dissociated from the level of intracellular GSH present at the time of lectin addition and that the inhibitory potential of 2-CHX exceeded that of BSO at comparable levels of soluble intracellular GSH. Thus, the inhibitory properties of 2-CHX cannot be accounted for solely on the basis of GSH depletion.
Article
Der Einfluß einer Methionin-Ergänzung zu einer eiweißfreien Ration auf die Stickstoffausscheidung bei Ratten und Schweinen 1. Sechs Gruppen von je 8 männlichen Albino-Ratten im Alter von 3 Monaten und von 260 bis 300 g Körpergewicht (KG) wurden während einer Vorversuchsperiode von 18 Tagen mit Rationen gefüttert, die Weizengluten oder mit Methionin angereichertes Kasein als einzige Eiweißquelle enthielten. Drei Eiweißniveaus wurden angewendet. Die N-Bilanz, bestimmt während der letzten 4 Tage der Vorversuchs-periode, betrug durchschnittlich -22, 98 und 229 mg N/kg KG0,75 entsprechend in den Gruppen mit niedrigem, mittlerem und hohem Eiweißniveau. 2. Jede Gruppe wurde anschließend in 2 Untergruppen eingeteilt, die während der 12 Tage langen Versuchsperiode mit einer eiweißfreien Ration, mit oder ohne Zusatz von 11 mg Methionin pro Tag und Tier, gefüttert wurden. Harn und Faeces wurden von jedem Tier gesondert gesammelt und der Gehalt an Gesamt-, Harnstoff- und Kreatinin-N bestimmt. 3. Die N-Ausscheidung in den Faeces war höher bei Ratten, die mit höheren Eiweißgaben vorgefüttert waren. Sie ging stufenweise zurück und war unabhängig von der Art des vorgefütterten Eiweißes und dem Methionin-Zusatz. 4. Der Methionin-Zusatz hatte keinen Einfluß auf die N-Ausscheidung im Harn von mit niedrigerem Eiweißniveau vorgefütterten Ratten, dagegen beeinflußte er significant die renale Ausscheidung von Gesamt- und Harnstoff-N in Gruppen, die mit mittlerem oder hohem Eiweißniveau vorgefüttert waren. Die Körpergewichtsverluste in diesen Gruppen wurden auch durch Methionin-Zusatz significant herabgesetzt. Daraus ist zu schließen, daß der Methionin-Zusatz zum eiweißfreien Futter die N-Ausscheidung im Harn nur bei Tieren mit nicht erschöpften Reserven von labilem Eiweiß herabsetzt und daß diese Wirkung mit der Resynthese dieser Reserven zusammenhängt. 5. In einem Vorversuch mit 4 bis 5 Monate alten Schweinen (Kastraten), die mit hohen Eiweißgaben gefüttert wurden, setzte auch ein Methionin-Zusatz (4 g täglich für Schweine von 76 kg Gewicht) die N-Ausscheidung im Harn significant herab. 6. Die Ausscheidung von Kreatinin-N bei Ratten war gleichmäßig und unabhängig von der vorgefütterten Futterration. Weder bei Ratten noch bei Schweinen wurde sie durch Methionin-Zusatz beeinflußt und war proportional zum jeweiligen Lebendgewicht (etwa 15 mg N/kg) nicht aber zu seiner Bruchpotenz.
Article
Lymphocytes may be stimulated to undergo transformation into blast cells by a variety of nonspecific chemical agents, or by antiglobulin or antilymphocyte antisera. Antigens act as mitogenic agents on lymphocytes from sensitized animals (1-3). We have now observed that the action of various mitogens on human and rabbit peripheral lymphoeytes is markedly enhsnced in the presence of the reducing agents, l-cysteine, glutathione, or sulfite. Rabbit peripheral lymphocytes were isolated, purified on a cotton-wool column and cultured as described previously (4). Human leukocytes were obtained from the blood of normal individuals by allowing the cells to settle in 10% Plasmagel (Laboratoire Roger Bellon, Neuilly, France). The leukocyte-rich layer was removed and the cells were washed several times with 20% decomplemented fetal calf serum in Eagle's minimal essential medium (MEM). Leukocytes were cultured (4) at a concentration of 106 cells/ml. Rabbit lymphoeytes were cultured for 48 hr and human leukocytes for 108 hr; 24 hr before harvest, 5 µCi of 3H-thymidine (6.7 Ci/mM) was added to the rabbit lymphocyte cultures and either 5 µCi or 2 µCi of 3H-thymidine to the human leukocyte cultures. Radioactivity incorporated into DNA was determined as described previously (4).
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This review is concerned with current progress in unraveling the biochemical bases of the physiological roles of this important compound. Detailed information is now available about glutathione synthesis and its metabolism by the reactions of the γ-glutamyl cycle, and its function in reductive processes that are essential for the synthesis (and the degradation) of proteins, formation of the deoxyribonucleotide precursors of DNA, regulation of enzymes, and protection of the cell against reactive oxygen compounds and free radicals. In addition, glutathione is a coenzyme for several reactions; it conjugates with foreign compounds (e.g. drugs) and with compounds formed in metabolism (e.g. estrogens, prostaglandins, leukotrienes), and thus participates in their metabolism. An important recent finding is that cellular turnover of glutathione is associated with its transport, in the form of GSH, out of cells. The functions of such GSH transport include formation by membrane-bound γ-glutamyl transpeptidase of γ-glutamyl amino acids, which can be transported into certain cells, and thus serve as one mechanism of amino acid transport. Transported GSH probably also functions in reductive reactions that may involve the cell membrane and the immediate environment of the cell. In the mammal, such transported GSH may enter the blood plasma and be transferred to other cells. Glutathione thus appears to be a storage form and a transport form of cysteine. Much of the new information about glutathione has arisen through studies with selective inhibitors of the enzymes involved in its metabolism. Thus, inhibition in vivo of γ-glutamyltranspeptidase, γ-glutamyl cyclotransferase, 5-oxoprolinase, and glutathione synthesis has been achieved, and the effects observed have contributed importantly to the understanding of glutathione metabolism and function. Studies on the inhibition of γ-glutamyl transpeptidase have elucidated the transport of GSH and the formation and transport of γ-glutamyl amino acids. Inhibition of glutathione synthesis by sulfoximine compounds that inactivate γ-glutamylcysteine synthetase has also contributed to such knowledge, as well as to information about the roles of glutathione in protection against both free radicals and reactive oxygen compounds, and in metabolism. These enzyme inhibitors, and other compounds that increase in vivo glutathione synthesis have opened the way to selective modulation of glutathione metabolism; this has made several new therapeutic approaches possible.
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Glutathione, a tripeptide thiol found in virtually all cells, functions in metabolism, transport, and cellular protection. It participates in the reduction of disulfides and other molecules, and conjugates with compounds of exogenous and endogenous origin. It protects cells against the destructive effects of reactive oxygen intermediates and free radicals. Modifications of glutathione metabolism may be achieved by administration of selective enzyme inhibitors, and also by giving compounds that increase glutathione synthesis. Such effects are useful in chemotherapy and radiation therapy and in protecting cells against the toxic effects of drugs, other foreign compounds, and oxygen.
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Activation of the CD28 surface receptor provides a major costimulatory signal for T cell activation resulting in enhanced production of interleukin-2 (IL-2) and cell proliferation. In primary T lymphocytes we show that CD28 ligation leads to the rapid intracellular formation of reactive oxygen intermediates (ROIs) which are required for CD28-mediated activation of the NF-kappa B/CD28-responsive complex and IL-2 expression. Delineation of the CD28 signaling cascade was found to involve protein tyrosine kinase activity, followed by the activation of phospholipase A2 and 5-lipoxygenase. Our data suggest that lipoxygenase metabolites activate ROI formation which then induce IL-2 expression via NF-kappa B activation. These findings should be useful for therapeutic strategies and the development of immunosuppressants targeting the CD28 costimulatory pathway.
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Plasma concentrations of 21 amino acids were determined for 20 control subjects and 20 subjects infected with human immunodeficiency virus type 1 (HIV). Compared with the control subjects, the HIV-infected group had lower cystine, tryptophan, and methionine (decreased 67%, 52%, and 32%, respectively, P < 0.001 for each) and increased taurine (230%, P < 0.001) and lysine concentrations (30%, P < 0.001). Other amino acid concentrations changed modestly. Amounts of cystine, tryptophan, methionine, taurine, and lysine did not differ significantly between subgroups of HIV-infected subjects with > 200 (n = 6) or < 200 (n = 14) CD4+ lymphocytes per microliter, suggesting that the concentrations decrease soon after infection and change little thereafter. Activation of metabolism of cystine to taurine may explain reciprocal changes in these amino acids and known depletion of cystine and glutathione. The selective changes in amino acid profiles observed during HIV infection differ from those recognized for malnutrition or other pathological processes.
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HIV-infected individuals and SIV-infected rhesus macaques have, on the average, decreased plasma cysteine and cystine concentrations and decreased intracellular glutathione levels. We show that the cysteine supply and the intracellular glutathione levels have a strong influence on the T cell system. A study of healthy human subjects revealed that persons with intracellular glutathione levels of 20-30 nmol/mg protein had significantly higher numbers of CD4+ T cells than persons with either lower or higher glutathione levels. Persons who moved during a 4-week observation period from the optimal to the suboptimal range (10-20 nmol/mg) experienced, on the average, a 30% decrease in CD4+ T cell numbers. This decrease was prevented by treatment with N-acetyl-cysteine (NAC). NAC caused this relative increase of CD4+ T cell numbers in spite of decreasing glutathione levels and not by increasing the glutathione level. Our studies suggest that the immune system may be exquisitely sensitive not only against a cysteine and glutathione deficiency but also against an excess of cysteine.
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Even a moderate increase in the cellular cysteine supply elevates the intracellular glutathione (GSH) and glutathione disulfide (GSSG) levels and potentiates immunological functions of lymphocytes in vitro. At low GSSG levels, T cells cannot optimally activate the immunologically important transcription factor NF kappa B, whereas high GSSG levels inhibit the DNA binding activity of NF kappa B. The effects of GSSG are antagonized by reduced thioredoxin (TRX). As the protein tyrosine kinase activities p56lck and p59fyn are activated in intact cells by hydrogen peroxide, they are likely targets for GSSG action. These redox-regulated enzymes trigger signal cascades for NF kappa B activation and transduce signals from the T cell antigen receptor, from CD4 and CD8 molecules, and from the IL-2 receptor beta-chain. The effector phase of cytotoxic T cell responses and IL-2-dependent functions are inhibited even by a partial depletion of the intracellular GSH pool. As signal transduction is facilitated by prooxidant conditions, we propose that the well-known immunological consequences of GSH depletion ultimately may be results of the accompanying GSSG deficiency. As HIV-infected patients and SIV-infected rhesus macaques have, on the average, significantly decreased plasma cyst(e)ine and intracellular GSH levels, we also hypothesize that AIDS may be the consequence of a GSSG deficiency as well.
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The transcription factors NF-kappa B and AP-1 have been implicated in the inducible expression of a variety of genes involved in responses to oxidative stress and cellular defense mechanisms. Here, we report that thioredoxin, an important cellular protein oxidoreductase with antioxidant activity, exerts different effects on the activation of NF-kappa B and AP-1. Transient expression or exogenous application of thioredoxin resulted in a dose-dependent inhibition of NF-kappa B activity, as demonstrated in gel shift and transactivation experiments. AP-1-dependent transactivation, in contrast was strongly enhanced by thioredoxin. A similar increase of AP-1 activity was also observed with other, structurally unrelated antioxidants such as pyrrolidine dithiocarbamate and butylated hydroxyanisole, indicating that the thioredoxin-induced increase of AP-1 activation was indeed based on an antioxidant effect. Moreover, the stimulatory effect on AP-1 activity was found to involve de novo transcription of the c-jun and c-fos components but to be independent of protein kinase C activation. These results suggest that thioredoxin plays an important role in the regulation of transcriptional processes and oppositely affects NF-kappa B and AP-1 activation.
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Oxidative conditions potentiate the activation of the nuclear transcription factor kappa B (NF kappa B) and the activator protein-1 (AP-1) in intact cells, but inhibit their DNA binding activity in vitro. We now show that both the activation of NF kappa B and the inhibition of its DNA binding activity is modulated in intact cells by the physiological oxidant glutathione disulphide (GSSG). NF kappa B activation in human T lineage cells (Molt-4) by 12-O-tetradecanoyl-phorbol 13-acetate was inhibited by dithiothreitol, and this was partly reversed by the glutathione reductase inhibitor 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) or by hydrogen peroxide, indicating that GSSG may be required for NF kappa B activation. These effects of BCNU and hydrogen peroxide were not seen in glutathione-depleted cells. However, NF kappa B and AP-1 activation were potentiated by dithiothreitol if added to cell cultures 1 h after the phorbol ester, indicating that a shift of redox conditions may support optimal oxidative activation with minimal inhibition of DNA binding. The elevation of intracellular GSSG levels by BCNU before stimulation suppressed the chloramphenicol acetyltransferase expression dependent on NF kappa B but increased that dependent on AP-1. This selective suppression of NF kappa B was also demonstrable by electrophoretic mobility shift assays. In vitro, GSSG inhibited the DNA binding activity of NF kappa B more effectively than that of AP-1, while AP-1 was inhibited more effectively by oxidized thioredoxin.
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We show that AP-1 is an antioxidant-responsive transcription factor. DNA binding and transactivation by AP-1 were induced in HeLa cells upon treatment with the antioxidants pyrrolidine dithiocarbamate (PDTC) and N-acetyl-L-cysteine (NAC), and upon transient expression of the antioxidative enzyme thioredoxin. While PDTC and NAC enhanced DNA binding and transactivation of AP-1 in response to phorbol ester, the oxidant H2O2 suppressed phorbol ester activation of the factor. H2O2 on its own was only a weak inducer of AP-1. Activation of AP-1 by PDTC was dependent on protein synthesis and involved transcriptional induction of c-jun and c-fos genes. Transcriptional activation of c-fos by PDTC was conferred by the serum response element, suggesting that serum response factor and associated proteins function as primary antioxidant-responsive transcription factors. In the same cell line, the oxidative stress-responsive transcription factor NF-kappa B behaved in a manner strikingly opposite to AP-1. DNA binding and transactivation by NF-kappa B were strongly activated by H2O2, while the antioxidants alone were ineffective. H2O2 potentiated the activation of NF-kappa B by phorbol ester, while PDTC and NAC suppressed PMA activation of the factor. PDTC did not influence protein kinase C (PKC) activity and PKC activation by PMA, indicating that the antioxidant acted downstream of and independently from PKC.
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In a double-blind placebo-controlled trial, human immunodeficiency virus (HIV)-seropositive patients with a CD4 lymphocyte cell count of more than 200 x 10(6) . l-1 were randomised to receive either 800 mg N-acetylcysteine (NAC) or placebo for 4 months. Before treatment low plasma cysteine levels, high free radical activity in neutrophils in the presence of autologous plasma-measured by the nitroblue tetrazolium (NBT) test- and increased tumor necrosis factor (TNF)-alpha levels were found in the HIV positive patients. After treatment the low plasma cysteine level in the NAC group increased to normal, and the decline of the CD4+ lymphocyte count before the study start, was less steep in the NAC group than in the placebo group after treatment. There was also a reduction in TNF-alpha level. However, NAC had no effect on the radical production by neutrophils, and although it did not increase the CD4+ cell count, it may have decreased the decline in CD4+ cells. Further controlled trials with NAC are needed to determine whether it has a beneficial effect in the treatment of asymptomatic HIV-infected individuals.
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To establish whether the low cysteine and glutathione levels in HIV-infected patients and SIV-infected rhesus macaques may be consequences of an abnormal cysteine catabolism, we analyzed sulfate and glutathione levels in macaques. Muscle tissue (m. vastus lateralis and m. gastrocnemius) of SIV-infected macaques (n = 25) had higher sulfate and lower glutathione and glutamate levels than that of uninfected controls (n =9). Hepatic tissue, in contrast, showed decreased sulfate and glutathione disulfide (GSSG) levels, and increased gamma-glutamylcysteine synthetase (gamma-GCS) activity. These findings suggest drainage of the cysteine pool by increased cysteine catabolism in skeletal muscle tissue, and by increased hepatic glutathione biosynthesis. Cachectic macaques also showed increased urea levels and decreased glutamine/urea ratios in the liver, which are obviously related to the abnormal urea excretion and negative nitrogen balance commonly observed in cachexia. As urea production and net glutamine synthesis in the liver are strongly influenced by proton-generating processes, the abnormal hepatic urea production may be the direct consequence of the cysteine deficiency and the decreased catabolic conversion of cysteine into sulfate and protons in the liver.
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Patients with skeletal muscle catabolism (cachexia) fail to conserve the skeletal muscle protein and release large amounts of nitrogen as urea. Previous studies suggest that the threshold for the conversion of amino acids into other forms of chemical energy and the concomitant production of urea are regulated by the plasma cystine level and hepatic cysteine catabolism. Studies of plasma amino acid exchange rates in the lower extremities now show that healthy young subjects regulate their plasma cystine level in a process that may be described as controlled constructive catabolism. The term controlled describes the fact that the release of cystine and other amino acids from the peripheral tissue is negatively correlated with (certain) plasma amino acid levels. The term constructive describes the fact that the release of cystine is correlated with an increase of the plasma cystine level. The regulation of the plasma cystine level is disturbed in conditions with progressive skeletal muscle catabolism including cancer, HIV infection, and old age. These conditions show also a low plasma glutamine:cystine ratio indicative of an impaired hepatic cystine catabolism. In HIV+ patients and SIV-infected macaques, a decrease of the plasma cystine level was found to coincide with the decrease of CD4+ T cells.
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Glutathione (GSH), a cysteine-containing tripeptide, is essential for the viability and function of virtually all cells. In vitro studies showing that low GSH levels both promote HIV expression and impair T cell function suggested a link between GSH depletion and HIV disease progression. Clinical studies presented here directly demonstrate that low GSH levels predict poor survival in otherwise indistinguishable HIV-infected subjects. Specifically, we show that GSH deficiency in CD4 T cells from such subjects is associated with markedly decreased survival 2-3 years after baseline data collection (Kaplan-Meier and logistic regression analyses, P < 0.0001 for both analyses). This finding, supported by evidence demonstrating that oral administration of the GSH prodrug N-acetylcysteine replenishes GSH in these subjects and suggesting that N-acetylcysteine administration can improve their survival, establishes GSH deficiency as a key determinant of survival in HIV disease. Further, it argues strongly that the unnecessary or excessive use of acetaminophen, alcohol, or other drugs known to deplete GSH should be avoided by HIV-infected individuals.
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Growing evidence has indicated that cellular reduction/oxidation (redox) status regulates various aspects of cellular function. Oxidative stress can elicit positive responses such as cellular proliferation or activation, as well as negative responses such as growth inhibition or cell death. Cellular redox status is maintained by intracellular redox-regulating molecules, including thioredoxin (TRX). TRX is a small multifunctional protein that has a redox-active disulfide/dithiol within the conserved active site sequence: Cys-Gly-Pro-Cys. Adult T cell leukemia-derived factor (ADF), which we originally defined as an IL-2 receptor alpha-chain/Tac inducer produced by human T cell lymphotrophic virus-I (HTLV-I)-transformed T cells, has been identified as human TRX. TRX/ADF is a stress-inducible protein secreted from cells. TRX/ADF has both intracellular and extracellular functions as one of the key regulators of signaling in the cellular responses against various stresses. Extracellularly, TRX/ADF shows a cytoprotective activity against oxidative stress-induced apoptosis and a growth-promoting effect as an autocrine growth factor. Intracellularly, TRX/ADF is involved in the regulation of protein-protein or protein-nucleic acid interactions through the reduction/oxidation of protein cysteine residues. For example, TRX/ADF translocates from the cytosol into the nucleus by a variety of cellular stresses, to regulate the expression of various genes through the redox factor-1 (Ref-1)/APEX. Further studies to clarify the regulatory roles of TRX/ADF and its target molecules may elucidate the intracellular signaling pathways in the responses against various stresses. The concept of "redox regulation" is emerging as an understanding of the novel mechanisms in the pathogenesis of several disorders, including viral infections, immunodeficiency, malignant transformation, and degenerative disease.
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Initial investigations demonstrated a deficiency of glutathione (GSH) in the epithelial lining fluid (ELF) of HIV-seropositive patients. In a recent study, our laboratory was unable to document such a deficiency. The current study was performed in an attempt to reconcile those disparate findings. To determine if ELF GSH decreases over time in asymptomatic HIV-seropositive subjects. Prospective, longitudinal study. Major university medical center. Thirty-three asymptomatic HIV-seropositive volunteers. None. BAL was performed on 33 asymptomatic HIV-seropositive subjects at baseline, 6 months later, and 12 months later. The volume of ELF and the concentration of GSH and oxidized GSH were determined. The concentration of total GSH in ELF was 689.0+/-100.4 microM. This significantly decreased when measured 6 and 12 months later (355.9+/-41.7 microM, and 397.9+/-52.7 microM, respectively, p=0.01, compared with baseline, both comparisons). Significant decreases were also noted in the HIV-seropositive subjects who smoked cigarettes (baseline--762.6+/-142.4 microM; 6 months--373.7+/-45.9 microM; 12 months--459.3+/-73.8 microM, p<0.03, for baseline vs 6 months, and baseline vs 12 months). In nonsmoking HIV-seropositive subjects, there was a decrease in ELF GSH over time, but it did not reach statistical significance (baseline--589.1+/-138.2 microM; 6 months--335.3+/-74.1 microM; 12 months--345.8+/-74.0 microM, p>0.1, all comparisons). The percentage of total GSH in the oxidized form was similar at all three time points (baseline--3.8+/-0.5%; 6 months--3.1+/-0.5%; 12 months--3.9+/-0.9%, p>0.1, all comparisons). The current study demonstrates that the GSH level in ELF is significantly decreased in HIV-seropositive subjects 6 and 12 months after the initial determination.