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Glutathione Disulfide (glutathione + disulfide)
Selected AbstractsMechanisms for sensitization to TNF-induced apoptosis by acute glutathione depletion in murine hepatocytesHEPATOLOGY, Issue 6 2003Katsuhiko Matsumaru We previously reported that depletion of glutathione in murine hepatocytes by diethylmaleate (DEM) or acetaminophen (APAP) leads to oxidative stress,dependent necrosis and sensitizes to tumor necrosis factor (TNF)-induced apoptosis in an oxidative stress,independent fashion, which could not be explained by interference with nuclear factor ,B (NF-,B) nuclear translocation. The present report explores the mechanisms of these effects. We observed that DEM led to necrosis when both mitochondrial and cytosol glutathione were depleted profoundly but sensitized to TNF-induced apoptosis when cytosol glutathione was depleted selectively. DEM and APAP lead to a significant decrease in reduced glutathione (GSH)/glutathione disulfide (GSSG) ratio. Glutathione depletion by DEM or APAP was associated with inhibition of TNF-induced NF-,B transactivation of anti-apoptotic genes, including inducible nitric oxide synthase (i-NOS). Provision of exogenous NO partially abrogated the sensitization to TNF in response to glutathione depletion. Glutathione depletion alone led to sustained increase in phospho-jun levels and c-Jun-N-terminal kinase (JNK) activity. JNK inhibitor partially blocked the sensitization to TNF-induced apoptosis accompanying glutathione depletion. In conclusion, these findings suggest that extramitochondrial glutathione depletion alters the thiol-disulfide redox state, leading to inhibition of NF-,B transactivation of survival genes and to sustained activation of JNK, both of which contribute to the sensitization to TNF-induced apoptosis. [source] Disposable Amperometric Sensors for Thiols with Special Reference to GlutathioneELECTROANALYSIS, Issue 18 2008Dipankar Bhattacharyay Abstract The antioxidant ,reduced glutathione' tripeptide is conventionally called glutathione (GSH). The oxidized form is a sulfur-sulfur linked compound, known as glutathione disulfide (GSSG). Glutathione is an essential cofactor for antioxidant enzymes; it provides protection also for the mitochondria against endogenous oxygen radicals. The ratio of these two forms can act as a marker for oxidative stress. The majority of the methods available for estimation of both the forms of glutathione are based on colorimetric and electrochemical assays. In this study, electrochemical sensors were developed for the estimation of both GSH and GSSG. Two different types of transducers were used: i) screen-printed three-electrode disposable sensor (SPE) containing carbon working electrode, carbon counter electrode and silver/silver chloride reference electrode; ii) three-electrode disposable system (CDE) consisting of three copper electrodes. 5,5,-dithiobis(2-nitrobenzoic acid) (DTNB) was used as detector element for estimation of total reduced thiol content. The enzyme glutathione reductase along with a co-enzyme reduced nicotinamide adenine dinucleotide phosphate was used to estimate GSSG. By combining the two methods GSH can also be estimated. The detector elements were immobilized on the working electrodes of the sensors by bulk polymerization of acrylamide. The responses were observed amperometrically. The detection limit for thiol (GSH) was less than 0.6,ppm when DTNB was used, whereas for GSSG it was less than 0.1,ppm. [source] The pentose-phosphate pathway in neuronal survival against nitrosative stressIUBMB LIFE, Issue 1 2010Juan P. Bolaños Abstract Neurons are thought to be particularly vulnerable cells against reactive oxygen and nitrogen species (RONS) damage (nitrosative stress), due in part to their weak antioxidant defense and low ability to compensate energy homeostasis. Intriguingly, nitrosative stress efficiently stimulates the rate of the antioxidant pentose-phosphate pathway (PPP), which generates NADPH a necessary cofactor for the reduction of glutathione disulfide. In fact, inhibition of PPP sensitizes cultured neurons to glutathione oxidation and apoptotic death, whereas its stimulation confers resistance to nitrosative stress. Furthermore, we recently described that neurons can preferentially use glucose through the PPP by inhibiting glycolysis, which is achieved by continuously degrading the glycolytic positive-effector protein, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 (Pfkfb3) by the action of the E3 ubiquitine ligase anaphase-promoting complex/cyclosome (APC/C)Cdh1. These results suggest that the antioxidant fragility of neurons may be compensated by the PPP at the expense of inhibiting bioenergetic glycolysis. © 2009 IUBMB IUBMB Life, 62(1):14,18, 2010 [source] Galactosamine-induced hepatotoxic effect and hepatoprotective role of a protein isolated from the herb Cajanus indicus L in vivo,JOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 1 2007Prasenjit Manna Abstract dd(+)-Galactosamine is a well-known experimental hepatotoxin. The present study was conducted to determine the protective role of a 43-kD protein isolated from the leaves of the herb Cajanus indicus L against dd(+)-galactosamine (GalN) induced liver damage in mice. Both preventive and curative effects of the protein have been investigated in the study. The protein was administered intraperitoneally at a dose of 2 mg/kg body weight for 4 days before and after GalN intoxication at a dose of 800 mg/kg body weight for 3 days. The increased activities of serum marker enzymes, alanine aminotransferase, and alkaline phosphatase because of GalN administration, were significantly reduced by the protein treatment. The protein also normalized the altered activities of antioxidant enzymes superoxide dismutase, catalase, glutathione reductase, and glutathione- S -transferase as well as the levels of cellular metabolites, reduced glutathione, glutathione disulfide, and total thiols. In addition, the enhanced hepatic lipid peroxidation because of GalN intoxication was also effectively inhibited by the protein treatment. Results suggest that GalN caused hepatic damages via oxidative insult and that the protein provided protection through its antioxidant mechanism. © 2007 Wiley Periodicals, Inc. J Biochem Mol Toxicol 21:13,23, 2007; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20154 [source] Aging induces cardiac diastolic dysfunction, oxidative stress, accumulation of advanced glycation endproducts and protein modificationAGING CELL, Issue 2 2005Shi-Yan Li Summary Evidence suggests that aging, per se, is a major risk factor for cardiac dysfunction. Oxidative modification of cardiac proteins by non-enzymatic glycation, i.e. advanced glycation endproducts (AGEs), has been implicated as a causal factor in the aging process. This study was designed to examine the role of aging on cardiomyocyte contractile function, cardiac protein oxidation and oxidative modification. Mechanical properties were evaluated in ventricular myocytes from young (2-month) and aged (24,26-month) mice using a MyoCam® system. The mechanical indices evaluated were peak shortening (PS), time-to-PS (TPS), time-to-90% relengthening (TR90) and maximal velocity of shortening/relengthening (± dL/dt). Oxidative stress and protein damage were evaluated by glutathione and glutathione disulfide (GSH/GSSG) ratio and protein carbonyl content, respectively. Activation of NAD(P)H oxidase was determined by immunoblotting. Aged myocytes displayed a larger cell cross-sectional area, prolonged TR90, and normal PS, ± dL/dt and TPS compared with young myocytes. Aged myocytes were less tolerant of high stimulus frequency (from 0.1 to 5 Hz) compared with young myocytes. Oxidative stress and protein oxidative damage were both elevated in the aging group associated with significantly enhanced p47phox but not gp91phox expression. In addition, level of cardiac AGEs was ,2.5-fold higher in aged hearts than young ones determined by AGEs-ELISA. A group of proteins with a molecular range between 50 and 75 kDa with pI of 4,7 was distinctively modified in aged heart using one- or two-dimension SDS gel electrophoresis analysis. These data demonstrate cardiac diastolic dysfunction and reduced stress tolerance in aged cardiac myocytes, which may be associated with enhanced cardiac oxidative damage, level of AGEs and protein modification by AGEs. [source] Maternal Alcohol Use During Pregnancy Causes Systemic Oxidation of the Glutathione Redox SystemALCOHOLISM, Issue 1 2010Theresa W. Gauthier Background:, Increased systemic oxidant stress contributes to a variety of maternal complications of pregnancy. Although the antioxidant glutathione (GSH) and its oxidized component glutathione disulfide (GSSG) have been demonstrated to be significantly altered in the adult alcoholic, the effects of maternal alcohol use during pregnancy on oxidant stress in the postpartum female remain under investigation. We hypothesized that maternal alcohol use would increase systemic oxidant stress in the pregnant female, evidenced by an oxidized systemic GSH redox potential. Methods:, As a subset analysis of a larger maternal language study, we evaluated the effects of alcohol consumption during pregnancy on the systemic GSH redox status of the postpartum female. Using an extensive maternal questionnaire, postpartum women where queried regarding their alcohol consumption during pregnancy. Any drinking, the occurrence of drinking >3 drinks/occasion, and heavy drinking of >5 drinks/occasion during pregnancy were noted. Using HPLC, maternal plasma samples were analyzed for GSH, oxidized GSSG and the redox potential of the GSH/GSSG antioxidant pair calculated. Results:, Maternal alcohol use occurred in 25% (83/321) of our study sample. Two in ten women reported consuming >3 drinks/occasion during pregnancy, while 1 in 10 women reported consuming alcohol at >5 drinks/occasion. Any alcohol use during pregnancy significantly decreased plasma GSH (p < 0.05), while alcohol at >3 drinks/occasion or >5 drinks/occasion significantly decreased plasma GSH concentration (p < 0.05), increased the percent of oxidized GSSG (p < 0.05), and substantially oxidized the plasma GSH redox potential (p < 0.05). Conclusions:, Alcohol use during pregnancy, particularly at levels >3 drinks/occasion, caused significant oxidation of the systemic GSH system in the postpartum women. The clinical ramifications of the observed alcohol-induced oxidation of the GSH redox system on high risk pregnancies or on the exposed offspring require more accurate identification and further investigation. [source] Carbon Monoxide has Antioxidative Properties in the Liver Involving p38 MAP Kinase Pathway in a Murine Model of Systemic InflammationMICROCIRCULATION, Issue 7 2010JÜRGEN BRUGGER Please cite this paper as: Brugger, Schick, Brock, Baumann, Muellenbach, Roewer and Wunder (2010). Carbon Monoxide has Antioxidative Properties in the Liver Involving p38 MAP Kinase Pathway in a Murine Model of Systemic Inflammation. Microcirculation17(7), 504,513. Abstract Objective:, Reactive oxygen species (ROS) are important in the hepatocellular injury process during a systemic inflammation. We examined the role of carbon monoxide (CO) on the hepatic generation of ROS with in-vivo and in-vitro models of systemic inflammation. Methods:, Using a murine model of bilateral hindlimb ischemia-reperfusion (I/R) we examined the effect of CO treatment on hepatic ROS formation, oxidative status, and cell injury. Cultured HUVEC were used to investigate intracellular pathways. Results:, CO treatment reduced hepatic lipid peroxidation, re-established total hepatic glutathione and glutathione disulfide (GSH/GSSG) levels and reduced hepatocellular injury. Inhibition of heme oxygenase (HO) during treatment with CO during hindlimb I/R failed to alter the antioxidant qualities provided by CO. The production of ROS after tumor necrosis factor-, (TNF-,) stimulation in HUVEC was diminished after exposure to CO. Treatment with CO during HO inhibition reduced both ROS formation and cell injury. Inhibiting the p38 MAPK (mitogen-activated protein kinase) pathway with pyridinyl imidazol (SB203580) revealed that the antioxidant potential of CO involved the activation of p38 MAPK. Conclusions:, CO has direct antioxidant potential independently of any HO activity during systemic inflammation. The antioxidant effects afforded by CO involve the activation of the p38 MAPK pathway. [source] Glutathione and adenosine triphosphate content of in vivo and in vitro matured porcine oocytesMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 4 2003A.M. Brad Abstract Glutathione (GSH) content in mature porcine oocytes is correlated with subsequent fertilization and developmental success. Adenosine triphosphate (ATP) is an important energy source for maintaining cellular activities and protein synthesis. The objective of this study was to compare GSH and ATP concentrations of in vivo and in vitro matured porcine oocytes. Ovulated, in vivo matured oocytes were frozen at ,80°C in groups of 10,20 (GSH) or 5,10 (ATP). In vitro oocytes were matured in either tissue culture medium-199 (TCM199) supplemented with polyvinyl alcohol (PVA) or hyaluronic acid (MAP5), or North Carolina State University-23 (NCSU23) supplemented with porcine follicular fluid (pFF) and frozen as described, or fertilized and cultured. GSH content was determined by the dithionitrobenzoic acid,glutathione disulfide (DTNB,GSSG) reductase recycling assay. ATP content was determined by using the Bioluminescent Somatic Cell Assay Kit. Oocytes matured in vitro in defined TCM199 with PVA or hyaluronic acid, or NCSU23 with pFF had significantly lower concentrations (P,<,0.05) of GSH (n,=,207, 9.82,±,0.71 pmol/oocyte; n,=,104, 9.73,±, 0.81 pmol/oocyte; n,=,108, 7.89,±,0.66 pmol/oocyte, respectively) compared to in vivo matured oocytes (n,=,217, 36.26,±,11.00 pmol/oocyte). Concentrations of ATP were not different between treatments (in vivo, n,=,70, 0.97,±,0.07 pmol/oocyte; TCM,PVA, n,=,117, 0.81,±,0.13 pmol/oocyte; TCM,MAP, n,=,107, 1.02,±,0.18 pmol/oocyte; NCSU,pFF, n,=,134, 0.71,±,0.08 pmol/oocyte). Intracellular ATP content does not appear to be related to developmental potential in porcine oocytes. Low intracellular GSH may be responsible, in part, for lower developmental competence observed in in vitro matured porcine oocytes. Mol. Reprod. Dev. 64: 492,498, 2003. © 2003 Wiley-Liss, Inc. [source] Determination of cellular redox status by stable isotope dilution liquid chromatography/mass spectrometry analysis of glutathione and glutathione disulfideRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 4 2008Peijuan Zhu Oxidation of glutathione (GSH) to glutathione disulfide (GSSG) occurs during cellular oxidative stress. The redox potential of the 2GSH/GSSG couple, which is determined by the Nernst equation, provides a means to assess cellular redox status. It is difficult to accurately quantify GSH and GSSG due to the ease with which GSH is oxidized to GSSG during sample preparation. To overcome this problem, a stable isotope dilution liquid chromatography/multiple reaction monitoring mass spectrometry (LC/MRM-MS) method has been developed using 4-fluoro-7-sulfamoylbenzofurazan (ABD-F) derivatization. ABD-F derivatization of the GSH thiol group was rapid, quantitative, and occurred at room temperature. The LC/MRM-MS method, which requires no sample clean-up, was validated within the calibration ranges of 5 to 400,nmol/mL in cell lysates for GSH and 0.5 to 40,nmol/mL in cell lysates for GSSG. Calibration curves prepared by adding known concentrations of GSH and GSSG to cell lysates were parallel to the standard curve prepared in buffers. GSH and GSSG concentrations were determined in two monocyte/macrophage RAW 267.4 cell lines with or without 15-LOX-1 expression (R15LO and RMock cells, respectively) after treatment with the bifunctional electrophile 4-oxo-2(E)-nonenal (ONE). R15LO cells synthesized much higher concentrations of the lipid hydroperoxide, 15(S)-hydroperoxyeicosatetraenoic acid (15-HPETE), which undergoes homolytic decomposition to ONE. GSH was depleted by ONE treatment in both RMock and R15LO cells, leading to significant increases in their redox potentials. However, R15LO cells had higher GSH concentrations (most likely through increased GSH biosynthesis) and had increased resistance to ONE-mediated GSH depletion than RMock cells. Consequently, R15LO cells had lower reduction potentials at all concentrations of ONE. GSSG concentrations were higher in R15LO cells after ONE treatment when compared with the ONE-treated RMock cells. This suggests that increased expression of 15(S)-HPETE modulates the activity of cellular GSH reductases or the transporters involved in removal of GSSG. Copyright © 2008 John Wiley & Sons, Ltd. [source] Glutathione transport in the endo/sarcoplasmic reticulum,BIOFACTORS, Issue 1-4 2003Miklós Csala Glutathione transport through the endo/sarcoplasmic reticulum (ER/SR) membrane might play a role in the maintenance of the thiol redox potential difference between the lumen and the cytosol. The transport of glutathione (both GSH and glutathione disulfide, GSSG) is entirely different in the ER and SR membranes. The transport measurements based on either rapid filtration or light scattering techniques revealed that the SR membrane transports glutathione much faster than the hepatic ER membrane or microsomal membranes prepared from heart or brain. The fastest transport has been measured in the membrane of muscle terminal cisternae, which is enriched in ryanodine receptor type 1 (RyR1). All the studied membranes have been found to be equally impermeable to various hydrophilic substances of similar size to glutathione, thus the glutathione transport in muscle microsomes and terminal cysternae as well as the correlation between the rate of glutathione transport and the abundance of RyR1 are specific. In both muscle microsomes and terminal cysternae, glutathione influx can be either inhibited or activated by antagonists and agonists of the ryanodine receptor, respectively, while these agents do not influence the transport of other small permeant molecules. These findings strongly suggest that the ryanodine receptor channel activity is directly associated with glutathione transport activity in the skeletal muscle sarcoplasmic reticulum membrane. [source] Determination of glutathione disulfide levels in biological samples using thiol,disulfide exchanging agent, dithiothreitolBIOMEDICAL CHROMATOGRAPHY, Issue 2 2009Burhan Ates Abstract A reverse-phase HPLC method incorporating dithiothreitol (DTT) reduction for quantitative determination of oxidized glutathione (GSSG) in biological samples is described here. This method is based on our previous enzymatic reduction technique that uses N-1-(pyrenyl) maleimide (NPM) as a derivatizing agent. In our earlier method, glutathione disulfide (GSSG) was measured by first reducing it to GSH with glutathione reductase (GR) in the presence of NADPH. However, this is a very costly and time-consuming technique. The method described here employs a common and inexpensive thiol,disulfide exchanging agent, DTT, for reduction of GSSG to GSH, followed by derivatization with NPM. The calibration curves are linear over a concentration range of 25,1250 nm (r2 > 0.995). The coefficients of variations for intra-run precision and inter-run precision range from 0.49 to 5.10% with an accuracy range of 1.78,6.15%. The percentage of relative recovery ranges from 97.3 to 103.2%. This new method provides a simple, efficient, and cost-effective way of determining glutathione disulfide levels with a 2.5 nm limit of detection per 5 µL injection volume. Copyright © 2008 John Wiley & Sons, Ltd. [source] MRP1/GS-X pump ATPase expression: is this the explanation for the cytoprotection of the heart against oxidative stress-induced redox imbalance in comparison to skeletal muscle cells?CELL BIOCHEMISTRY AND FUNCTION, Issue 1 2007Maurício S. Krause Abstract Striated muscle activity is always accompanied by oxidative stress (OxStress): the more intense muscle work and/or its duration, the more a redox imbalance may be attained. In spite of cardiac muscle functioning continuously, it is well known that the heart does not suffer from OxStress-induced damage over a broad physiological range. Although the expression of antioxidant enzymes may be of importance in defending heart muscle against OxStress, a series of combined antioxidant therapeutic approaches have proved to be mostly ineffective in avoiding cellular injury. Hence, additional mechanisms may be involved in heart cytoprotection other than antioxidant enzyme activities. The strong cardiotoxic effect of doxorubicin-induced cancer chemotherapy shed light on the possible role for multidrug resistance-associated proteins (MRP) in this context. Muscle activity-induced ,physiological' OxStress enhances the production of glutathione disulfide (GSSG) thus increasing the ratio of GSSG to glutathione (GSH) content inside the cells, which, in turn, leads to redox imbalance. Since MRP1 gene product (a GS-X pump ATPase) is a physiological GSSG transporter, adult Wistar rats were tested for MRP1 expression and activity in the heart and skeletal muscle (gastrocnemius), in as much as the latter is known to be extremely sensitive to muscle activity-induced OxS. MRP1 expression was completely absent in skeletal muscle. In contrast, the heart showed an exercise training-dependent induction of MRP1 protein expression which was further augmented (2.4-fold) as trained rats were challenged with a session of acute exercise. On the other hand, inducible expression of the 70-kDa heat shock protein (HSP70), a universal marker of cellular stress, was completely absent in the heart of sedentary and acutely exercised rats, whereas skeletal muscle showed a conspicuous exercise-dependent HSP70 expression, which decreased by 45% with exercise training. This effect was paralleled by a 58% decrease in GSH content in skeletal muscle which was even higher (an 80%-fall) after training thus leading to a marked redox imbalance ([GSSG]/[GSH] raised up to 38-fold). In the heart, GSH contents and [GSSG]/[GSH] ratio remained virtually unchanged even after exercise challenges, while GS-X pump activity was found to be 20% higher in the heart related to skeletal muscle. These findings suggest that an intrinsic higher capacity to express the MRP1/GS-X pump may dictate the redox status in the heart muscle thus protecting myocardium by preventing GSSG accumulation in cardiomyocytes as compared to skeletal muscle fibres. Copyright © 2006 John Wiley & Sons, Ltd. [source] | |||||||||||||||||||||||||