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Intracellular GSH (intracellular + gsh)
Terms modified by Intracellular GSH Selected AbstractsCadmium-induced astroglial death proceeds via glutathione depletionJOURNAL OF NEUROSCIENCE RESEARCH, Issue 2 2006Joo-Young Im Abstract Cadmium is a heavy metal that accumulates in the body, and its accumulation in the brain damages both neurons and glial cells. In the current study, we explored the mechanism underlying cadmium toxicity in primary cortical astroglia cultures. Chronic treatment with 10 ,M cadmium was sufficient to cause 90% cell death in 18 hr. However, unlike that observed in neurons, cadmium-induced astroglial toxicity was not attenuated by the antioxidants trolox (100 ,M), caffeic acid (1 mM), and vitamin C (1 mM). In contrast, extracellular 100 ,M glutathione (GSH; ,-Glu-Cys-Gly) or 100 ,M cysteine almost completely blocked cadmium-induced astroglial death, whereas 300 ,M oxidized GSH (GSSG) or 300 ,M cystine, which do not have the free thiol group, were ineffective. In addition, cadmium toxicity was noticeably inhibited or enhanced when intracellular GSH was, respectively, increased by using the cell-permeable glutathione ethyl ester (GSH-EE) or depleted by using buthionine sulfoximine (BSO), an inhibitor of ,-glutamylcysteine synthetase. In agreement with these data, intracellular GSH levels were found to be depressed in cadmium-treated astrocytes. These results suggest that the toxic effect of cadmium on primary astroglial cells involves GSH depletion and, furthermore, that GSH administration can potentially be used to counteract cadmium-induced astroglial cell death therapeutically. © 2005 Wiley-Liss, Inc. [source] Enhanced glutathione production by using low-pH stress coupled with cysteine addition in the treatment of high cell density culture of Candida utilisLETTERS IN APPLIED MICROBIOLOGY, Issue 5 2008G. Liang Abstract Aims:, To investigate the effects of pH stress coupled with cysteine addition on glutathione (GSH) production in the treatment of high cell density culture of Candida utilis. Methods and Results:, We have previously observed that most Candida utilis cells remained viable after being subjected to pH at 1·2 for 3 h and that some intracellular GSH leaked into the medium. A cysteine addition strategy was applied in fed-batch production of GSH. A single cysteine addition resulted in higher GSH yield than two separate additions without pH stress. An increase in intracellular GSH content triggered inhibition of ,-glutamylcysteine synthetase (,-GCS). A strategy that combines cysteine addition with low-pH stress was developed to relieve the inhibition of ,-GCS. Conclusion:, Without pH stress, single shot and double shot cysteine addition yielded a total GSH of 1423 and 1325 mg l,1. In comparison, a low-pH stress counterpart resulted in a total GSH of 1542 and 1730 mg l,1, respectively. With low-pH stress, we observed GSH secretion into the medium at 673 and 558 mg l,1 and an increase in the ,-GCS activity by 1·2- and 1·5-fold, respectively. The specific GSH production yield increased from 1·76% to 1·91% (w/w) for single shot, and 1·64% to 2·14% for double shots. Significance and Impact of the Study:, Low-pH shift was applied to alleviate the feedback inhibition of intracellular GSH on ,-GCS activity by secreting GSH into the medium. This strategy is coupled with cysteine addition to enhance GSH production in Candida utilis. [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] Azathioprine hepatotoxicity and the protective effect of liquorice and glycyrrhizic acidPHYTOTHERAPY RESEARCH, Issue 8 2006Yue-Ting Wu Abstract This study aimed to evaluate the responses of human hepatocytes to azathioprine hepatotoxicity in comparison with the well-studied azathioprine hepatotoxicity in rat hepatocytes and the effects of protective agents to suppress azathioprine hepatotoxicity. Azathioprine presented its hepatotoxicity at clinically relevant concentrations (lower than 10 µm) in primary rat hepatocytes after 48 h of treatment as shown by a severe decrease in cell viability as well as intracellular GSH depletion. However, primary human hepatocytes exhibited only significant intracellular GSH depletion after treatment with azathioprine at these clinically relevant concentrations, while a reduction in cell viability by 29% was only evidenced after 48 h of treatment with azathioprine at the high concentration of 50 µm. In addition, a monolayer culture of primary rat hepatocytes was used as an in vitro model to examine the protective effects of antihepatotoxic drugs including glutathione (GSH), N-acetylcysteine (NAC, a GSH precursor), liquorice and glycyrrhizic acid (GA), a major bioactive component of liquorice, against hepatotoxicity of 1 µm azathioprine. It was found that both liquorice and GA showed substantial protection according to assays of cell viability and intracellular GSH, while neither GSH nor NAC had such a protective function. Similarly, GA protected human hepatocytes from intracellular GSH depletion on exposure to 1 µm azathioprine. These results implied that GA or liquorice could be considered as potent protection agents against azathioprine hepatotoxicity. Copyright © 2006 John Wiley & Sons, Ltd. [source] Intracellular glutathione in stretch-induced cytokine release from alveolar type-2 like cellsRESPIROLOGY, Issue 1 2004Behrouz Jafari Objective: Ventilator-induced lung injury (VILI) is characterized by release of inflammatory cytokines, but the mechanisms are not well understood. We hypothesized that stretch-induced cytokine production is dependent on oxidant release and is regulated by intracellular glutathione (GSH) inhibition of nuclear factor ,B (NF-,B) and activator protein-1 (AP-1) binding. Methodology: Type 2-like alveolar epithelial cells (A549) were exposed to cyclic stretch at 15% strain for 4 h at 20 cycles/min with or without N-acetylcysteine (NAC) or glutathione monoethylester (GSH-e) to increase intracellular GSH, or buthionine sulfoximine (BSO), to deplete intracellular GSH. Results: Cyclic stretch initially caused a decline in intracellular GSH and a rise in the levels of isoprostane, a marker of oxidant injury. This was followed by a significant increase in intracellular GSH and a decrease in isoprostane. Stretch-induced IL-8 and IL-6 production were significantly inhibited when intracellular GSH was further increased by NAC or GSH-e (P < 0.0001). Stretch-induced IL-8 and IL-6 production were augmented when intracellular GSH was depleted by BSO (P < 0.0001). NAC blocked stretch-induced NF-,B and AP-1 binding and inhibited IL-8 mRNA expression. Conclusions: We conclude that oxidant release may play a role in lung cell stretch-induced cytokine release, and antioxidants, which increase intracellular GSH, may protect lung cells against stretch-induced injury. [source] Redox regulation of ascorbic acid transport: Role of transporter and intracellular sulfhydrylsBIOFACTORS, Issue 4 2004James M. May Abstract Ascorbic acid is one of the most sensitive cellular defenses against oxidant damage. However, it requires a sodium- and energy-dependent transporter to enter cells against a concentration gradient. To test the hypothesis that ascorbate transport is sensitive to redox stress, we studied changes in transport of the vitamin in response to sulfhydryl modification of the protein and to GSH depletion in cultured endothelial cells. Transport of ascorbic acid, measured as the uptake of radiolabeled ascorbate, was inhibited by the membrane-impermeant sulfhydryl reagents thorin, p -chloromercuribenzene sulfonic acid, and 5,5,-dithiobis-(2-nitrobenzoic acid) in a dose-dependent manner without significant depletion of intracellular GSH. Sulfhydryl reagents capable of penetrating the plasma membrane, including phenylarsine oxide, p -chloromercuribenzoic acid, and N-ethylmaleimide, inhibited transport and lowered cellular GSH. Diamide, which induces disulfide formation, increased ascorbate transport over a narrow concentration range under conditions in which GSH was not depleted. On the other hand, specific depletion of intracellular GSH by several different mechanisms did inhibit transport. Together, these results suggest that the ascorbate transporter is sensitive to redox modulation. This relates in part to sulfhydryl groups exposed on the exofacial ascorbate transporter, and to sulfhydryl groups that are sensitive to changes in the redox state of intracellular GSH. [source] Separation and quantification of N -acetyl- l -cysteine and N -acetyl-cysteine-amide by HPLC with fluorescence detectionBIOMEDICAL CHROMATOGRAPHY, Issue 5 2006Wei Wu Abstract N- acetyl- l -cysteine (NAC) is a well-known antioxidant that is capable of facilitating glutathione (GSH) biosynthesis and replenishing intracellular GSH under oxidatively challenging circumstances. N- acetyl-cysteine-amide (NACA), the amide form of NAC, is a newly designed and synthesized thiol-containing compound which is believed to be more lipophilic and permeable through cell membranes than NAC. The metabolic and antioxidant effects of these compounds in vitro and in vivo are under investigation. However, an analytical method that can separate and quantify both compounds simultaneously is not yet available, to the best of our knowledge. Because of their structural similarities, the two compounds are difficult to separate using earlier HPLC methods which were designed for NAC quantification. Therefore, the goal of this work was to develop an HPLC method with fluorescence detection for simultaneous quantification of NAC and NACA in biological blood and tissue samples. A gradient HPLC program with fluorescence detection (,ex = 330 nm, ,em = 376 nm) using N -(1-pyrenyl)maleimide (NPM) as the derivatizing agent was developed. The calibration curves were linear over a concentration range of 25,5000 nm (r2 > 0.997). The coefficients of variation for within-run precision and between-run precision ranged from 0.67 to 5.23% and for accuracy ranged from 0.98 to 10.54%; the percentage relative recovery ranged from 94.5 to 102.8%. This new method provides satisfactory separation of NAC and NACA, along with other biological thiols, in 20 min with a 5 nm limit of detection (LOD) per 5 µL injection volume. Copyright © 2005 John Wiley & Sons, Ltd. [source] | |||||||||||||