Simultaneous determination of reduced and oxidized glutathione in freshly isolated rat hepatocytes and cardiomyocytes by HPLC with electrochemical detection

Remião, Fernando; Carmo, Helena; Carvalho, Félix; Bastos, Maria de Lourdes

doi:10.1002/1099-0801(200011)14:7<468::aid-bmc10>3.0.co;2-c

Cited by 23 publications

(5 citation statements)

References 33 publications

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“…GSH and GSSG were quantified as before (24). Briefly, perchloric acid (5% final acid concentration) was added to aliquots of cells (2.5 × 10 4 ) or supernatant (100 µL).…”

Section: Methodsmentioning

confidence: 99%

Cu²⁺-Induced Isoproterenol Oxidation into Isoprenochrome in Adult Rat Calcium-Tolerant Cardiomyocytes

Remião

Carvalho

Carmo

et al. 2002

Chem. Res. Toxicol.

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Sustained high levels of circulating catecholamines may induce cardiotoxicity. There is increasing evidence that this could result from catecholamine oxidation into aminochromes, which is catalyzed by transition metals. In fact, it has already been shown that copper-induced oxidation of the beta-agonist isoproterenol decreases the viability of isolated cardiomyocytes. Thus, the aim of this work was to contribute for the clarification of the mechanisms underlying the toxic effects of isoproterenol, Cu2+ and their concomitant effect in isolated rat cardiomyocytes. Freshly isolated calcium-tolerant cardiomyocytes from adult rat were incubated with 1 mM isoproterenol, 20 microM Cu2+ or with both during 4 h. Isoproterenol and its aminochrome (isoprenochrome), and reduced and oxidized glutathione were measured at each hour in the incubation medium and in the cells. The intracellular activities of the selenium-dependent glutathione peroxidase, glutathione reductase, and glutathione-S-transferase were determined after 4 h of incubation. Isoprenochrome was found in both cells and incubation medium in samples incubated with isoproterenol alone. However, in the isoproterenol plus Cu2+ samples, a greater depletion of isoproterenol accompanied by a proportional increase of isoprenochrome was observed. This higher ISO oxidation resulted in the depletion of intracellular glutathione and in the release of oxidized glutathione to the incubation medium. The content of total glutathione (intra- and extracellular) and the intracellular activity of the selenium-dependent glutathione peroxidase, glutathione reductase, and glutathione-S-transferase were also decreased in the isoproterenol plus Cu2+ samples. These results seem to indicate that the oxidative stress resulting from catecholamine/transition metal association may contribute to catecholamine cardiotoxicity.

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“…GSH and GSSG were quantified as before (24). Briefly, perchloric acid (5% final acid concentration) was added to aliquots of cells (2.5 × 10 4 ) or supernatant (100 µL).…”

Section: Methodsmentioning

confidence: 99%

Cu²⁺-Induced Isoproterenol Oxidation into Isoprenochrome in Adult Rat Calcium-Tolerant Cardiomyocytes

Remião

Carvalho

Carmo

et al. 2002

Chem. Res. Toxicol.

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“…GSH measurement was performed by HPLC with a dual-electrode coulometric detection (Remia˜o et al 2000). Briefly, liver samples were homogenized in 5% perchloric acid with an UltraTurrax device.…”

Section: Biochemical Analysismentioning

confidence: 99%

Effect of 3,4-methylenedioxymethamphetamine ("ecstasy") on body temperature and liver antioxidant status in mice: influence of ambient temperature

Carvalho

Remião

et al. 2002

Archives of Toxicology

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The consumption of 3,4-methylenedioxymethamphetamine (MDMA; ecstasy) is known to cause severe hyperthermia and liver damage in humans. The thermogenic response induced by MDMA is complex and partially determined by the prevailing ambient temperature (AT). This is of extreme importance since ecstasy is often consumed at "rave" parties, where dancing takes place in a warm environment, which may exacerbate the effect of MDMA on thermoregulation. In view of the fact that hyperthermia is a well-known pro-oxidant aggressive condition, its potential role in ecstasy-induced hepatocellular toxicity should be further studied. Thus, the present study was performed in order to evaluate the influence of AT on the effects of single administration of MDMA on body temperature and liver toxicity in Charles River mice. Animals were given an acute intraperitoneal dose of MDMA (5, 10 or 20 mg/kg) and placed in AT of 20+/-2 degrees C or 30+/-2 degrees C for 24 h. Body temperature was measured during the study using implanted transponders and a temperature probe reading device. Plasma and liver samples were used for biochemical analysis. Liver sections were also taken for histological examination. The parameters evaluated were (1) plasma levels of transaminases and alkaline phosphatase, (2) hepatic glutathione (GSH), (3) hepatic lipid peroxidation, (4) activity of hepatic antioxidant enzymes (catalase, glutathione peroxidase, glutathione reductase, glutathione- S-transferase, copper/zinc superoxide dismutase and manganese superoxide dismutase), and (5) liver histology. The hyperthermic response elicited by MDMA was clearly dose-related and potentiated by high AT. Administration of MDMA produced some evidence of oxidative stress, expressed as GSH depletion at both ATs studied, as well as by lipid peroxidation and decreased catalase activity at high AT. High AT, by itself, decreased glutathione peroxidase activity. Histological examination of the liver revealed abnormalities of a dose- and AT-dependent nature. These changes included vacuolation of the hepatocytes, presence of blood clots and loss of typical hepatic cord organisation. The results obtained in the present study suggest that oxidative stress plays a part in the first stage of MDMA-induced liver damage and that liver antioxidant status is aggravated by increased AT. Thus, these findings are in accordance with the hypothesis that high AT may potentiate ecstasy-induced hepatotoxicity by increasing body hyperthermia.

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“…However, the concentration of GSH and GSSH were measured in diluted human plasma, without induced oxidative stress, and without any comparison with other analytical techniques. Also, the simultaneous detection of GSH and GSSG using HPLC and electrochemical detection has been demonstrated for rat hepatocytes and cardiomyocytes . Finally, the simultaneous detection of GSH and acetaminophen has also been previously reported …”

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confidence: 89%

Simultaneous Electrochemical Speciation of Oxidized and Reduced Glutathione. Redox Profiling of Oxidative Stress in Biological Fluids with a Modified Carbon Electrode

Moya¹,

Alfaro²,

Kazemi

et al. 2017

Anal. Chem.

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The simultaneous electrochemical quantification of oxidized (GSSG) and reduced glutathione (GSH), biomarkers of oxidative stress, is demonstrated in biological fluids. The detection was accomplished by the development of a modified carbon electrode and was applied to the analysis of biological fluids of model organisms under oxidative stress caused by lead intoxication. Nanocomposite molecular material based on cobalt phthalocyanine (CoPc) and multiwalled carbon nanotubes functionalized with carboxyl groups (MWCNT) was developed to modify glassy carbon electrodes (GCE) for the detection of reduced and oxidized glutathione. The morphology of the nanocomposite film was characterized by scanning electron microscopy (SEM) and profilometry. The electrochemical behavior of the modified electrode was assessed by cyclic voltammetry (CV) to determine the surface coverage (Γ) by CoPc. The electrocatalytic behavior of the modified electrode toward reduced (GSH) and oxidized (GSSG) forms of glutathione was assessed by CV studies at physiological pH. The obtained results show that the combined use of CoPc and MWCNT results in an electrocatalytic activity for GSH oxidation and GSSG reduction, enabling the simultaneous detection of both species. Differential pulse voltammetry reveals detection limits of 100 μM for GSH and 8.3 μM for GSSG, respectively. The potential interference from ascorbic acid, cysteine, glutamic acid, and glucose was also studied, and the obtained results show limited effects from these species. Finally, the hybrid electrode was used for the determination of GSH and GSSG in rat urine and plasma samples, intoxicated or not by lead. Both glutathione forms were detected in these complex biological matrixes without any pretreatment. Our results portray the role of GSH and GSSG as markers of oxidative stress in live organisms under lead intoxication.

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Simultaneous determination of reduced and oxidized glutathione in freshly isolated rat hepatocytes and cardiomyocytes by HPLC with electrochemical detection

Cited by 23 publications

References 33 publications

Cu²⁺-Induced Isoproterenol Oxidation into Isoprenochrome in Adult Rat Calcium-Tolerant Cardiomyocytes

Cu²⁺-Induced Isoproterenol Oxidation into Isoprenochrome in Adult Rat Calcium-Tolerant Cardiomyocytes

Effect of 3,4-methylenedioxymethamphetamine ("ecstasy") on body temperature and liver antioxidant status in mice: influence of ambient temperature

Simultaneous Electrochemical Speciation of Oxidized and Reduced Glutathione. Redox Profiling of Oxidative Stress in Biological Fluids with a Modified Carbon Electrode

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Simultaneous determination of reduced and oxidized glutathione in freshly isolated rat hepatocytes and cardiomyocytes by HPLC with electrochemical detection

Cited by 23 publications

References 33 publications

Cu2+-Induced Isoproterenol Oxidation into Isoprenochrome in Adult Rat Calcium-Tolerant Cardiomyocytes

Cu2+-Induced Isoproterenol Oxidation into Isoprenochrome in Adult Rat Calcium-Tolerant Cardiomyocytes

Effect of 3,4-methylenedioxymethamphetamine ("ecstasy") on body temperature and liver antioxidant status in mice: influence of ambient temperature

Simultaneous Electrochemical Speciation of Oxidized and Reduced Glutathione. Redox Profiling of Oxidative Stress in Biological Fluids with a Modified Carbon Electrode

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Cu²⁺-Induced Isoproterenol Oxidation into Isoprenochrome in Adult Rat Calcium-Tolerant Cardiomyocytes

Cu²⁺-Induced Isoproterenol Oxidation into Isoprenochrome in Adult Rat Calcium-Tolerant Cardiomyocytes