Cellular glutathione and the response of adult rat heart myocytes to oxidant stress

Ap, Timerman; Ra, Altschuld; Cm, Hohl; Gp, Brierley; Aj, Merola

doi:10.1016/0022-2828(90)90958-5

Cited by 36 publications

(7 citation statements)

References 43 publications

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“…To address these objectives, a cellular model of H,O,-induced cardiomyocyte oxidative stress has been defined. Neonatal-rat cardiomyocytes in primary monolayer culture were selected as the study object, for these cultures constitute a homogeneous population of viable, beating cells representative of the basic unit of myocardial structure and function, allowing changes in cardiomyocyte status to be quantified directly and reliably (Janero et al, 1988;Timerman et al, 1990). Reagent H,Oz was employed to enable precise definition and titration of the cardiomyocyte oxidative insult and to avoid the ambiguities inherent with the use of radical "scavengers" to help identify the H20, component of a chemically heterogenous oxidative insult (VerDonck et al, 1988).…”

Section: Discussionmentioning

confidence: 99%

“…Cardiomyocyte GSH was quantified in HC10, extracts by a technique modified from Timerman et al (1990) with the anion-exchange HPLC procedure of Reed et al (1980). Briefly, cardiomyocyte monolayers were rinsed with 15.0 ml PBS, scraped into 0.7 ml ice-cold PBS, and centrifuged for 1.0 min a t 4°C in a Beckman microfuge B through 0.35 ml dibutylphthalate and into 0.25 ml 10.0% (wiv) HC10, containing 25.0 pM a-glutamyl glutamic acid internal standard and 1.0 mM EDTA.…”

Section: Gsh Quantificationmentioning

confidence: 99%

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Hydrogen peroxide‐induced oxidative stress to the mammalian heart‐muscle cell (cardiomyocyte): Lethal peroxidative membrane injury

Janero¹,

Hreniuk²,

Sharif³

1991

Journal Cellular Physiology

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Oxidative stress induced by hydrogen peroxide (H2O2) may contribute to the pathogenesis of ischemic-reperfusion injury in the heart. For the purpose of investigating directly the injury potential of H2O2 on heart muscle, a cellular model of H2O2-induced myocardial oxidative stress was developed. This model employed primary monolayer cultures of intact, beating neonatal-rat cardiomyocytes and discrete concentrations of reagent H2O2 in defined, supplement-free culture medium. Cardiomyocytes challenged with H2O2 readily metabolized it such that the culture content of H2O2 diminished over time, but was not depleted. The consequent H2O2-induced oxidative stress caused lethal sarcolemmal disruption (as measured by lactate dehydrogenase release), and cardiomyocyte integrity could be preserved by catalase. During oxidative stress, a spectrum of cellular derangements developed, including membrane phospholipid peroxidation, thiol oxidation, consumption of the major chain-breaking membrane antiperoxidant (alpha-tocopherol), and ATP loss. No net change in the protein or phospholipid contents of cardiomyocyte membranes accompanied H2O2-induced oxidative stress, but an increased turnover of these membrane constituents occurred in response to H2O2. Development of lethal cardiomyocyte injury during H2O2-induced oxidative stress did not require the presence of H2O2 itself; a brief "pulse" exposure of the cardiomyocytes to H2O2 was sufficient to incite the pathogenic mechanism leading to cell disruption. Cardiomyocyte disruption was dependent upon an intracellular source of redox-active iron and the iron-dependent transformation of internalized H2O2 into products (e.g., the hydroxyl radical) capable of initiating lipid peroxidation, since iron chelators and hydroxyl-radical scavengers were cytoprotective. The accelerated turnover of cardiomyocyte-membrane protein and phospholipid was inhibited by antiperoxidants, suggesting that the turnover reflected molecular repair of oxidized membrane constitutents. Likewise, the consumption of alpha-tocopherol and the oxidation of cellular thiols appeared to be epiphenomena of peroxidation. Antiperoxidant interventions coordinately abolished both H2O2-induced lipid peroxidation and sarcolemmal disruption, demonstrating that an intimate pathogenic relationship exists between sarcolemmal peroxidation and lethal compromise of cardiomyocyte integrity in response to H2O2-induced oxidative stress. Although sarcolemmal peroxidation was causally related to cardiomyocyte disruption during H2O2-induced oxidative stress, a nonperoxidative route of H2O2 cytotoxicity was also identified, which was expressed in the complete absence of cardiomyocyte-membrane peroxidation. The latter mode of H2O2-induced cardiomyocyte injury involved ATP loss such that membrane peroxidation and cardiomyocyte disruption on the one hand and cellular de-energization on the other could be completely dissociated.(ABSTRACT TRUNCATED AT 400 WORDS)

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Section: Discussionmentioning

confidence: 99%

Section: Gsh Quantificationmentioning

confidence: 99%

Hydrogen peroxide‐induced oxidative stress to the mammalian heart‐muscle cell (cardiomyocyte): Lethal peroxidative membrane injury

Janero¹,

Hreniuk²,

Sharif³

1991

Journal Cellular Physiology

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“…Both the activity of glutathione peroxidase and radicals induce GSH oxidation. All studies in hepatocytes (Jewell et al, 1986;Shertzer et al, 1994;Fernandes et al, 1995) or cardiomyocytes (Timerman et al, 1990) with t-BHP report depletion of GSH and GSSG production. Thus, the observed GSH oxidation and the increased GSSG/GSH balance are in accordance with the literature and reflect the t-BHP oxidative action.…”

Section: Discussionmentioning

confidence: 99%

“…Oxidative stress induced by t-BHP has been widely studied in isolated hepatocytes (Eklow et al, 1984;Jewell et al, 1986;Joyeux et al, 1990;Shertzer et al, 1994;Fernandes et al, 1995) as well as in isolated cardiomyocytes (Timerman et al, 1990;Daly et al, 1991;Vlessis et al, 1991;Castro and Bhatnagar, 1993;Andersson et al, 1996). Loss of cell viability, increase of intracellular free Ca 2 , nucleotide degradation, GSH oxidation and lipid peroxidation are the highlights of t-BHP effect.…”

Section: Introductionmentioning

confidence: 99%

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

Remião

Carmo

Carvalho

et al. 2000

Biomed. Chromatogr.

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Glutathione and glutathione disulphide constitute an essential thiol redox system present in the cell. The balance in favour of the latter is an indication of oxidative stress. Glutathione and glutathione disulphide quantification in isolated cells may therefore be essential for the evaluation of mechanistic and comparative studies of toxic xenobiotics. In this study, a rapid and sensitive isocratic reverse-phase high-performance liquid chromatographic method using coulometric detection was implemented for the simultaneous detection of glutathione and glutathione disulphide, in freshly isolated hepatocytes and cardiomyocytes of the rat. The method implemented proved to be effective for the measurement of glutathione and glutathione disulphide in control conditions and for the detection of variations in this redox system, induced by tert-butylhydroperoxide. tert-Butylhydroperoxide is an organic peroxide, which has been used as a model molecule for inducing oxidative stress in isolated cells. A comparative study with a previously published HPLC-electrochemical detection method was performed.

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“…tert-Butylhydroperoxide (t-BHP) is an organic peroxide, which is similar to short-chain lipidic peroxides produced as a result of normal cellular metabolism or oxidative injury elicited by xenobiotics. Oxidative stress induced by t-BHP has been widely studied in isolated hepatocytes (Eklow et al, 1984;Jewell et al, 1986;Joyeux et al, 1990;Shertzer et al, 1994;Fernandes et al, 1995) as well as in isolated cardiomyocytes (Timerman et al, 1990;Daly et al, 1991;Vlessis et al, 1991;Castro and Bhatnagar, 1993;Andersson et al, 1996). Loss of cell viability, increase of intracellular free Ca 2 , nucleotide degradation, GSH oxidation and lipid peroxidation are the highlights of t-BHP effect.…”

Section: Introductionmentioning

confidence: 99%

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

Remião

Carmo

Carvalho

et al. 2000

Biomed. Chromatogr.

View full text Add to dashboard Cite

show abstract

Cellular glutathione and the response of adult rat heart myocytes to oxidant stress

Cited by 36 publications

References 43 publications

Hydrogen peroxide‐induced oxidative stress to the mammalian heart‐muscle cell (cardiomyocyte): Lethal peroxidative membrane injury

Hydrogen peroxide‐induced oxidative stress to the mammalian heart‐muscle cell (cardiomyocyte): Lethal peroxidative membrane injury

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

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

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