The role of glutathione reductase in maintaining human granulocyte function and sensitivity to exogenous H2O2

Cohen, Harvey J.; Tape, Elizabeth H.; Novak, John; Chovaniec, Margaret E.; Liegey, P; Whitin, John C.

doi:10.1182/blood.v69.2.493.493

Cited by 27 publications

(1 citation statement)

References 14 publications

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“…Since depletion of intracellular GSH impairs neutro-phi1 functions such as phagocytosis and bactericidal activity [ 16,17,20], impaired transsulfuration and an impaired ability to re-synthesize the GSH in certain disease states such as cirrhosis [34,35] may be an important determinant of PMN function and could at least in part account for the impaired antimicrobial defence in some patients with liver disease [36]. The fate of GSH following activation of PMN is difficult to assess since intracellular GSH disappeared and neither GSH nor GSSG were recovered in the medium of activated PMN.…”

Section: Discussionmentioning

confidence: 99%

Glutathione metabolism in activated human neutrophils: stimulation of glutathione synthesis and consumption of glutathione by reactive oxygen species

Bilzer

Lauterburg

1991

Eur J Clin Investigation

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Since glutathione (GSH) is involved in the modulation of the function of polymorphonuclear leucocytes (PMN) such as phagocytosis and production of reactive oxygen species, the metabolism of GSH was studied in human PMN. The concentration of GSH in resting PMN amounted to 13.3 nmol 10(-7) PMN and remained stable over 100 min of incubation. Upon activation of PMN with phorbol myristate acetate intracellular GSH decreased to 50% of the resting concentration within 80 min. In the presence of buthionine sulfoximine, which inhibits the synthesis of GSH, the depletion of intracellular GSH was dramatically accelerated, indicating that activation of PMN is associated with a marked stimulation of GSH synthesis. Since a similar depletion of GSH was seen in the presence of propargylglycine, an inhibitor of the cystathionine pathway, most of the cysteine required for the resynthesis of GSH must originate from methionine and not from cysteine generated by the catabolism of GSH. Further studies showed that GSH is sequentially oxidized by O2-. and HOCl, first to GSSG and then to an unidentified compound, most likely a chloramine. In the presence of an adequate supply of GSH and NADPH which is required for the reduction of GSSG by glutathione reductase this further oxidation of GSSG was prevented. Thus, the highly toxic HOCl generated by PMN can be detoxified by the glutathione reducatase system. The capacity of PMN to re-synthesize GSH may be an important determinant of PMN function.

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Section: Discussionmentioning

confidence: 99%

Glutathione metabolism in activated human neutrophils: stimulation of glutathione synthesis and consumption of glutathione by reactive oxygen species

Bilzer

Lauterburg

1991

Eur J Clin Investigation

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Flow Cytometric Analysis of Respiratory Burst Activity in Phagocytes With Hydroethidine and 2′,7′-Dichlorofluorescin

Rothe

Valet

1990

Journal of Leukocyte Biology

810

484

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Hydroethidine (HE) and 2',7'-dichlorofluorescin (DCFH) were used for the flow cytometric measurement of reactive oxygen metabolites in leukocytes. Hydroethidine and DCFH were both rapidly oxidized in a cell-free cuvette assay to ethidium bromide (EB) and 2',7'-dichlorofluorescein (DCF) by H2O2 and peroxidase, but not by H2O2 alone, while only HE was oxidized by KO2, a source of O2-. Quiescent lymphocytes, monocytes, and neutrophils spontaneously oxidized HE to EB, while DCFH was only oxidized to a low degree. Neutrophils increased 6.9-fold in EB red fluorescence and 12.5-fold in DCF green fluorescence during the respiratory burst induced by phorbol 12-myristate 13-acetate or 6.1-fold and 4.7-fold, respectively, during the respiratory burst induced by Escherichia coli bacteria. The HE or DCFH oxidation during the respiratory burst, unlike the spontaneous HE oxidation, was not inhibitable by 10 mM NaNe indicating a non-mitochondrial source of cellular oxidants during the respiratory burst such as NADPH oxidase, which produces O2-. The oxidation of DCFH, but not of HE, was decreased in stimulated neutrophils, which were simultaneously loaded with HE and DCFH. Intracellular DCFH oxidation induced by incubation of resting neutrophils with extracellular H2O2 was not influenced by the presence of HE. This indicates that HE is oxidized at an earlier step in the reactive oxygen metabolism of neutrophils than DCFH, i.e., by early oxygen metabolites like O2-, while DCFH is oxidized in part by H2O2 and phagosomal peroxidases. The differential oxidation of HE and DCFH during simultaneous cellular staining permits the analysis of up to three functionally different neutrophil populations in septic patients. This is of interest for the determination of disease-related alterations of oxygen metabolism in quiescent and stimulated leukocytes.

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Diamide primes neutrophils for enhanced release of superoxide anion: relationship to S-thiolation of cellular proteins

Moriguchi

Seres

Ravichandran

et al. 1996

Journal of Leukocyte Biology

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Stimulation of the respiratory burst in phagocytes induces the formation of mixed disulfides between sulfhydryl groups of proteins and low-molecular-weight thiols. We hypothesized that this process (S-thiolation) might be involved in turning off the respiratory burst. However, induction of S-thiolation by pretreatment of neutrophils with diamide, a direct thiol oxidizing agent, actually primed the cells for a two- to fivefold increase in total release and fourfold increase in rate of release of 02- on stimulation by f-Met-Leu-Phe. Generation of intracellular oxidants (hydroethidine fluorescence) was increased ninefold. Priming and S-thiolation were apparent at 1 min of incubation and peaked at 5-10 min. Diamide pretreatment also reduced the lag time between addition of phorbol diester and release of 02- by a mean of 23 s (41%). Dithioerythritol, a sulfhydryl-reducing agent, abolished both the S-thiolation and priming mediated by diamide. H202 also induced priming and S-thiolation; and these were eliminated by dithioerythritol. In contrast to the effect of endotoxin, diamide priming did not affect Ca2+ homeostasis of the neutrophils. Diamide did not significantly alter NADPH oxidase activity in a cell-free system. These findings suggest that sulfhydryl groups on one or more proteins play an important role in modulating the respiratory burst.

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The role of glutathione reductase in maintaining human granulocyte function and sensitivity to exogenous H2O2

Cited by 27 publications

References 14 publications

Glutathione metabolism in activated human neutrophils: stimulation of glutathione synthesis and consumption of glutathione by reactive oxygen species

Glutathione metabolism in activated human neutrophils: stimulation of glutathione synthesis and consumption of glutathione by reactive oxygen species

Flow Cytometric Analysis of Respiratory Burst Activity in Phagocytes With Hydroethidine and 2′,7′-Dichlorofluorescin

Diamide primes neutrophils for enhanced release of superoxide anion: relationship to S-thiolation of cellular proteins

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