Redox thiol status plays a central role in the mobilization and metabolism of nitric oxide in human red blood cells

Authors

  • José Pedro de Almeida,

    Corresponding author
    1. Santa Maria Hospital, University of Lisbon Medical School, Av Prof. Egas Moniz, 1649-028 Lisbon, Portugal
    2. Institute of Biochemistry, Institute of Molecular Medicine, University of Lisbon Medical School, Av Prof. Egas Moniz, 1649-028 Lisbon, Portugal
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  • Filomena Almeida Carvalho,

    1. Institute of Biochemistry, Institute of Molecular Medicine, University of Lisbon Medical School, Av Prof. Egas Moniz, 1649-028 Lisbon, Portugal
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  • Ana Santos Silva-Herdade,

    1. Institute of Biochemistry, Institute of Molecular Medicine, University of Lisbon Medical School, Av Prof. Egas Moniz, 1649-028 Lisbon, Portugal
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  • Teresa Santos-Freitas,

    1. Institute of Biochemistry, Institute of Molecular Medicine, University of Lisbon Medical School, Av Prof. Egas Moniz, 1649-028 Lisbon, Portugal
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  • Carlota Saldanha

    1. Institute of Biochemistry, Institute of Molecular Medicine, University of Lisbon Medical School, Av Prof. Egas Moniz, 1649-028 Lisbon, Portugal
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Instituto de Bioquímica, Instituto de Medicina Molecular, Faculdade de Medicina de Lisboa, Edifício Egas Moniz, Av Prof Egas Moniz, 1649-028 Lisboa, Portugal. Tel.: +351 91 8985450; fax: +351 21 7999477. E-mail addresses: jpedro.gla@gmail.com

Abstract

We assessed the redox thiol status influence on nitric oxide (NO) metabolism and efflux in erythrocytes stimulated with acetylcholinesterase substrate (acetylcholine, ACh) and inhibitor (velnacrine maleate, VM). Erythrocyte suspensions from healthy donors were incubated with increasing concentrations of dithiothreitol (1–50 μM), in the presence and absence of acetylcholine/velnacrine (10 μM). Levels of NO, nitrite/nitrate, S-nitrosohemoglobin, peroxynitrite and S-nitrosoglutathione were determined by spectrofluorimetric and spectrophotometric methods.

Dithiothreitol significantly mobilized NO toward nitrite/nitrate and S-nitrosoglutathione, and decreased the amount of NO efflux. Both ACh/VM induce changes on the levels of erythrocyte nitrite/nitrate dependent on the DTT concentration. Higher levels of peroxynitrite and S-nitrosoglutathione were seen with velnacrine in presence of DTT 1 and 50 μM.

We concluded that dithiothreitol-induced activation of erythrocyte thiol status decreases NO efflux and allows greater intracellular NO mobilization onto different derivative molecules, both in the absence and presence of acetylcholinesterase substrate and inhibitor.

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