Redox thiol status plays a central role in the mobilization and metabolism of nitric oxide in human red blood cells.
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 (1-50microM), in the presence and absence of acetylcholine/velnacrine (10microM). 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 50microM. 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.
Lopes de Almeida
José Pedro
JP
Santa Maria Hospital, University of Lisbon Medical School, Portugal. jpedro.gla@gmail.com
Carvalho
Filomena Almeida
FA
Silva-Herdade
Ana Santos
AS
Santos-Freitas
Teresa
T
Saldanha
Carlota
C
D000109:Acetylcholine / Q000494:pharmacology; D000110:Acetylcholinesterase / Q000378:metabolism; D002800:Cholinesterase Inhibitors / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology*; D004912:Erythrocytes / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D009569:Nitric Oxide / Q000378:metabolism*; D010084:Oxidation-Reduction; D030421:Peroxynitrous Acid / Q000378:metabolism; D026422:S-Nitrosoglutathione / Q000378:metabolism; D013619:Tacrine / Q000031:analogs & derivatives / Q000494:pharmacology
D002800:Cholinesterase Inhibitors; D030421:Peroxynitrous Acid; D009569:Nitric Oxide; D013619:Tacrine; D026422:S-Nitrosoglutathione; D000110:Acetylcholinesterase; D000109:Acetylcholine; D004229:Dithiothreitol; C056424:velnacrine