RESUMO
Activation of K+/Cl- cotransport was studied after exposure of normal human erythrocytes to the oxidative action of acetylphenylhydrazine (APH), menadione sodium bisulfite (MSB), hydrogen peroxide (H2O2) or phenazine metasulfate (PMS). In order to better define the relative contributions of K+/Cl- cotransport on ouabain and bumetanide-resistant (OBR) K+ efflux induced by oxidation, we used (dihydroindenyl)oxyalkanoic acid (DIOA) and carbocyanine as specific inhibitors, respectively, of cotransport system and Ca(2+)-activated K+ channel. APH, MSB and - to much less extent - H2O2 promoted a K+ efflux pathway with features corresponding to those of K+/Cl- cotransport. This pathway showed: (i) kinetics of efflux compatible with a specific cation transport system; (ii) requirement for chloride anion; (iii) resistance to ouabain, bumetanide and carbocyanine inhibition; (iv) stimulation by hypotonic challenge; (v) susceptibility to inhibition by DIOA. Dithiothreitol (DTT) or 2-mercaptoethanol (2-ME) decreased K+/Cl- cotransport activation, suggesting that oxidative mechanisms affected crucial SH groups of the transporter. These data suggest that oxidation represents a factor capable of modulating activation of K+/Cl- cotransport. Its possible contribution in situations with high oxidative risk, such as sickle-cell anaemia or beta thalassemia, is discussed.
Assuntos
Cloretos/metabolismo , Eritrócitos/efeitos dos fármacos , Oxidantes/farmacologia , Potássio/metabolismo , Transporte Biológico/efeitos dos fármacos , Senescência Celular , Eritrócitos/metabolismo , Hemoglobinas/metabolismo , Humanos , Peróxido de Hidrogênio/farmacologia , Concentração de Íons de Hidrogênio , Íons , Oxirredução , Fenil-Hidrazinas/farmacologia , Vitamina K/análogos & derivados , Vitamina K/farmacologia , Vitamina K 3RESUMO
The in vivo biosynthesis of thromboxane B2 (TXB2) in man is currently evaluated by measuring urinary excretion of its major urinary metabolites, 11-dehydro-TXB2 and 2,3-dinor-TXB2. 11-Dehydro-2,3-dinor-TXB2, another prominent metabolite of exogenous TXB2 in man, has never been measured in human urine. We measured urinary 11-dehydro-2,3-dinor-TXB2 in parallel with 11-dehydro-TXB2 and 2,3-dinor-TXB2 by immunoaffinity extraction/gas chromatography-mass spectrometry in healthy non-smokers (n = 12) and age-matched smokers (n = 11). In non-smokers, urinary excretion of 11-dehydro-2,3-dinor-TXB2, 11-dehydro-TXB2 and 2,3-dinor-TXB2 was 29.7 +/- 11.1, 53.6 +/- 15.0 and 13.5 +/- 2.8 ng/h (mean +/- SD), respectively. In smokers, only urinary excretion of 2,3-dinor-TXB2 was significantly different (19.7 +/- 6.7 ng/h, p < 0.01). Selective inhibition of platelet thromboxane biosynthesis by chronic low-dose aspirin (30 mg/day for 8 days, 4 subjects) comparably reduced platelet-derived metabolites and 11-dehydro-2,3-dinor-TXB2, suggesting that the latter also derives from platelets in healthy subjects.