RESUMO
To clarify the presence of cross-talk between H(2)S and NO, we investigated effect of NaHS, an H(2)S donor, on activity of recombinant NO synthase (NOS) isoforms. Activity of all nNOS, iNOS and eNOS was inhibited by NaHS (IC(50): 0.13-0.21 mM). In contrast, Na(2)SO(3), L-cysteine and threo-1,4-dimercapto-2,3-butanediol, a reductant, exerted poor inhibition of NOS activity. Increasing concentrations of tetrahydrobiopterin (BH(4)) reversed the NaHS inhibition of nNOS and eNOS, but not iNOS. Our data thus demonstrate inhibition of three NOS isoforms by NaHS/H(2)S, and suggest involvement of interaction of NaHS/H(2)S with BH(4) in inhibition of nNOS and eNOS, but not iNOS.
Assuntos
Poluentes Atmosféricos/toxicidade , Inibidores Enzimáticos , Sulfeto de Hidrogênio/toxicidade , Óxido Nítrico Sintase Tipo III/antagonistas & inibidores , Óxido Nítrico Sintase Tipo II/antagonistas & inibidores , Óxido Nítrico Sintase Tipo I/antagonistas & inibidores , Biopterinas/análogos & derivados , Biopterinas/metabolismo , Cisteína/toxicidade , Ditiotreitol/toxicidade , Humanos , NADP/metabolismo , Proteínas Recombinantes , Sulfatos/toxicidadeRESUMO
This study examines the application of Ishikawa human endometrial adenocarcinoma cells to measure the estrogenic activity of fractionated extracts of sediments from Tokyo Bay, Japan. Estrogen stimulates alkaline phosphatase activity in this cell line. The results of these assays were compared with those of a yeast estrogen screen (YES) assay. The Ishikawa cell line bioassay showed higher sensitivity to 17beta-estradiol (median effective concentration [EC50], 10.7 pM) than did the YES assay (EC50, 480 pM). Fractionation of sediment extracts (all samples collected from 5 sites) showed that the nonpolar fraction was poisonous to yeast cells; the estrogenic activity of this fraction, therefore, could not be measured by YES. However, the nonpolar fraction did not kill the Ishikawa cells. The 17beta-estradiol-equivalent values of 15 extracts (3 fractions from each of 5 sediment samples) ranged from 5.7 to 697 pg/g dry weight according to the Ishikawa cell line bioassay. Chemical analysis using gas chromatography-mass spectrometry revealed that the highest concentrations of endocrine-disrupting chemicals were observed at the sampling station near the sewage treatment plant. The results support that the Ishikawa cell line bioassay is suitable for measuring the estrogenic activity of sediment samples.