Vicinal dithiol-binding agent, phenylarsine oxide, inhibits inducible nitric-oxide synthase gene expression at a step of nuclear factor-kappaB DNA binding in hepatocytes.
Inflammatory cytokine interleukin 1beta induces inducible nitric-oxide synthase (iNOS) mRNA and its protein, which are followed by increasing the production of nitric oxide, in primary cultures of rat hepatocytes. Nuclear factor-kappaB (NF-kappaB), an important transcription factor for iNOS gene expression, is also activated and translocated to the nucleus. In the present study, we found that vicinal dithiol-binding agent, phenylarsine oxide (PAO), inhibited the induction of iNOS protein and mRNA as well as the release of nitrite (nitric oxide metabolite) into the culture medium. Simultaneous addition of a vicinal dithiol compound, 2, 3-dimercaptopropanol, with PAO completely abolished these inhibitions. PAO could not prevent either degradation of an inhibitory protein, IkappaB, of NF-kappaB or translocation of NF-kappaB to the nucleus. However, electrophoretic mobility shift assay demonstrated that PAO decreased the interaction between NF-kappaB and its binding consensus oligonucleotide. Transfection experiments with iNOS promoter-luciferase construct revealed that PAO inhibited NF-kappaB binding to DNA. These results indicate that PAO inhibits iNOS gene expression at a step of NF-kappaB binding to DNA by modifying its vicinal dithiol moiety, which may play a crucial role for the iNOS regulation in hepatocytes.
Oda
M
M
First Department of Surgery, Kansai Medical University, Moriguchi, Osaka 570-8506, Japan.
Sakitani
K
K
Kaibori
M
M
Inoue
T
T
Kamiyama
Y
Y
Okumura
T
T
D000818:Animals; D001152:Arsenicals / Q000494:pharmacology*; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004112:Dimercaprol / Q000494:pharmacology; D004790:Enzyme Induction / Q000187:drug effects; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D017930:Genes, Reporter; D007375:Interleukin-1 / Q000494:pharmacology; D008099:Liver / Q000201:enzymology; D008297:Male; D016328:NF-kappa B / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics*; D052247:Nitric Oxide Synthase Type II; D012333:RNA, Messenger / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar; D013438:Sulfhydryl Compounds / Q000737:chemistry*; D015533:Transcriptional Activation / Q000187:drug effects; D014162:Transfection
D001152:Arsenicals; D004268:DNA-Binding Proteins; D007375:Interleukin-1; D016328:NF-kappa B; D012333:RNA, Messenger; D013438:Sulfhydryl Compounds; D004112:Dimercaprol; C029341:oxophenylarsine; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat