Induction of glutathione peroxidase in response to inactivation by nitric oxide.
To determine effect of nitric oxide (NO) on cellular glutathione peroxidase (GPX) level in living cells, we measured the activity, protein and mRNA of GPX in rat kidney (KNRK) cells under a high NO condition. Combined treatment of lipopolysaccharide (LPS, 1 microgram/ml) and tumor necrosis factor-alpha (TNF-alpha, 50 ng/ml) synergistically enhanced (23-folds) nitrite production from KNRK cells. This was suppressed by an inducible NO synthase (iNOS) inhibitor (aminoguanidine, N-nitro-L-arginine methylester hydrochloride) and arginase. iNOS expression was detected by RT-PCR in the treated cells. GPX was inactivated irreversibly when the cells had been homogenized before exposure to a NO donor, S-nitroso-N-acetylpenicillamine (SNAP). In living KNRK cells, SNAP and LPS + TNF-alpha exerted a transient effect on the GPX activity. The treatment with SNAP (200 microM) or sodium nitroprusside (200 microM) enhanced GPX gene expression, which was blocked by a NO scavenger, 2-phenyl-4,4,5,5,-tetramethylimidazoline-1-oxyl-3-oxide. GPX mRNA was markedly increased by the treatment with LPS + TNF-alpha, and aminoguanidine blocked the effect. In cells metabolically labeled with 75Se, LPS + TNF-alpha accelerated the incorporation of radioactivity into GPX molecule by 2.1-fold. These results suggest that inactivation of GPX by NO triggers a signal for inducing GPX gene expression in KNRK cells, thereby restoring the intracellular level of this indispensable enzyme.
Dobashi
K
K
Department of Pediatrics, Yamanashi Medical University, Yamanashi, 409-3898 Japan. kdobashi@res.yamanashi-med.ac.jp
Asayama
K
K
Nakane
T
T
Kodera
K
K
Hayashibe
H
H
Nakazawa
S
S
D000818:Animals; D002461:Cell Line, Transformed; D002470:Cell Survival / Q000187:drug effects; D004790:Enzyme Induction; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005979:Glutathione Peroxidase / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D007668:Kidney / Q000187:drug effects* / Q000201:enzymology; D008070:Lipopolysaccharides / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism / Q000494:pharmacology*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D052247:Nitric Oxide Synthase Type II; D010396:Penicillamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D004791:Enzyme Inhibitors; D008070:Lipopolysaccharides; D020030:Nitric Oxide Donors; D012333:RNA, Messenger; C110881:S-nitro-N-acetylpenicillamine; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide; D005979:Glutathione Peroxidase; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D010396:Penicillamine