Expression of citrulline-nitric oxide cycle in lipopolysaccharide and cytokine-stimulated rat astroglioma C6 cells.
Nitric oxide (NO) is involved in many physiological and pathological processes in the brain. NO is synthesized from arginine by nitric oxide synthase (NOS), with citrulline generated as a by-product of the reaction. Thus, citrulline can by recycled to arginine by argininosuccinate synthetase (AS) and argininosuccinate lyase (AL) via the citrulline-NO cycle. Rat astroglioma C6 cells were treated with bacterial lipopolysaccharide (LPS), interferon-gamma (IFNgamma) and tumor necrosis factor-alpha, and the expression of the enzymes of the citrulline-NO cycle was investigated by RNA blot and immunoblot analyses. NO production from arginine and citrulline was also assessed. iNOS mRNA and protein were induced 6-12 h after stimulation with LPS and cytokines and decreased at 24 h. AS mRNA increased up to 12 h and decreased at 24 h. AS protein increased gradually up to 48 h. On the other hand, AL mRNA remained unchanged by stimulation. NO production from arginine was enhanced by the treatment with LPS and cytokines. NO production was also observed when arginine was replaced by citrulline. These results indicate that NO production is enhanced in LPS- and cytokine-stimulated C6 cells due to induction of iNOS and that the citrulline-arginine recycling is important for NO production.
Zhang
W Y
WY
Department of Molecular Genetics, Kumamoto University School of Medicine, Honjo 2-2-1, Kumamoto, Japan.
Takiguchi
M
M
Koshiyama
Y
Y
Gotoh
T
T
Nagasaki
A
A
Iwase
K
K
Yamamoto
K
K
Takeshima
H
H
Negi
A
A
Mori
M
M
D000818:Animals; D001120:Arginine / Q000378:metabolism; D001254:Astrocytoma; D002956:Citrulline / Q000378:metabolism*; D016207:Cytokines / Q000494:pharmacology*; D004790:Enzyme Induction; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D007371:Interferon-gamma / Q000494:pharmacology; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology*; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000096:biosynthesis / Q000235:genetics*; D052247:Nitric Oxide Synthase Type II; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D011994:Recombinant Proteins; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D019323:omega-N-Methylarginine / Q000494:pharmacology
D016207:Cytokines; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D019323:omega-N-Methylarginine; D002956:Citrulline; D009569:Nitric Oxide; D007371:Interferon-gamma; D001120:Arginine; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D019331:NG-Nitroarginine Methyl Ester