Induction of nitric oxide synthase mRNA expression. Suppression by exogenous nitric oxide.
The reactive nitrogen species, nitric oxide (NO), plays an important role in the pathogenesis of neurodegenerative diseases. The suppression of NO production may be fundamental for survival of neurons. Here, we report that pretreatment of human ramified microglial cells with nearly physiological levels of exogenous NO prevents lipopolysaccharide (LPS)/tumor necrosis factor alpha (TNF alpha)-inducible NO synthesis, because by affecting NF-kappa B activation it inhibits inducible Ca(2+)-independent NO synthase isoform (iNOS) mRNA expression. Using reverse transcriptase polymerase chain reaction, we have found that both NO donor sodium nitroprusside (SNP) and authentic NO solution are able to inhibit LPS/TNF alpha-inducible iNOS gene expression; this effect was reversed by reduced hemoglobin, a trapping agent for NO. The early presence of SNP during LPS/TNF alpha induction is essential for inhibition of iNOS mRNA expression. Furthermore, SNP is capable of inhibiting LPS/TNF alpha-inducible nitrite release, as determined by Griess reaction. Finally, using electrophoretic mobility shift assay, we have shown that SNP inhibits LPS/TNF alpha-elicited NF-kappa B activation. This suggests that inhibition of iNOS gene expression by exogenous NO may be ascribed to a decreased NF-kappa B availability.
Colasanti
M
M
Department of Biology, III University of Rome, Italy.
Persichini
T
T
Menegazzi
M
M
Mariotto
S
S
Giordano
E
E
Caldarera
C M
CM
Sogos
V
V
Lauro
G M
GM
Suzuki
H
H
D001483:Base Sequence; D001665:Binding Sites; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology; D017628:Microglia / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism; D009422:Nervous System Diseases / Q000209:etiology; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics*; D009599:Nitroprusside / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000235:genetics*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D008070:Lipopolysaccharides; D016328:NF-kappa B; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D009599:Nitroprusside; D009569:Nitric Oxide; D004247:DNA; D019001:Nitric Oxide Synthase