Inhibition of nitric oxide synthase expression by PPM-18, a novel anti-inflammatory agent, in vitro and in vivo.
We studied the effect of PPM-18, a chemically synthesized naphthoquinone derivative and also an anti-inflammatory agent, on the lipopolysaccharide (LPS)-activated inducible NO synthase (iNOS) expression in rat alveolar macrophages. Pretreatment of macrophages with PPM-18 (0.1-10 microM) significantly inhibited nitrite production, iNOS protein expression and iNOS mRNA accumulation. PPM-18 did not directly affect the enzymic activities of iNOS and other constitutive NOS forms. The LPS-induced increase in nuclear transcription factor kappaB (NF-kappaB) p65 and p50 in nucleus was suppressed by PPM-18 (10 microM). Moreover electrophoretic mobility-shift assays demonstrated that PPM-18 inhibited DNA binding to NF-kappaB induced by LPS in whole cells but not when added in the nuclear extract, suggesting that PPM-18 did not interfere directly with the binding of NF-kappaB to DNA and that some events had to be processed before NF-kappaB could bind DNA. Examination of NF-kappaB showed that PPM-18 stabilized the NF-kappaB inhibitor, IkappaBalpha, by preventing its degradation from NF-kappaB. Therefore the stabilization of IkappaBalpha might have contributed to the inhibition of NF-kappaB activation. These results also indicate strongly that NF-kappaB is involved in the production of NO on stimulation by LPS. PPM-18 significantly decreased the production of tumour necrosis factor alpha in response to LPS. PPM-18 protects mice against LPS-induced lethal toxicity. These results also indicate that PPM-18 is a potent inhibitor of iNOS expression by blocking the binding of NF-kappaB to promoter and exerts a beneficial effect in the mouse model of sepsis.
Yu
S M
SM
Department of Pharmacology, Chang Gung Medical College, 259 Wen-Hwa 1st Road, Kwei-San, Tao-Yuan, Taiwan.
Wu
J F
JF
Lin
T L
TL
Kuo
S C
SC
D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004790:Enzyme Induction / Q000187:drug effects; D007022:Hypotension / Q000517:prevention & control; D020672:I-kappa B Proteins; D008070:Lipopolysaccharides / Q000633:toxicity; D016676:Macrophages, Alveolar / Q000201:enzymology*; D051379:Mice; D008807:Mice, Inbred BALB C; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism; D009285:Naphthoquinones / Q000494:pharmacology*; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000096:biosynthesis* / Q000235:genetics; D009573:Nitrites / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D051381:Rats; D017208:Rats, Wistar; D012772:Shock, Septic / Q000517:prevention & control; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D000894:Anti-Inflammatory Agents, Non-Steroidal; D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; D009285:Naphthoquinones; C000606205:Nfkbia protein, mouse; C000606206:Nfkbia protein, rat; D009573:Nitrites; D014409:Tumor Necrosis Factor-alpha; C109682:alpha-benzoylamino-1,4-naphthoquinone; D000072000:NF-KappaB Inhibitor alpha; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase