Inhibitory effect of E3330, a novel quinone derivative able to suppress tumor necrosis factor-alpha generation, on activation of nuclear factor-kappa B.
(2E)-3-[5-(2,3-Dimethoxy-6-methyl-1,4-benzoquinoyl)]-2-nonyl-2- propenoic acid (E3330), is a novel agent with hepatoprotective activity. We report the effect of E3330 on transcriptional activation of tumor necrosis factor (TNF)-alpha gene and on nuclear factor (NF)-kappa B activation. Nuclear run-on experiments showed that E3330 decreases transcriptional activation of TNF-alpha gene induced by lipopolysaccharide (LPS) stimulation in human peripheral monocytes. To investigate the inhibitory mechanisms, we constructed a secreted-type placental alkaline phosphatase (PLAP) reporter gene whose transcription is controlled by a 1.4-kb human TNF-alpha promoter. A stable transformant of the PLAP reporter gene derived from human monocytic cell line showed very little activity on the promoter before stimulation, whereas LPS stimulation led to a dramatic increase in PLAP activity. E3330 inhibited this induced promoter activity in a dose-dependent manner. There are four putative NF-kappa B binding sites (kappa B-1, kappa B-2, kappa B-3, kappa B-4) in human TNF-alpha promoter. By using mutated promoter-PLAP plasmids, we established that these NF-kappa B sites were necessary for induction of TNF-alpha transcription on stimulation with LPS. A gel retardation experiment with synthetic double-stranded oligonucleotides showed that activated NF-kappa B consisting of p50/ p65 heterodimer bound to all four putative NF-kappa B DNA probes, suggesting that all four putative NF-kappa B recognition sites play an important role in inducible TNF-alpha expression. E3330 decreased activated NF-kappa B in nuclei, suggesting that E3330 inhibits NF-kappa B activation and/or translocation of the nuclei. Western blotting analysis with anti-I kappa B-alpha antibody indicated that E3330 inhibited degradation of I kappa B-alpha, which is an inhibitory protein of NF-kappa B, in LPS-stimulated monocytes. E3330 may suppress the production of active oxygen species serving as common messengers to activate NF-kappa B.[1]References
- Inhibitory effect of E3330, a novel quinone derivative able to suppress tumor necrosis factor-alpha generation, on activation of nuclear factor-kappa B. Goto, M., Yamada, K., Katayama, K., Tanaka, I. Mol. Pharmacol. (1996) [Pubmed]
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