title: Inhibition of lipopolysaccharide-induced I-kappaB degradation and tumor necrosis factor-alpha expression by acriflavine, an antimicrobial agent.
abstract: Acriflavine neutral (ACF) has been used for treatment of microbial infections for humans and fishes. Effects of ACF on the nuclear factor-kappaB (NF-kappaB) activation and tumor necrosis factor-alpha (TNF-alpha) production by lipopolysaccharide (LPS), an endotoxin, were examined in rat and RAW264.7 cells. Gel retardation analysis revealed that LPS (1 microg/kg) activated NF-kappaB in the liver, whereas pretreatment of rats with ACF (10 mg/kg) completely prevented the NF-kappaB activation. Selectivity of the NF-kappaB DNA binding was confirmed by immunodepletion with anti-p65 and anti-p50 antibodies. Translocation of NF-kappaB to the nucleus is preceded by phosphorylation and proteolytic degradation of inhibitor-kappaBalpha (I-kappaBalpha) subunit. Whereas the level of I-kappaBalpha protein was rapidly decreased after treatment of rats with LPS (1 microg/kg), ACF treatment prior to LPS attenuated the decrease in I-kappaBalpha protein level. LPS-induced increase in the production of TNF-alpha, the principal inflammatory mediator, was prevented by ACF pretreatment by 80%. Stimulation of RAW264.7 cells with 1 microg/ml of LPS caused an increase in DNA binding activity of NF-kappaB, which was 80% inhibited by 1 microg/ml of ACF. LPS reduced I-kappaBalpha level in RAW264.7 cells by 77%. ACF attenuated LPS-induced decrease in I-kappaBalpha protein in a concentration-dependent manner. Production of TNF-alpha by LPS from RAW264.7 cells was decreased by 84% in the presence of ACF. Data showed that ACF inhibited LPS-induced NF-kappaB activation through inhibition of I-kappaBalpha degradation and TNF-alpha production in both rat and RAW264.7 cells. Inhibition of NF-kappaB activation and TNF-alpha production may be associated with the anti-inflammatory activity of ACF.
authors.affiliation: College of Pharmacy, Seoul National University, South Korea.
authors.forename: S H
authors.initials: SH
authors.lastname: Choi
authors.forename: J Y
authors.initials: JY
authors.lastname: Cho
authors.forename: Y S
authors.initials: YS
authors.lastname: Chung
authors.forename: E
authors.initials: E
authors.lastname: Hong
authors.forename: Y
authors.initials: Y
authors.lastname: Han
authors.forename: S G
authors.initials: SG
authors.lastname: Kim
mesh_terms: D000167:Acriflavine / Q000494:pharmacology*; D000818:Animals; D000891:Anti-Infective Agents, Local / Q000494:pharmacology*; D002460:Cell Line; D015342:DNA Probes; D003720:Densitometry; D004797:Enzyme-Linked Immunosorbent Assay; D020672:I-kappa B Proteins / Q000378:metabolism*; D015151:Immunoblotting; D008070:Lipopolysaccharides / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D013347:Subcellular Fractions / Q000187:drug effects / Q000378:metabolism; D013778:Tetrazolium Salts; D013844:Thiazoles; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis*
chemical_list: D000891:Anti-Infective Agents, Local; D015342:DNA Probes; D020672:I-kappa B Proteins; D008070:Lipopolysaccharides; D013778:Tetrazolium Salts; D013844:Thiazoles; D014409:Tumor Necrosis Factor-alpha; D000167:Acriflavine; C022616:thiazolyl blue