Geldanamycin inhibits NF-kappaB activation and interleukin-8 gene expression in cultured human respiratory epithelium.
Geldanamycin is a benzoquinone ansamycin with multiple pharmacologic properties. Recent data demonstrated that geldanamycin conferred protection in an animal model of inflammation-associated acute lung injury. In the current study, we investigated the effects of geldanamycin on interleukin (IL)-8 gene expression and nuclear factor (NF)-kappaB activation. Geldanamycin inhibited tumor necrosis factor (TNF)-alpha-mediated IL-8 gene expression in A549 human respiratory epithelial cells as measured by enzyme-linked immunosorbent assay and Northern blot analyses. In cells transiently transfected with an IL-8 promoter-luciferase reporter plasmid, geldanamycin inhibited TNF-alpha-mediated luciferase activity. Geldanamycin inhibited TNF-alpha-mediated NF-kappaB activation as measured by electromobility shift assays and transient transfections with a NF-kappaB-dependent luciferase reporter plasmid. In contrast, geldanamycin did not affect TNF-alpha-mediated degradation of the NF-kappaB inhibitory protein IkappaBalpha and did not block nuclear translocation of the NF-kappaB p65 subunit as measured by Western blot analyses. Geldanamycin added directly to nuclear extracts of TNF-alpha-treated cells reduced the formation of the NF-kappaB/DNA complex. These results demonstrate that geldanamycin inhibits TNF-alpha-mediated IL-8 gene expression in A549 cells by inhibiting activation of the IL-8 promoter. The mechanism of inhibition involves inhibition of NF-kappaB activation, which is independent of IkappaBalpha degradation or p65 nuclear translocation. Geldanamycin appears to directly inhibit the ability of NF-kappaB to bind DNA. The observed in vitro effects could account, in part, for the anti-inflammatory properties of geldanamycin observed in vivo.
Malhotra
V
V
Division of Critical Care Medicine, Children's Hospital Medical Center and Children's Hospital Research Foundation, 3333 Burnet Ave., Cincinnati, OH 45229, USA.
Shanley
T P
TP
Pittet
J F
JF
Welch
W J
WJ
Wong
H R
HR
D016227:Benzoquinones; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D015342:DNA Probes; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D006868:Hydrolysis; D016209:Interleukin-8 / Q000235:genetics*; D047029:Lactams, Macrocyclic; D008168:Lung / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D016328:NF-kappa B / Q000037:antagonists & inhibitors* / Q000378:metabolism; D021381:Protein Transport; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011809:Quinones / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000494:pharmacology
D016227:Benzoquinones; D015342:DNA Probes; D004791:Enzyme Inhibitors; D016209:Interleukin-8; D047029:Lactams, Macrocyclic; D016328:NF-kappa B; D011809:Quinones; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D011505:Protein-Tyrosine Kinases; C001277:geldanamycin