Molecular mechanisms of repression of eotaxin expression with fluticasone propionate in airway epithelial cells.
BACKGROUND
Glucocorticoids are known to repress the expression of CC chemokine eotaxin in airway epithelial cells. We focused our study on the molecular mechanisms of the glucocorticoid, fluticasone, in the inhibition of the expression of the eotaxin gene in the cells.


METHODS
The airway epithelial cell line, BEAS-2B, was stably transfected with signal transducers and activators of transcription 6 (STAT6)-expressing vector and used in the following experiments to clarify the function of STAT6. Levels of eotaxin mRNA and protein expression were determined with RT-PCR and ELISA. Mechanisms of transcriptional regulation were assessed by the electrophoretic mobility shift assay and dual luciferase assay using eotaxin promoter-luciferase reporter plasmids.


RESULTS
Fluticasone significantly inhibited the induction of eotaxin protein stimulated with TNF-alpha and IL-4 in the cells. Fluticasone also repressed the induction of eotaxin mRNA with these stimuli. It partially inhibited the activity of eotaxin promoter; however, it did not inhibit the nuclear translocation and binding of transcription factors, nuclear factor-kappa B (NF-kappaB) or STAT6, to the DNA derived from the proximal promoter region of the eotaxin gene. Moreover, the inhibitory effect was also conserved in the experiments using the reporter plasmid of which the putative glucocorticoid-responsive element was mutated.


CONCLUSIONS
Fluticasone inhibits the expression of eotaxin gene in airway epithelial cells in part through repression of the transcription. However, the mechanisms depend neither on the inhibition of transcription factors' translocation into nuclei nor the function of the putative glucocorticoid-responsive element in the promoter, indicating that other mechanisms would be related to the transcriptional repression of the eotaxin gene in airway epithelial cells.
Matsukura
Satoshi
S
First Department of Internal Medicine, Showa University School of Medicine, Tokyo, Japan. smatsuku@med.showa-u.ac.jp
Kokubu
Fumio
F
Kurokawa
Masatsugu
M
Kawaguchi
Mio
M
Kuga
Hideki
H
Ieki
Koushi
K
Odaka
Miho
M
Suzuki
Shintaro
S
Watanabe
Shin
S
Takeuchi
Hiroko
H
Schleimer
Robert P
RP
Schindler
Ulrike
U
Adachi
Mitsuru
M
D000730:Androstadienes / Q000494:pharmacology*; D001993:Bronchodilator Agents / Q000494:pharmacology*; D002461:Cell Line, Transformed; D054413:Chemokine CCL11; D019742:Chemokines, CC / Q000096:biosynthesis*; D024202:Electrophoretic Mobility Shift Assay; D004797:Enzyme-Linked Immunosorbent Assay; D000068298:Fluticasone; D006801:Humans; D008168:Lung / Q000187:drug effects / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D020545:Respiratory Mucosa / Q000187:drug effects* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050800:STAT6 Transcription Factor; D015534:Trans-Activators / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection
D000730:Androstadienes; D001993:Bronchodilator Agents; C515456:CCL11 protein, human; D054413:Chemokine CCL11; D019742:Chemokines, CC; D016328:NF-kappa B; D050800:STAT6 Transcription Factor; C494101:STAT6 protein, human; D015534:Trans-Activators; D000068298:Fluticasone