Biochemical and Biophysical Research Communications
Endogenous metalloprotease solubilizes IL-13 receptor α2 in airway epithelial cells☆
Section snippets
Materials and methods
Cell culture. BEAS-2B cells, a human airway epithelial cell line transformed with the SV40 virus, were cultured in DMEM/F-12 medium with 10% FBS, penicillin, and streptomycin.
Adenovirus transfection and stimulation. A replication-defective ΔE1ΔE3 adenoviral vector expressing the IL-13Rα2 chain gene under the CMV promoter (Ad-IL-13Rα2) was constructed as described previously [18]. Another replication-defective adenoviral vector expressing the β-galactosidase gene (Ad-LacZ) was constructed in the
IL-13Rα2 gene induction in airway epithelial cells and secretion of soluble L-13Rα2
IL-13Rα2 mRNA levels were barely detectable in BEAS-2B cells. To establish airway epithelial cells with a high expression of IL-13Rα2, BEAS-2B cells were transfected with a full-length IL-13Rα2 using Ad-IL-13Rα2. Ad-IL-13Rα2 transfection, but not Ad-LacZ, induced IL-13 Rα2 mRNA in a multiplicity of infection (moi)-dependent manners (Fig. 1A). Maximal induction of IL-13Rα2 was observed at 30-moi transfection of Ad-IL-13Rα2. We also analyzed the time kinetics of IL-13Rα2 mRNA expression. The high
Discussion
We demonstrate that the transfection of full-length IL-13Rα2 induces the release of soluble IL-13Rα2 from airway epithelial cells and that the transfer of conditioned media from IL-13Rα2-transfected cells inhibits IL-13-induced eotaxin production and STAT6 phosphorylation in other non-transfected cells. PMA enhances the release of soluble IL-13Rα2, and metalloprotease inhibitors block this PMA-induced release of IL-13Rα2. These findings suggest that the upregulation of membrane-bound IL-13Rα2
Acknowledgments
We thank Ayako Hashizume, Tomoko Yoshimura, Yuki Yoshiura, and the Morphology Core, Faculty of Medicine, Kyushu University, for technical assistance.
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This work was supported in part by Grant-in-Aid for Scientific Research from the Japan Society for the Promotion of Science.