title: High-dose tobramycin inhibits lipopolysaccharide-induced MUC5AC production in human lung epithelial cells.
abstract: Tobramycin inhalation therapy is an effective therapy for cystic fibrosis as well as severe bronchiectasis that is colonized with Pseudomonas aeruginosa. The mechanism responsible for the efficacy of tobramycin in the treatment of severe chronic infectious diseases has not been elucidated. We demonstrate that high-dose tobramycin decreases MUC5AC gene expression and protein production in NCI-H292 cell stimulated with lipopolysaccharide of P. aeruginosa. MUC5AC protein of NCI-H292 cell stimulated by lipopolysaccharide was analyzed by an enzyme-linked immunosorbent assay and MUC5AC expression at the mRNA level was analyzed by RT-PCR. Western blot was performed to examine a potential role for the signaling molecules upstream of NFκB. High-dose tobramycin (500μg/ml) decreased the level of MUC5AC protein released into the supernatant of the NCI-H292 cell line at 24h after lipopolysaccharide stimulation (P<0.001). The tobramycin treatment also inhibited MUC5AC mRNA expression at 12h after lipopolysaccharide stimulation (P<0.05) and suppressed NFκB activation 60min after lipopolysaccharide stimulation (P<0.001). Tobramycin suppressed the phosphorylation of extracellular signal-regulated protein kinase, p38 MAP kinase. These results suggest that high-dose tobramycin, such as inhalation therapy, can inhibit MUC5AC gene expression and MUC5AC protein production in NCI-H292 cells, in part through the mitogen-activated protein kinase pathway. Thus, the activation of TLR4 and the subsequent immune/inflammatory responses induced by chronic infections such as P. aeruginosa might be modulated by tobramycin. Our data may reveal one of the mechanisms responsible for the clinical effect of tobramycin inhalation therapy against severe chronic respiratory diseases due to P. aeruginosa.
authors.affiliation: Second Department of Internal Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Shigeki
authors.initials: S
authors.lastname: Nakamura
authors.affiliation: Second Department of Internal Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan; Department of Laboratory Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Katsunori
authors.initials: K
authors.lastname: Yanagihara
authors.affiliation: Department of Laboratory Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Nobuko
authors.initials: N
authors.lastname: Araki
authors.affiliation: Second Department of Internal Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan; Department of Laboratory Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Koichi
authors.initials: K
authors.lastname: Yamada
authors.affiliation: Second Department of Internal Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan; Department of Laboratory Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Yoshitomo
authors.initials: Y
authors.lastname: Morinaga
authors.affiliation: Second Department of Internal Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Koichi
authors.initials: K
authors.lastname: Izumikawa
authors.affiliation: Second Department of Internal Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Masafumi
authors.initials: M
authors.lastname: Seki
authors.affiliation: Second Department of Internal Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kakeya
authors.affiliation: Second Department of Internal Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Yamamoto
authors.affiliation: Department of Laboratory Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Shimeru
authors.initials: S
authors.lastname: Kamihira
authors.affiliation: Second Department of Internal Medicine, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan; Division of Molecular & Clinical Microbiology, Department of Molecular Microbiology & Immunology, Nagasaki University Graduate School of Medical Science, Nagasaki, Japan.
authors.forename: Shigeru
authors.initials: S
authors.lastname: Kohno
mesh_terms: D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects* / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008168:Lung / Q000166:cytology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D055271:Mucin 5AC / Q000096:biosynthesis* / Q000235:genetics; D016328:NF-kappa B / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014031:Tobramycin / Q000494:pharmacology*
chemical_list: D008070:Lipopolysaccharides; D055271:Mucin 5AC; D016328:NF-kappa B; D012333:RNA, Messenger; D020928:Mitogen-Activated Protein Kinases; D014031:Tobramycin