title: Autophagy is required for toll-like receptor-mediated interleukin-8 production in intestinal epithelial cells.
abstract: Autophagy is an evolutionarily conserved process that maintains cellular homeostasis via synthesis, degradation, and subsequent recycling of cellular products under various physiological conditions. However, the link between autophagy and the innate immune system remains unknown. In the present study, we evaluated Toll-like receptor (TLR)-mediated autophagy induction in intestinal epithelial cells (IECs) and its relationship to interleukin (IL)-8 production. IEC-6, HCT-15, RAW264.7, and THP-1 cells were cultured with or without various TLR ligands, followed by evaluation of the expressions of pro-inflammatory cytokines [IL-8, cytokine-induced neutrophil chemoattractants (CINC)-2β, macrophage inflammatory protein (MIP)-2] by real-time PCR and ELISA. To reveal the status of autophagy in IECs and macrophages, light chain 3 (LC3)-II expression was examined using Western blotting and immunofluorescence with confocal microscopy. Also, to evaluate the influence of TLR ligands on autophagy-mediated innate-immune responses, autophagy-related gene (Atg)7 specific siRNA was transfected into intestinal epithelial cells and IL-8 expression was determined following exposure to various TLR ligands. Cells treated with the TLR ligands produced considerable amounts of pro-inflammatory cytokines (IL-8, CINC-2β, MIP-2). Furthermore, the basal levels of LC3-II were markedly higher in IECs as compared to those in macrophages. Our findings indicated that autophagy induction following TLR ligand stimulation was not significantly evident in IECs as compared to macrophages. In addition, Atg7 gene expression silencingled to down-regulation of TLR-mediated IL-8 expression in IECs, which indicates a potential role of autophagy in generating innate-immune responses. In conclusion, autophagy may be an important intracellular machinery for inducing the innate immune system in IECs.
authors.affiliation: Department of Internal Medicine II, Shimane University School of Medicine, Izumo, Shimane, Japan.
authors.forename: Yong-Yu
authors.initials: YY
authors.lastname: Li
authors.forename: Shunji
authors.initials: S
authors.lastname: Ishihara
authors.forename: M Monowar
authors.initials: MM
authors.lastname: Aziz
authors.forename: Akihiko
authors.initials: A
authors.lastname: Oka
authors.forename: Ryusaku
authors.initials: R
authors.lastname: Kusunoki
authors.forename: Yasumasa
authors.initials: Y
authors.lastname: Tada
authors.forename: Takafumi
authors.initials: T
authors.lastname: Yuki
authors.forename: Yuji
authors.initials: Y
authors.lastname: Amano
authors.forename: Mesbah Uddin
authors.initials: MU
authors.lastname: Ansary
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Kinoshita
mesh_terms: D000818:Animals; D001343:Autophagy / Q000502:physiology*; D000071193:Autophagy-Related Protein 7; D002460:Cell Line; D054426:Chemokine CXCL2 / Q000096:biosynthesis / Q000276:immunology; D019743:Chemokines, CXC / Q000096:biosynthesis / Q000276:immunology; D004847:Epithelial Cells / Q000276:immunology / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D007113:Immunity, Innate / Q000502:physiology; D016209:Interleukin-8 / Q000096:biosynthesis* / Q000276:immunology; D007413:Intestinal Mucosa / Q000276:immunology / Q000378:metabolism*; D008264:Macrophages / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000096:biosynthesis / Q000276:immunology; D051381:Rats; D051193:Toll-Like Receptors / Q000276:immunology / Q000378:metabolism*; D044764:Ubiquitin-Activating Enzymes / Q000096:biosynthesis / Q000276:immunology
chemical_list: C502661:Atg7 protein, mouse; C508606:CXCL8 protein, human; D054426:Chemokine CXCL2; D019743:Chemokines, CXC; C404915:Cxcl2 protein, mouse; C513296:Cxcl2 protein, rat; C513294:Cxcl3 protein, rat; D016209:Interleukin-8; D008869:Microtubule-Associated Proteins; D051193:Toll-Like Receptors; C509349:ATG7 protein, human; D000071193:Autophagy-Related Protein 7; D044764:Ubiquitin-Activating Enzymes