title: Intestinal epithelial cells with impaired autophagy lose their adhesive capacity in the presence of TNF-α.
abstract: OBJECTIVE
  Genome-wide association studies have revealed a link between autophagy-related (ATG) genes and susceptibility to Crohn's disease. This suggests underlying involvement of autophagy impairment in the pathogenesis of Crohn's disease. This study was performed to investigate the pathophysiological importance of autophagy impairment in intestinal epithelial cells exposed to TNF-α.
  
  
  METHODS
  Human colonic epithelial cells (HT-29) and rat small intestinal epithelial cells (IEC-18) were used. Formation of phosphatidylethanolamine-conjugated microtubule-associated protein light chain 3 (LC3-II) was monitored as a marker of autophagy. Autophagy was inhibited using 3-methyladenine or short interfering RNA targeting ATG5 and ATG16L1.
  
  
  RESULTS
  TNF-α treatment elicited a significant dose-dependent increase in LC3-II protein levels, thus autophagy is induced in the presence of TNF-α. Combined autophagy inhibition and TNF-α treatment induced a marked increase in the number of detached cells and a decrease in activated integrin β1 protein levels. Trypan blue staining indicated 70-80 % of the detached cells were alive, suggesting that these cells became detached not because they were killed but because of dysfunction of cellular adhesion.
  
  
  CONCLUSIONS
  This is the first study indicating that intestinal epithelial cells with impaired autophagy lose their adhesive capacity in the presence of TNF-α. These observations indicate that impairment of autophagy leads to disruption of the intestinal epithelial cell layers in TNF-α-rich environments.
authors.affiliation: Department of Medicine and Clinical Oncology (K1), Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba-shi 260-8670, Japan.
authors.forename: Masaya
authors.initials: M
authors.lastname: Saito
authors.forename: Tatsuro
authors.initials: T
authors.lastname: Katsuno
authors.forename: Tomoo
authors.initials: T
authors.lastname: Nakagawa
authors.forename: Toru
authors.initials: T
authors.lastname: Sato
authors.forename: Yoshiko
authors.initials: Y
authors.lastname: Noguchi
authors.forename: Sayuri
authors.initials: S
authors.lastname: Sazuka
authors.forename: Keiko
authors.initials: K
authors.lastname: Saito
authors.forename: Makoto
authors.initials: M
authors.lastname: Arai
authors.forename: Koutaro
authors.initials: K
authors.lastname: Yokote
authors.forename: Osamu
authors.initials: O
authors.lastname: Yokosuka
mesh_terms: D000818:Animals; D001343:Autophagy; D002448:Cell Adhesion; D003424:Crohn Disease / Q000209:etiology; D004847:Epithelial Cells / Q000166:cytology; D019204:GTP-Binding Proteins / Q000378:metabolism; D019073:HT29 Cells; D006801:Humans; D019012:Integrin beta1 / Q000378:metabolism; D007413:Intestinal Mucosa / Q000166:cytology / Q000502:physiology*; D051381:Rats; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology*; D033921:Vesicular Transport Proteins
chemical_list: D019012:Integrin beta1; D014409:Tumor Necrosis Factor-alpha; D033921:Vesicular Transport Proteins; C577013:Atg16L1 protein, rat; D019204:GTP-Binding Proteins