title: Autophagy Induced by Intracellular Infection of Propionibacterium acnes.
abstract: BACKGROUND
  Sarcoidosis is caused by Th1-type immune responses to unknown agents, and is linked to the infectious agent Propionibacterium acnes. Many strains of P. acnes isolated from sarcoid lesions cause intracellular infection and autophagy may contribute to the pathogenesis of sarcoidosis. We examined whether P. acnes induces autophagy.
  
  
  METHODS
  Three cell lines from macrophages (Raw264.7), mesenchymal cells (MEF), and epithelial cells (HeLa) were infected by viable or heat-killed P. acnes (clinical isolate from sarcoid lymph node) at a multiplicity of infection (MOI) of 100 or 1000 for 1 h. Extracellular bacteria were killed by washing and culturing infected cells with antibiotics. Samples were examined by colony assay, electron-microscopy, and fluorescence-microscopy with anti-LC3 and anti-LAMP1 antibodies. Autophagy-deficient (Atg5-/-) MEF cells were also used.
  
  
  RESULTS
  Small and large (≥5 μm in diameter) LC3-positive vacuoles containing few or many P. acnes cells (LC3-positive P. acnes) were frequently found in the three cell lines when infected by viable P. acnes at MOI 1000. LC3-positive large vacuoles were mostly LAMP1-positive. A few small LC3-positive/LAMP1-negative vacuoles were consistently observed in some infected cells for 24 h postinfection. The number of LC3-positive P. acnes was decreased at MOI 100 and completely abolished when heat-killed P. acnes was used. LC3-positive P. acnes was not found in autophagy-deficient Atg5-/- cells where the rate of infection was 25.3 and 17.6 times greater than that in wild-type Atg5+/+ cells at 48 h postinfection at MOI 100 and 1000, respectively. Electron-microscopic examination revealed bacterial cells surrounded mostly by a single-membrane including the large vacuoles and sometimes a double or multi-layered membrane, with occasional undigested bacterial cells in ruptured late endosomes or in the cytoplasm.
  
  
  CONCLUSIONS
  Autophagy was induced by intracellular P. acnes infection and contributed to intracellular bacterial killing as an additional host defense mechanism to endocytosis or phagocytosis.
authors.affiliation: Department of Human Pathology, Graduate School and Faculty of Medicine, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
authors.forename: Teruko
authors.initials: T
authors.lastname: Nakamura
authors.affiliation: Department of Human Pathology, Graduate School and Faculty of Medicine, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
authors.forename: Asuka
authors.initials: A
authors.lastname: Furukawa
authors.affiliation: Division of Surgical Pathology, Tokyo Medical and Dental University Hospital, Tokyo 113-8510, Japan.
authors.forename: Keisuke
authors.initials: K
authors.lastname: Uchida
authors.affiliation: Department of Human Pathology, Graduate School and Faculty of Medicine, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
authors.forename: Tomohisa
authors.initials: T
authors.lastname: Ogawa
authors.affiliation: Division of Surgical Pathology, Tokyo Medical and Dental University Hospital, Tokyo 113-8510, Japan.
authors.forename: Tomoki
authors.initials: T
authors.lastname: Tamura
authors.affiliation: Department of Human Pathology, Graduate School and Faculty of Medicine, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Sakonishi
authors.affiliation: Department of Human Pathology, Graduate School and Faculty of Medicine, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
authors.forename: Yuriko
authors.initials: Y
authors.lastname: Wada
authors.affiliation: Department of Human Pathology, Graduate School and Faculty of Medicine, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
authors.forename: Yoshimi
authors.initials: Y
authors.lastname: Suzuki
authors.affiliation: Department of Human Pathology, Graduate School and Faculty of Medicine, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Ishige
authors.affiliation: Department of Clinical Engineering, School of Health Sciences, Tokyo University of Technology, Tokyo 144-8650, Japan.
authors.forename: Junko
authors.initials: J
authors.lastname: Minami
authors.affiliation: Division of Surgical Pathology, Tokyo Medical and Dental University Hospital, Tokyo 113-8510, Japan.
authors.forename: Takumi
authors.initials: T
authors.lastname: Akashi
authors.affiliation: Department of Human Pathology, Graduate School and Faculty of Medicine, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.|Division of Surgical Pathology, Tokyo Medical and Dental University Hospital, Tokyo 113-8510, Japan.
authors.forename: Yoshinobu
authors.identifier: http://orcid.org/0000-0002-0521-1670
authors.initials: Y
authors.lastname: Eishi
mesh_terms: D000818:Animals; D001343:Autophagy; D000071187:Autophagy-Related Protein 5 / Q000172:deficiency; D004847:Epithelial Cells / Q000166:cytology* / Q000382:microbiology / Q000473:pathology; D016908:Gram-Positive Bacterial Infections / Q000378:metabolism / Q000473:pathology*; D006367:HeLa Cells; D006801:Humans; D052118:Lysosomal-Associated Membrane Protein 1 / Q000378:metabolism; D008264:Macrophages / Q000166:cytology* / Q000382:microbiology / Q000473:pathology; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000382:microbiology / Q000473:pathology; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008954:Models, Biological; D011425:Propionibacterium acnes / Q000472:pathogenicity*; D000067996:RAW 264.7 Cells; D012507:Sarcoidosis / Q000378:metabolism / Q000382:microbiology* / Q000473:pathology; D014617:Vacuoles / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D000071187:Autophagy-Related Protein 5; D052118:Lysosomal-Associated Membrane Protein 1; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins