title: Autophagy Activation by Rapamycin Before Hypoxia-Reoxygenation Reduces Endoplasmic Reticulum Stress in Alveolar Epithelial Cells.
abstract: BACKGROUND
  To determine potential effects of autophagy activation on hypoxia-reoxygenation (H/R) induced damage of a rat alveolar epithelial cell line.
  
  
  METHODS
  CCL149 cells were subjected to autophagy agonist (rapamycin, Rap), autophagy inhibitor (3-methyladenine, 3-MA) or PBS for 1 h before H/R treatment for 2 h, 4 h and 6 h. The optimal concentration of Rap (150 nM, 200 nM and 250 nM) or 3-MA (5 mM, 10 mM and 15 mM) was obtained from MTT assay. Autophagy was determined by fluorescence microscopy of eRFP-LC3 positive cells, transmission electron microscopy of autophagosome, western blot of LC3, AMPK, Beclin-1, HDAC6 and p62 proteins. Endoplasmatic reticulum stress was indicated by detecting expressions of BIP, XBP-1 and CHOP via western blot.
  
  
  RESULTS
  Rap at concentration of 250 nM before H/R increased the autophagy formation with more eRFP-LC3 positive cells and higher expressions of LC3-II, Beclin-1, HDAC6 and p62, but lower expressions of BIP, XBP-1 and CHOP in H/R treated CCL149. This effect seemed to be still obvious after H/R exposure for 6 h. The contrary results were obtained by treatment with 5 mM 3-MA.
  
  
  CONCLUSIONS
  Rap might be a promising agent before mechanical ventilation or reperfusion to prevent re-damage in hypoxia related lung diseases.
authors.forename: Tao
authors.initials: T
authors.lastname: Fan
authors.forename: Lei
authors.initials: L
authors.lastname: Chen
authors.forename: Zhixin
authors.initials: Z
authors.lastname: Huang
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.forename: Boyou
authors.initials: B
authors.lastname: Zhang
authors.forename: Yao
authors.initials: Y
authors.lastname: Xu
authors.forename: Zhangfan
authors.initials: Z
authors.lastname: Mao
authors.forename: Hao
authors.initials: H
authors.lastname: Hu
authors.forename: Qing
authors.initials: Q
authors.lastname: Geng
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000633:toxicity; D056809:Alveolar Epithelial Cells; D000818:Animals; D001343:Autophagy / Q000187:drug effects*; D000071186:Beclin-1 / Q000235:genetics / Q000378:metabolism; D015687:Cell Hypoxia; D002460:Cell Line; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects*; D007150:Immunohistochemistry; D046529:Microscopy, Electron, Transmission; D008856:Microscopy, Fluorescence; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism / Q000494:pharmacology; D051381:Rats; D020123:Sirolimus / Q000633:toxicity*; D051742:Transcription Factor CHOP / Q000235:genetics / Q000378:metabolism; D000071717:X-Box Binding Protein 1 / Q000235:genetics / Q000378:metabolism
chemical_list: D000225:Adenine; D000071186:Beclin-1; D008869:Microtubule-Associated Proteins; D010100:Oxygen; D020123:Sirolimus; D051742:Transcription Factor CHOP; D000071717:X-Box Binding Protein 1; C497251:Ddit3 protein, rat; C526109:Anxa3 protein, rat; C000605545:Xbp1 protein, rat; C025946:3-methyladenine
keywords: Alveolar hypoxia-reoxygenation; Autophagy; Endoplasmatic reticulum stress; Rapamycin; Unfolded protein response