title: Control of photoreceptor autophagy after retinal detachment: the switch from survival to death.
abstract: OBJECTIVE
  To examine whether calpain inhibition following retinal detachment would prolong autophagy and result in reduced photoreceptor apoptosis.
  
  
  METHODS
  Retinal detachments were created in Brown-Norway rats by subretinal injection of 1% hyaluronic acid and simulated in vitro by Fas-receptor activation of 661W cells, a cone cell line. Protein levels of LC3 and autophagy-related gene 5 (Atg5), both of which are involved in the creation of the autophagosome, were assayed by Western blot. Calpain 1, the protease responsible for Atg5 cleavage and transitioning photoreceptors from autophagy to apoptosis, activity was monitored by α-spectrin cleavage. Various calpain inhibitors were added either to the subretinal space or cell culture media. Apoptosis was assessed in vitro by caspase-8 activity assays and in vivo via TUNEL assays. Cell counts were assessed in vivo at 2 months following detachment.
  
  
  RESULTS
  Following retinal detachment or Fas-receptor activation of 661W cells, there was an increase in Atg5 and LC3-II that peaked at 3 days and decreased by 7-days postdetachment. Calpain 1 activity level peaked at 7 days and was associated with decreased autophagy. Calpain inhibition led to increased autophagy, a decrease in caspase-8 activation, reduced TUNEL-positive photoreceptors, and increased photoreceptor cell survival.
  
  
  CONCLUSIONS
  Our data suggest that calpain activation, which peaks at 7-days postdetachment, is a key step in triggering photoreceptors to shift from cell survival to death. Prolonging autophagy through calpain inhibition leads to significantly reduced photoreceptor apoptosis and increased cell survival.
authors.affiliation: Department of Ophthalmology and Visual Sciences, W.K. Kellogg Eye Center, University of Michigan, Ann Arbor, Michigan.
authors.forename: Nicholas D
authors.initials: ND
authors.lastname: Chinskey
authors.forename: Qiong-Duon
authors.initials: QD
authors.lastname: Zheng
authors.forename: David N
authors.initials: DN
authors.lastname: Zacks
mesh_terms: D000818:Animals; D017209:Apoptosis; D001343:Autophagy / Q000502:physiology*; D000071187:Autophagy-Related Protein 5; D015153:Blotting, Western; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism; D053181:Caspase 8 / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000502:physiology; D004195:Disease Models, Animal; D004791:Enzyme Inhibitors / Q000494:pharmacology; D020287:In Situ Nick-End Labeling; D018942:Macrolides / Q000494:pharmacology; D008297:Male; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D051381:Rats; D011914:Rats, Inbred BN; D017949:Retinal Cone Photoreceptor Cells / Q000378:metabolism / Q000473:pathology*; D012163:Retinal Detachment / Q000378:metabolism / Q000473:pathology*; D019014:fas Receptor / Q000378:metabolism
chemical_list: D000071187:Autophagy-Related Protein 5; D002154:Calpain; D053181:Caspase 8; D004791:Enzyme Inhibitors; D018942:Macrolides; D008869:Microtubule-Associated Proteins; D011506:Proteins; D019014:fas Receptor; C545865:Atg5 protein, rat; C526109:Anxa3 protein, rat; C057620:bafilomycin A; C505528:Casp8 protein, rat
keywords: apoptosis; autophagy; retinal detachment