title: Activation of the mitochondrial caspase pathway and subsequent calpain activation in monkey RPE cells cultured under zinc depletion.
abstract: OBJECTIVE
  Decreased zinc levels in the macula are reported in patients with age-related macular degeneration, and the zinc chelator N,N,N',N'-tetrakis (2- pyridylmethyl) ethylenediamine) (TPEN) causes death of human retinal pigment epithelial (RPE) cells. The purpose of the present study was to investigate signal transduction pathways during cell death initiated by TPEN, using monkey RPE cells.
  
  
  METHODS
  RPE cells were cultured with TPEN. Activation of calpains and caspases, and proteolysis of their substrates were detected by immunoblotting. Incubation of calpain inhibitor SNJ-1945 or caspase inhibitor z-VAD-fmk was used to confirm activation of specific proteases.
  
  
  RESULTS
  TPEN caused a time-dependent decrease in viable RPE cells. Cell death was accompanied by activation of calpain-1, caspase-9, and caspase-3. SNJ-1945 inhibited calpain activation and slightly inhibited caspase-9 activation. z-VAD-fmk inhibited caspases and calpain-1 activation. TPEN did not activate caspase-12.
  
  
  CONCLUSIONS
  Relative zinc deficiency in RPE cells causes activation of cytosolic calpain and mitochondrial caspase pathways without ER stress.
authors.affiliation: 1] Senju Laboratory of Ocular Sciences, Senju Pharmaceutical Corporation Limited, Portland, OR, USA [2] Department of Integrative Biosciences, Oregon Health & Science University, Portland, OR, USA.
authors.forename: E
authors.initials: E
authors.lastname: Nakajima
authors.affiliation: 1] Senju Laboratory of Ocular Sciences, Senju Pharmaceutical Corporation Limited, Portland, OR, USA [2] Department of Integrative Biosciences, Oregon Health & Science University, Portland, OR, USA.
authors.forename: K B
authors.initials: KB
authors.lastname: Hammond
authors.affiliation: Department of Integrative Biosciences, Oregon Health & Science University, Portland, OR, USA.
authors.forename: T R
authors.initials: TR
authors.lastname: Shearer
authors.affiliation: 1] Senju Laboratory of Ocular Sciences, Senju Pharmaceutical Corporation Limited, Portland, OR, USA [2] Department of Integrative Biosciences, Oregon Health & Science University, Portland, OR, USA.
authors.forename: M
authors.initials: M
authors.lastname: Azuma
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002154:Calpain / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002219:Carbamates / Q000494:pharmacology; D061945:Caspase Inhibitors / Q000494:pharmacology; D020169:Caspases / Q000378:metabolism*; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D004789:Enzyme Activation; D005029:Ethylenediamines / Q000494:pharmacology; D008253:Macaca mulatta; D008858:Microscopy, Phase-Contrast; D008928:Mitochondria / Q000201:enzymology*; D009842:Oligopeptides / Q000494:pharmacology; D055213:Retinal Pigment Epithelium / Q000378:metabolism* / Q000473:pathology; D015398:Signal Transduction / Q000502:physiology; D013997:Time Factors; D015032:Zinc / Q000172:deficiency*
chemical_list: C515164:((1S)-1-((((1S)-1-benzyl-3-cyclopropylamino-2,3-di-oxopropyl)amino)carbonyl)-3-methylbutyl)carbamic acid 5-methoxy-3-oxapentyl ester; D002219:Carbamates; D061945:Caspase Inhibitors; D002614:Chelating Agents; D005029:Ethylenediamines; D009842:Oligopeptides; C517629:benzyloxycarbonyl-valyl-alanyl-aspartic acid; D002154:Calpain; D020169:Caspases; D015032:Zinc; C044387:N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine