title: The endoplasmic reticulum stress and the HIF-1 signalling pathways are involved in the neuronal damage caused by chemical hypoxia.
abstract: OBJECTIVE
  Hypoxia inducible factor-1 (HIF-1) promotes transitory neuronal survival suggesting that additional mechanisms such as the endoplasmic reticulum (ER) stress might be involved in determining neuronal survival or death. Here, we examined the involvement of ER stress in hypoxia-induced neuronal death and analysed the relationship between ER stress and the HIF-1 pathways.
  
  
  METHODS
  Cultures of rat cortical neurons were exposed to chemical hypoxia induced by 200 μM CoCl2 , and its effect on neuronal viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and counting apoptotic nuclei. Protein levels were determined by Western blot analysis. RT-PCR was performed to analyse the content and the t1/2 of HIF-1α mRNA.
  
  
  RESULTS
  Chemical hypoxia induced neuronal apoptosis in a time-dependent manner and activated the ER stress PRK-like endoplasmic reticulum kinase (PERK)-dependent pathway. At later stages, chemical hypoxia increased the expression of the C/EBP homologous protein (CHOP) and caspase 12 activity. CoCl2 reduced HIF-1α mRNA t1/2 leading to a decrease in HIF-1α mRNA and protein content, simultaneously activating the ER stress PERK-dependent pathway. Salubrinal, a selective inhibitor of phospho-eIF2α phosphatase, protected neurons from chemical hypoxia by reducing CHOP levels and caspase 12 activity, and increasing the t1/2 of HIF-1α mRNA and the levels of HIF-1α protein. Knocking down HIF-1α blocked the neuroprotective effects of salubrinal.
  
  
  CONCLUSIONS
  Neuronal apoptosis induced by chemical hypoxia is a process regulated by HIF-1α stabilization early on and by ER stress activation at later stages. Our data also suggested that HIF-1α levels were regulated by ER stress.
authors.affiliation: Departamento de Ciencias Médicas, Unidad Asociada Neurodeath CSIC-Universidad de Castilla-La Mancha, Albacete, Spain.
authors.forename: Beatriz
authors.initials: B
authors.lastname: López-Hernández
authors.affiliation: Departamento de Ciencias Médicas, Unidad Asociada Neurodeath CSIC-Universidad de Castilla-La Mancha, Albacete, Spain.|CIBERNED, Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: Valentin
authors.initials: V
authors.lastname: Ceña
authors.affiliation: Departamento de Ciencias Médicas, Unidad Asociada Neurodeath CSIC-Universidad de Castilla-La Mancha, Albacete, Spain.|CIBERNED, Instituto de Salud Carlos III, Madrid, Spain.
authors.forename: Inmaculada
authors.initials: I
authors.lastname: Posadas
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D053454:Caspase 12 / Q000187:drug effects / Q000378:metabolism; D002470:Cell Survival / Q000235:genetics; D002478:Cells, Cultured; D002934:Cinnamates / Q000494:pharmacology; D003035:Cobalt / Q000633:toxicity; D059865:Endoplasmic Reticulum Stress / Q000235:genetics*; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism; D055785:Gene Knockdown Techniques; D000860:Hypoxia / Q000139:chemically induced / Q000235:genetics* / Q000378:metabolism; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics* / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D013890:Thiourea / Q000031:analogs & derivatives / Q000494:pharmacology; D051742:Transcription Factor CHOP / Q000187:drug effects / Q000378:metabolism; D019892:eIF-2 Kinase / Q000378:metabolism
chemical_list: D053454:Caspase 12; D002934:Cinnamates; D003035:Cobalt; D015852:Eukaryotic Initiation Factor-2; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D012333:RNA, Messenger; D013890:Thiourea; D051742:Transcription Factor CHOP; D019892:eIF-2 Kinase; C497251:Ddit3 protein, rat; C497444:Hif1a protein, rat; C496827:salubrinal; C506443:Casp12 protein, rat; C018021:cobaltous chloride