title: Autophagy-mediated stress response in motor neurons after hypothermic spinal cord ischemia in rabbits.
abstract: OBJECTIVE
  The development of spinal cord injury is believed to be related to the vulnerability of spinal motor neurons to ischemia. However, the mechanisms underlying this vulnerability have not been fully investigated. Previously, we reported that spinal motor neurons are lost likely due to autophagy and that local hypothermia prevents such spinal motor neuron death. Therefore, we investigated the role of autophagy in normothermic and hypothermic spinal cord ischemia using an immunohistochemical analysis of Beclin 1 (BCLN1; B-cell leukemia 2 protein [Bcl-2] interacting protein), Bcl-2, and γ-aminobutyric acid type-A receptor-associated protein (GABARAP), which are considered autophagy-related proteins.
  
  
  METHODS
  We used rabbit normothermic and hypothermic transient spinal cord ischemia models using a balloon catheter. Neurologic function was assessed according to the Johnson score, and the spinal cord was removed at 8 hours and 1, 2, and 7 days after reperfusion, and morphologic changes were examined using hematoxylin and eosin staining. A Western blot analysis and histochemical study of BCLN1, Bcl-2, and GABARAP, and double-labeled fluorescent immunocytochemical studies were performed.
  
  
  RESULTS
  There were significant differences in the physiologic function between the normothermic model and hypothermic model after the procedure (P < .05). In the normothermic model, most of the motor neurons were selectively lost at 7 days of reperfusion (P < .001 compared with the sham group), and they were preserved in the hypothermic model (P = .574 compared with the sham group). The Western blot analysis revealed that the sustained expression of the autophagy markers, BCLN1 and GABARAP, was observed (P < .001 compared with the sham group) and was associated with neuronal cell death in normothermic ischemic conditions. In hypothermic ischemic conditions, the autophagy inhibitory protein Bcl-2 was powerfully induced (P < .001 compared with the sham group) and was associated with blunted expression of BCLN1 and GABARAP and neuronal cell survival. The double-label fluorescent immunocytochemical study revealed that immunoreactivitiy for BCLN1, Bcl-2, and GABARAP was induced in the same motor neurons.
  
  
  CONCLUSIONS
  These data suggest that the prolonged induction of autophagy might be a potential factor responsible for delayed motor neuron death, and the induction of the autophagy inhibitory protein Bcl-2 using hypothermia might limit autophagy and protect against delayed motor neuron death.
authors.affiliation: Department of Cardiovascular Surgery, Kyushu University Graduate School of Medicine, Fukuoka, Japan. Electronic address: sfujita.heart.med@gmail.com.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Fujita
authors.affiliation: Department of Clinical Research, National Hospital Organization Sendai Medical Center, Sendai, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Sakurai
authors.affiliation: Department of Cardiovascular Surgery, Kyushu University Graduate School of Medicine, Fukuoka, Japan.
authors.forename: Hironori
authors.initials: H
authors.lastname: Baba
authors.affiliation: Department of Neurology, Okayama University Graduate School of Medicine, Okayama, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Abe
authors.affiliation: Department of Cardiovascular Surgery, Kyushu University Graduate School of Medicine, Fukuoka, Japan.
authors.forename: Ryuji
authors.initials: R
authors.lastname: Tominaga
mesh_terms: D000818:Animals; D001343:Autophagy; D004195:Disease Models, Animal; D007036:Hypothermia, Induced; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009046:Motor Neurons / Q000378:metabolism / Q000473:pathology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011817:Rabbits; D020760:Spinal Cord Ischemia / Q000378:metabolism / Q000473:pathology* / Q000503:physiopathology; D013131:Spine / Q000378:metabolism / Q000473:pathology* / Q000503:physiopathology; D013312:Stress, Physiological; D013997:Time Factors
chemical_list: D008869:Microtubule-Associated Proteins; D019253:Proto-Oncogene Proteins c-bcl-2