title: Ischemic preconditioning enhances autophagy but suppresses autophagic cell death in rat spinal neurons following ischemia-reperfusion.
abstract: Autophagy serves to eliminate damaged proteins and organelles under normal physiological conditions and can be accelerated by pathological stress, possibly as a cytoprotective mechanism. Brief periods of ischemia (ischemic preconditioning or IPC) can reduce neuronal death in response to subsequent severe ischemic insults. Ischemic preconditioning also induces autophagy, but the contribution of autophagy to IPC-associated neuroprotection remains unclear. We investigated the contribution of autophagy to IPC-mediated neuroprotection in rats subjected to ischemic spinal cord injury. Fifty adult rats were randomly assigned to either (1) a sham group receiving anesthesia and surgical preparation (n=5), (2) an ischemia/reperfusion (I/R) group (n=20) subjected to 0.5 h ischemia followed by 3, 6, 12, or 24 h reperfusion, (3) an IPC group receiving three cycles of 5 min ischemia followed by 5 min of reperfusion (n=5), or (4) an IPC+I/R group (n=20). Hematoxylin-eosin (HE) and immunohistochemical staining were performed to evaluate spinal neuron survival in the four treatment groups. Autophagic activity was investigated by electron microscopy and by immunohistochemical and Western blot analyses of the autophagosome marker LC3-II and the autophagy-associated BH3 protein Beclin-1. Changes in Bcl-2/Beclin-1 complex association and Bcl-2 phosphorylation (p-Bcl-2) were examined by co-immunoprecipitation and Western blot analyses. In the I/R group, LC3-II was significantly elevated after 3h of reperfusion, but declined significantly by 24 h. At 24 h, I/R rats exhibited extensive spinal damage and decreased neuronal survival. In the IPC+IR group, neuronal death was reduced and expression of LC3-II sustained throughout the 24 h reperfusion period. In the I/R group, expression of (inactive) p-Bcl-2(Ser70) was increased significantly during reperfusion and was accompanied by dissociation of the Bcl-2/Beclin-1 complex and increased Beclin-1 expression. Preconditioning inhibited these changes in p-Bcl-2, Beclin-1, and Bcl-2/Beclin-1 complex expression. Ischemic preconditioning appears to sustain the beneficial effects of autophagic lysosomal degradation during I/R while inhibiting autophagic cell death.
authors.affiliation: Department of Orthopaedics, the First Affiliated Hospital of Nanjing Medical University, Jiangsu 210029, China.
authors.forename: Jin
authors.initials: J
authors.lastname: Fan
authors.affiliation: Department of Orthopaedics, the Affiliated Drum Tower Hospital of Nanjing University Medical School, Jiangsu 210008, China.
authors.forename: Zitao
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Center for Musculoskeletal Research, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA.
authors.forename: Xie
authors.initials: X
authors.lastname: Chao
authors.affiliation: Xi Shan People׳s Hospital, WuXi, Jiangsu 214194, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Gu
authors.affiliation: Department of Orthopaedics, the First Affiliated Hospital of Nanjing Medical University, Jiangsu 210029, China.
authors.forename: Weihua
authors.initials: W
authors.lastname: Cai
authors.affiliation: Department of Orthopaedics, the First Affiliated Hospital of Nanjing Medical University, Jiangsu 210029, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhou
authors.affiliation: Department of Orthopaedics, the First Affiliated Hospital of Nanjing Medical University, Jiangsu 210029, China. Electronic address: guoyong_yin2005nanjing@yahoo.com.
authors.forename: Guoyong
authors.initials: G
authors.lastname: Yin
authors.affiliation: Department of Orthopaedics, the First Affiliated Hospital of Nanjing Medical University, Jiangsu 210029, China. Electronic address: liqingqing_1999@163.com.
authors.forename: Qingqing
authors.initials: Q
authors.lastname: Li
mesh_terms: D000818:Animals; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D001343:Autophagy / Q000502:physiology*; D000071186:Beclin-1; D016923:Cell Death; D002470:Cell Survival; D004195:Disease Models, Animal; D019194:Ischemic Preconditioning; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009474:Neurons / Q000473:pathology / Q000502:physiology; D010766:Phosphorylation; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D011897:Random Allocation; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000473:pathology / Q000503:physiopathology*; D013116:Spinal Cord / Q000098:blood supply* / Q000473:pathology / Q000503:physiopathology*; D020760:Spinal Cord Ischemia / Q000473:pathology / Q000503:physiopathology*; D013997:Time Factors
chemical_list: D051017:Apoptosis Regulatory Proteins; D000071186:Beclin-1; D008869:Microtubule-Associated Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; C512829:Becn1 protein, rat; C526109:Anxa3 protein, rat
keywords: Autophagic cell death; Autophagy; Ischemia/reperfusion; Ischemic preconditioning (IPC); Spinal cord