title: Proteomic analysis of primary cultured rat cortical neurons in chemical ischemia.
abstract: To elucidate the molecular events involved in early ischemic neuronal death, we performed two-dimensional proteome profiling of primary cultures of rat cortical neurons following chemical ischemia induced by the administration of sodium azide under glucose-free conditions. Using a lactic dehydrogenase assay and Western blot analysis of dephosporylation of the voltage-gated potassium channel Kv2.1, we determined duration of chemical ischemia of 2 h to be the relevant time-point for early ischemic neuronal death. Sixty-one proteins were differentially expressed, and 26 different proteins were identified by MALDI-TOF with Mascot database searching. The proteome data indicated that chemical ischemia altered the expression of 20 proteins that are involved in stress response/chaperone, brain development, cytoskeletal/structural proteins, metabolic enzymes, and calcium ion homeostasis. Western blotting and immunocytochemical studies of the 6-most functionally significant proteins showed that, in the ischemia-treated group, the expression of glucose-related protein 78, heat shock protein 90 alpha, and α-enolase was significantly increased, while the expression of inositol triphosphate receptor 1 and ATP synthase beta subunit was decreased. In addition, the expression of dihydropyrimidinase-like 3 showed a truncated pattern in the ischemia group. The changes in the expression of these proteins might be significant indicators of early ischemic neuronal death.
authors.affiliation: Department of Physiology, Biomedical Science Institute and Medical Research Center, School of Medicine, Kyung Hee University, Seoul, 130-701, South Korea.
authors.forename: Jung-Woo
authors.initials: JW
authors.lastname: Seo
authors.forename: Younghoon
authors.initials: Y
authors.lastname: Kim
authors.forename: Jinyoung
authors.initials: J
authors.lastname: Hur
authors.forename: Kang-Sik
authors.initials: KS
authors.lastname: Park
authors.forename: Young-Wuk
authors.initials: YW
authors.lastname: Cho
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D007511:Ischemia / Q000139:chemically induced*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D010766:Phosphorylation; D040901:Proteomics; D051381:Rats; D019810:Sodium Azide / Q000008:administration & dosage*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D009419:Nerve Tissue Proteins; D019810:Sodium Azide