title: Ischemia-reperfusion model in rat spinal cord: cell viability and apoptosis signaling study.
abstract: This work aimed at determining the ideal ischemia time in an in vitro ischemia-reperfusion model of spinal cord injury. Rat spinal cord slices were prepared and then exposed or not to oxygen deprivation and low glucose (ODLG) for 30, 45, 60, 75 and 90 minutes. Cell viability was assessed by triphenyltetrazolium (TTC), lactate dehydrogenase (LDH) release, and fluorochrome dyes specific for cell dead (ethidium homodimer) using the apotome system. Glutamate release was enzymatically measured by a fluorescent method. Gene expression of apoptotic factors was assessed by real time RT-PCR. Whereas spinal cord slices exposed to ODLG exhibited mild increase in fluorescence for 30 minutes after the insult, the 45, 60, 75 and 90 minutes caused a 2-fold increase. ODLG exposure for 45, 60, 75 or 90 minutes, glutamate and LDH release were significantly elevated. nNOS mRNA expression was overexpressed for 45 minutes and moderately increased for 60 minutes in ODLG groups. Bax/bcl-xl ratio, caspase 9 and caspase 3 mRNA expressions were significantly increased for 45 minutes of ODLG, but not for 30, 60, 75 and 90 minutes. Results showed that cell viability reduction in the spinal cord was dependent on ischemic time, resulting in glutamate and LDH release. ODLG for 45 minutes was adequate for gene expression evaluation of proteins and proteases involved in apoptosis pathways.
authors.affiliation: Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais, Brasil ; Departamento de Ciências Agrárias e Ambientais, Universidade Estadual de Santa Cruz Ilhéus, Bahia, Brasil.
authors.forename: Mário Sérgio Lima
authors.initials: MS
authors.lastname: de Lavor
authors.affiliation: INCT, Laboratório de Neurociências, Faculdade de Medicina, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais.
authors.forename: Nancy Scardua
authors.initials: NS
authors.lastname: Binda
authors.affiliation: Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais, Brasil.
authors.forename: Fabíola Bono
authors.initials: FB
authors.lastname: Fukushima
authors.affiliation: Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais, Brasil.
authors.forename: Fátima Maria Caetano
authors.initials: FM
authors.lastname: Caldeira
authors.affiliation: INCT, Laboratório de Neurociências, Faculdade de Medicina, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais.
authors.forename: Juliana Figueira
authors.initials: JF
authors.lastname: da Silva
authors.affiliation: Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais, Brasil.
authors.forename: Carla Maria Osório
authors.initials: CM
authors.lastname: Silva
authors.affiliation: Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais, Brasil.
authors.forename: Karen Maciel
authors.initials: KM
authors.lastname: de Oliveira
authors.affiliation: Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais, Brasil.
authors.forename: Bernardo de Caro
authors.initials: Bde C
authors.lastname: Martins
authors.affiliation: Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais, Brasil.
authors.forename: Bruno Benetti Junta
authors.initials: BB
authors.lastname: Torres
authors.affiliation: Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais, Brasil.
authors.forename: Isabel Rodrigues
authors.initials: IR
authors.lastname: Rosado
authors.affiliation: INCT, Laboratório de Neurociências, Faculdade de Medicina, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais.
authors.forename: Renato Santiago
authors.initials: RS
authors.lastname: Gomez
authors.affiliation: INCT, Laboratório de Neurociências, Faculdade de Medicina, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais.
authors.forename: Marcus Vinícius
authors.initials: MV
authors.lastname: Gomez
authors.affiliation: Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária, Universidade Federal de Minas Gerais Belo Horizonte, Minas Gerais, Brasil.
authors.forename: Eliane Gonçalves
authors.initials: EG
authors.lastname: de Melo
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology; D002470:Cell Survival / Q000502:physiology; D004195:Disease Models, Animal; D009924:Organ Culture Techniques; D051381:Rats; D017208:Rats, Wistar; D060888:Real-Time Polymerase Chain Reaction; D015427:Reperfusion Injury / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D020760:Spinal Cord Ischemia / Q000378:metabolism*; D013997:Time Factors
keywords: Apoptosis; cell viability; excitotoxicity; oxygen deprivation low glucose; spinal cord ischemia