Culture of glial lineage progenitor cells derived from white matter and establishment of its oxygen-glucose deprivation models

Abstract

Objective To in vitro culture glial lineage progenitor cells derived from periventricular white matter in neonatal rats,and establish oxygen-glucose deprivation models of progenitor cells to further explore the mechanism of neurogenesis in the ischemic white matter injury.Methods The progenitor cells were isolated and cultured from the white matter of 3-d-old neonatal rats.Cells were passaged every 3 or 4 d,and were further induced differentiation.The primary and passaged neurospheres and their differentiated cells were respectively identified by immunocytochemical stanning of NG2 (Neurospheres marker) and O4 (oligodendrocyte precursor marker).The third-or fourth-generation progenitor cells were used to establish the ischemic model of oxygen-glucose deprivation (OGD).The influence of different OGD periods (30,45 and 60 min,2 and 3 h) in the survival rates of progenitor cells was assessed 24 h after success of model making by both CCK-8 and Hoechst 33342/PI stainning.Results The progenitor cells obtained from the white matter had the capacity of forming neurospheres and reproduction which showed NG2 immunoreactive positivity,and could be induced differentiation into O4-positive oligodendrocyte precursors.No apoptotic or necrotic cells were observed in normal cultured NG2-positive progenitor cells.The apoptotic and necrotic cells tended to increase significantly with the extension of OGD period gradually.The survival rates of progenitor cells in turn were (85.94±3.06) ％ for 30 min OGD,(62.68±2.66) ％ for 45 min OGD,(45.09±2.24) ％ for 60 min OGD,(36.70±2.84) ％ for 2 h OGD and (22.01±3.00) ％ for 3 h OGD,respectively,with significant differences between each two groups (P＜0.05).Conclusion The oxygen-glucose deprivation models of glial lineage progenitor cells derived from periventricular white matter of neonatal rats are successfully established,which may provide the foundation for further exploring the mechanism of neurogenesis in the ischemic white matter injury. Key words: Glial lineage progenitor cell; Cell culture; Oxygen-glucose deprivation; Neurogenesis