缺氧缺糖海马神经元模型最佳造模时间的选择

The Best Time of Establishing the Hippocampal Neurons Model Lacking Oxygen and Sugar

目的:建立可靠合理的海马神经元缺氧缺糖(OGD)模型。方法:用培养5 d的海马神经元细胞,采用三气培养箱联合EBSS液,进行OGD造模;于培养0.5、1、2及3 h时收集海马神经元细胞,倒置显微镜下观察海马细胞的形态,并采用MTT法通过检测海马细胞的吸光度(OD)值绘制海马神经元OGD损伤时效曲线,筛选海马神经元OGD最佳造模时间。结果:倒置显微镜下可见随着OGD时间的延长,神经元细胞受损越严重,生长状态越差,胞体从正常的饱满圆润,折光性、立体感强,神经元细胞轴突长,形状粗壮,数量多,与树突相互交集呈网络状;细胞贴壁牢固,变成大部分神经元胞体与自身轴突、树突连接断裂,有的胞体,细胞直接从培养板壁上脱落,脱落的细胞堆积在一起;从海马神经元OGD损伤时效曲线可发现前5 d海马神经元生长迅速,第5天到达高峰期,第5~7天为平台期,第8天后活力稍下降;与OGD 0 h(正常)OD值比较,OGD 0.5、1、2及3 h的OD值减低(P<0.05),表明所采用的OGD造模方法是成功的,OGD 2 h时细胞的损伤程度为正常细胞组30%,可作为正式实验OGD模型。结论:三气培养箱联合EBSS液可进行体外OGD造模,最佳的细胞模型为培养2 h。

Objective： To establish a reliable and reasonable hippocampal neurons model lacking oxygen and sugar. Methods： The ceils of hippocampal neuron were cultivated t＇or five days. The three- air-incubator jointed EBSS liquid to build the model lacking oxygen or sugar and selected half an hour, 1 hour, 2 hours, 3 hours as well as the four periods by MT17 method atld inverted microscope to deler- mine the most appropriate time. Results： By the MTF method and inverted microscope, the three-air- incubator jointed EBSS liquid lacking oxygen and sugar to build the best time for 2 hours, and the, norreal cells were damaged about 30%. Conclusion： The Three-air-incubator jointed EBSS liquid to build the model in vitro, and the best cell model for the study of diseases related to glucose deprivalion and hypoxia was cultivated for 2 hours.