贵州医科大学学报

2017, v.42;No.202(07) 778-782

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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

杨辉;鲁利香;肖政华;徐春梅;沈慧;侯琼琼;戴家庆;祁兰兰;
YANG Hui;LU lixiang;XIAO Zhenghua;XU Chunmei;SHEN Hui;HOU Qiongqiong;DAI Jiaqing;QI Lanlan;China Academy of Traditional Chinese Medicine Post-doctoral Research Stations;The Second Clinical Medical College of Guiyang Traditional Chinese Medicine university;Guiyang Traditional Chinese Medicine University;

摘要(Abstract):

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 cells of hippocampal neuron were cultivated for five days. The threeair-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 MTT method and inverted microscope to determine the most appropriate time. Results: By the MTT method and inverted microscope,the three-airincubator jointed EBSS liquid lacking oxygen and sugar to build the best time for 2 hours,and the normal 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 deprivation and hypoxia was cultivated for 2 hours.

关键词(KeyWords): 神经元;海马;缺氧缺糖;MTT;模型,细胞
neurons;hippocamp;oxygen-glucose deprivation;MTT;model;cell

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金资助项目(81260601);; 贵州省科学技术基金项目[黔科合J字(2008)2174号];; 贵州省科技计划基金资助项目[黔科合中药字(2011)LKZ7003号];; 贵州省教育厅自然科学研究项目(黔教科2008031号);; 贵州省中医药管理局资助课题项目(QZYY2010-48)

作者(Author): 杨辉;鲁利香;肖政华;徐春梅;沈慧;侯琼琼;戴家庆;祁兰兰;
YANG Hui;LU lixiang;XIAO Zhenghua;XU Chunmei;SHEN Hui;HOU Qiongqiong;DAI Jiaqing;QI Lanlan;China Academy of Traditional Chinese Medicine Post-doctoral Research Stations;The Second Clinical Medical College of Guiyang Traditional Chinese Medicine university;Guiyang Traditional Chinese Medicine University;

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DOI: 10.19367/j.cnki.1000-2707.2017.07.008

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