摘要
目的建立裸鼹鼠星形胶质细胞体外培养方法,观察不同氧浓度对裸鼹鼠星形胶质细胞增殖及生长状态的影响,寻找最适宜体外裸鼹鼠细胞增殖和生长的氧浓度。方法利用机械法辅以化学法分离新生裸鼹鼠脑内胶质细胞,利用差速贴壁法进行星形胶质细胞纯化,通过设置3%、5%、10%以及20%等不同氧浓度环境,利用CCK8染色法分析不同氧浓度下裸鼹鼠星形胶质细胞增殖速率,利用免疫细胞化学染色法观察不同氧浓度下裸鼹鼠细胞胞体伸长突起的状况从而进一步判断不同氧浓度下裸鼹鼠星形胶质细胞生长状态。结果利用机械及化学法分离裸鼹鼠胶质细胞并结合差速贴壁法能够成功建立体外裸鼹鼠星形胶质细胞模型。不同氧浓度环境下裸鼹鼠生长速率不同,随着氧浓度的升高,裸鼹鼠进入快速生长期所用的时间越短。氧浓度为20%时,裸鼹鼠星形胶质细胞较小,突起少,状态不佳,但是10%以下的氧浓度环境中,裸鼹鼠能保持较多的细胞突起,状态较好。结论机械及化学法相结合,合并差速贴壁法能够成功建立裸鼹鼠星形胶质细胞体外培养模型,10%氧浓度下既可以保证裸鼹鼠星形胶质细胞适宜的生长速率又能保证其处于良好的生长状态。
Objective The aim of this study is to establish the in vitro culture method for primary naked mole rat astrocytes,and observe effects of different oxygenous concentration to naked mole rat astrocyte proliferation and growth status,and find the most convenient concentration of O2 for naked mole rat culture in vitro. Method Astrocytes isolated from new-born naked mole rat brain with the aid of mechanical and enzyme digestion method were purified by means of shaking-off method. For further investigation,naked mole rat astrocytes incubated in a series of different O2 concentration of 3%,5%,10%,and 20% were processed for the analysis of proliferation with CCK8 staining and growth status with immunocytochemistry according to the morphology of the cells at scheduled time-points. Result Mechanical and enzyme digestion method combined with shaking-off method were feasible for naked mole rat astrocyte in vitro culture. Naked mole rat astrocyte proliferation in vitro mainly depends on the concentration of O2. As the incensement of O2 concentration,naked mole rat astrocyte grow faster. For morphological investigation,under 20% O2 concentration,soma of naked mole rat astrocytes shrink and possesses fewer cellular processes,While,when O2 concentration was below 10%,naked mole rat astrocytes possesses more processes,Conclusion Mechanical and enzyme digestion method, combined with shaking-off method were appropriate for naked mole rat astrocyte culture in vitro,and 10% O2 concentration will ensure the proliferation and growth status in high quality of the cells.
出处
《实验动物科学》
2016年第3期20-26,共7页
Laboratory Animal Science
基金
国家科技部支撑计划(No.2015BAI09B02)
2014年度军队实验动物专项科研课题面上项目(No.SYDW[2014]005号)联合资助
关键词
裸鼹鼠
星形胶质细胞
增殖
生长状态
naked mole rat
astrocyte
proliferation
growth status
