bcl-2基因转染PC12细胞及其神经保护作用的研究  

THE STUDY OF NEUROPROTECTION OF bcl-2 GENE BY TRANSFECTING IT INTO PC12 CELL LINE

作　　者：吴永青[1] 刘彦文[2] 邱鹏新[3] 张清秀[4] 葛坚 

机构地区：[1]南京大学医学院附属鼓楼医院眼科,南京210008 [2]广州中山大学动物实验中心,广州510089 [3]广州中山大学药理学教研室,广州510089 [4]广州中山大学生物化学教研室,广州510089 [5]中山眼科中心,广州510060

出　　处：《解剖学报》2002年第4期346-349,共4页Acta Anatomica Sinica

基　　金：国家自然科学基金资助项目 ( 39770 789)

摘　　要：目的　研究基因bcl 2对神经元的生物活性作用。　方法　构建真核表达载体pcDNA3 bcl 2 ,采用脂质体介导将重组质粒导入PC12细胞 ,Westernblot和原位免疫组织化学检测外源基因表达 ;10 μmol L、5 0 μmol L和10 0 μmol L顺铂处理转染重组质粒的实验A组细胞 ,同样条件处理转染空质粒的对照B组细胞 ,72h后比较两者存活的细胞数 ;流式细胞仪检测分析两者的细胞周期指数。　结果　成功构建了真核表达载体pcDNA3 bcl 2 ,并用脂质体介导的方法获得了稳定表达Bcl 2的细胞克隆 ;10 μmol L、5 0 μmol L和 10 0 μmol L顺铂处理后 ,实验A组存活的细胞数分别为 2 76± 13、185± 11和 10 8± 10 ,而对照B组中存活的细胞数分别为 10 0± 9、12± 3和 2± 2 ,两者相比有显著性差异 ;流式细胞仪检测显示 ,实验A组S期细胞所占百分比为 8 81% ,比对照B组 2 5 79%明显减少。　结论　bcl 2能够拮抗顺铂对神经元的损害作用 ,促进神经细胞存活 。Objective To investigate the biological effects of bcl-2 gene on neurons. Methods The recombinant expression plasmid pc DNA3-bcl-2 was constructed from pSFFV-bcl-2,then it was introduced into PC12 cell line by liposome method.Western blotting and immunohistochemistry in situ were applied to exam the exogenous gene expression.The two groups of cells(Group A,PC12 transfected by pcDNA3-bcl-2 and Group B,PC12 transfected by pcDNA3) were exposed to cisplatin with the concentration of 10*!μmol/L,50*!μmol/L, and 100*!μmol/L 72 hours later,the survival cells were estimated.Cell cycle indexes between these two groups were also studied by FCM. Results The recombinant expression plasmid pcDNA 3-bcl-2 was constructed successfully and PC12 cell line transfected by the plasmid could express Bcl-2 protein effectively.Compared with the control(25 79%),there was a significant decrease of cells from the S phase in PC12 with bcl-2 gene(8.81%).After exposed with 10*!μmol/L,50*!μmol/L,and 100*!μmol/L cisplatin,the surviving cells in group A were 276±13,185±11 and 108±10 respectively,which increased much more than in group B while they were 100±9,12±3 and 2±2 accordingly.Conclusions bcl-2 can protect PC12 cells against cytotoxic insults of cisplatin,and it suggested that it might act via cell cycle controlling.

关 键 词：BCL-2基因 PC12细胞 基因表达 神经保护 神经系统疾病 基因转染 

分 类 号：R74[医药卫生—神经病学与精神病学]