CD/5-FC自杀基因治疗系统体外杀伤恶性人脑胶质瘤细胞作用的研究  

Effects of CD/5-FC suicide gene therapy system on human malignant glioma cells in vitro

作　　者：吕胜青[1] 刘运生[1] 杨辉[2] 陈祖林[2] 黄河[2] 

机构地区：[1]中南大学湘雅医院神经外科,长沙410008 [2]第三军医大学新桥医院神经外科,重庆400037

出　　处：《中南大学学报（医学版）》2004年第2期174-176,180,共4页Journal of Central South University ：Medical Science

摘　　要：目的 :探讨胞嘧啶脱氨酶 / 5 氟胞嘧啶 (CD/ 5 FC)自杀基因治疗系统对恶性人脑胶质瘤细胞的杀伤作用。方法 :采用Lipofectamine2 0 0 0脂质体介导法将CD基因转染U2 5 1恶性人脑胶质瘤细胞 ,G4 1 8筛选获得抗性克隆 (取名为U2 5 1 /CD细胞 ) ,使用不同浓度的 5 FC作用于U2 5 1 /CD细胞 ,MTT法测定活性细胞比率。采用高效液相色谱法 (HPLC)检测 5 FC培养液内 5 FU的浓度。结果 :U2 5 1细胞获得了质粒的成功转染。基因转染使G4 1 8抗性细胞 (U2 5 1 /CD细胞 )对 5 FC高度敏感。未转染的U2 5 1细胞对 5 FC不敏感 ,IC50 约为 6 5 0 0 μmol/L ,而转染基因后IC50 约为 1 0 μmol/L。并且加入不同浓度的 5 FC后 ,U2 5 1 /CD细胞培养液内均能检测到 5 FU。结论 :5 FC对CD基因修饰U2 5 1细胞具有杀伤作用 ;Objective To investigate the killing effects of cytosine deaminase/5 fluorocytosine (CD/5 FC) suicide gene therapy system on human malignant glioma cells in vitro.Methods The pCMVCD plasmid was transfected into the U251 human malignant glioma cells using Lipofectamine 2000 mediated method. Resistant clones (named U251/CD cells) were isolated by screening with G418 presence. U251/CD cells were incubated with 5 FC in different concentrations to determine the number of viability cells (or cytotoxicity assay),and measured by the 3 (4,5 dimethylthiazol 2 yl) 2,5 diphenyltetrazolium bromide (MTT) assay. The concentrations of 5 fluorouracil (5 FU) in the media were measured by the high performance liquid chromatography (HPLC) analysis.Results The untreated U251 cells were insensitive to 5 FC, with the IC 50 about 6 500 μmol/L. After the transfection, the IC 50 was dramatically reduced to about 10 μmol/L. Therefore, gene transfection made G418 resistant clones (U251/CD cells) highly sensitive to 5 FC. HPLC analysis showed that 5 FU was detected in U251/CD cell medium. Conclusion The killing effects of U251 cells modified by CD gene were induced by 5 FC, and the CD/5 FC system was feasible to treat glioma.

关 键 词：胞嘧啶脱氨酶 5-氟胞嘧啶 基因治疗 恶性脑胶质瘤 细胞杀伤作用 

分 类 号：R739.41[医药卫生—肿瘤]