反义RNA逆转耐药肝癌基因工程细胞株多药耐药性的实验研究  

Reversing effect of multidrug resistance in hepatocellular carcinoma engineering cell line with antisence RNA

作　　者：李波[1] 叶田[2] 李德华 陈永兵[4] 张孟瑜[1] 贺凯[1] 冯春红[1] 陈星[1] 徐松波[1] 倪建彬[1] 罗亮[1] 夏先明[1] 

机构地区：[1]泸州医学院附属第一医院肝胆外科,四川泸州646000 [2]泸州医学院附属第二医院中医外科,四川泸州646000 [3]成都地奥制药集团有限公司基因工程药物研究室,四川成都610041 [4]首都医科大学附属佑安医院肝胆外科,北京100069

出　　处：《中国现代医学杂志》2009年第19期2907-2910,2914,共5页China Journal of Modern Medicine

基　　金：泸州医学院和四川省科技厅科研资助项目(No:05JY029-146)

摘　　要：目的通过重组腺病毒表达出mdr1基因的反义RNA,观察反义RNA对人肝癌基因工程细胞株HepG2/R多药耐药性的逆转效应。方法利用真核表达载体构建可以稳定表达mdr1基因和P糖蛋白的肝癌基因工程细胞株HepG2/R,通过腺病毒载体AdEasy系统生成重组腺病毒pAd Easy-GFP-ASmdr1,将此病毒感染HepG2/R,用RT-PCR检测mdr1 mRNA的表达强度,流式细胞仪检测细胞膜P-糖蛋白的表达和柔红霉素的蓄积率,HepG2/R细胞对阿霉素的耐药性用MTT法检测。结果HepG2/R细胞感染重组腺病毒后,RT-PCR检测表明mdr1 mRNA表达下降,流式细胞仪检测证实P-gp表达下降和柔红霉素的蓄积率增加,MTT法表明HepG2/R细胞对阿霉素的IC50从25μg/mL下降到3μg/mL。结论反义RNA通过抑制mdr1mRNA和P-gp的表达。[ Objective ] To investigate the reversal effect of multidrug resistance with antisense RNA of mdrl gene delivered by recombinant adenoviruses in human engineering HCC cell line HepG2/R. [ Methods ] The recombinant adenoviruse was transfected into the engineering cell line HepG2/R. In order to investgate the reversal of the multidrug resistance phenotype, the expression of mdrl mRNA was measured by RT-PCR, the production of P-glyco- protein and the accumulation of the daunorubicin （DNR） was determinated by flow eytometry. The sensitivitie of adriamyein （ADM） for HepG2/R cells was examined by MTT analysis. [ Results ] Compared with the parental HepG2 cells expressing low-level mdrl mRNA and P-glycoprotein, engineering cell line HepG2/R was only stably expressing mdrl gene and P-glyeoprotion. The transfeetion of antisence RNA into HepG2/R cells resulted in decreases of mdrl mRNA and P-glycoprotein levels. The sensitivity of transfected HepG2/R cells to ADM was reduced from 25μg/mL to 3μg/mL in IC50 level. The DNR accumulation was increased in transfected HepG2/R cells. [Conclusion] This study demonstrates that mdrl antisense RNA can increase the sensitivities of HepG2/R cells to anticaneer drug by decreasing the expression of the mdrl gene and inhibiting P-glyeoprotein expression.

关 键 词：多药耐药性 多药耐药基因 肝癌细胞株 工程细胞株 

分 类 号：R735.7[医药卫生—肿瘤]