以多药耐药基因为靶标的siRNA对耐药肿瘤细胞药物敏感性和凋亡的影响  

Study on the change of drug resistance and apoptosis of carcinoma cells after transfecting siRNA targeting multidrug resistance related protein

作　　者：胡海燕[1] 郭坤元[1] 刘革修[2] 马秀君[2] 姜铧[2] 马文丽[1] 

机构地区：[1]南方医科大学基因工程研究所,广东广州510515 [2]暨南大学医学院,广东广州510632

出　　处：《暨南大学学报（自然科学与医学版）》2007年第6期550-553,共4页Journal of Jinan University(Natural Science & Medicine Edition)

基　　金：博士后基金项目(2006039793)

摘　　要：目的:研究应用以多药耐药相关蛋白(MRP)基因为靶标的siRNA(small interference RNA)抑制相应蛋白表达后,耐药肿瘤细胞药物敏感性和凋亡率的改变。方法:通过脂质体将以MRP基因为靶标的siRNA(100μmol/L)转入柔红霉素(DNR)0.8、1.6、3.2、6.4、12.8μg/mL处理的肺癌耐药细胞株SW1573/R20;阿霉素(ADM)1.6、3.2、6.4、12.8、25.6μg/mL处理的白血病耐药细胞株K562/ADM;顺铂(DDP)0.125、0.25、0.5、1.0、2.0、4.0μmol/mL处理的鼻咽癌耐药细胞株CNE2/DDP;以单纯化疗处理组和未处理组为对照。于转染后24、48、72h,用MTT法检测各组细胞生长抑制率,算出各细胞IC50。转染siRNA联合IC50剂量化疗处理6 h后流式细胞仪检测各细胞凋亡率和死亡率,24 h后RT-PCR检测相应基因mRNA表达水平,48 h后检测MRP蛋白表达率。结果:以MRP为靶标的siRNA明显抑制各肿瘤细胞靶基因蛋白和mRNA的表达,与未处理组相比均有统计学差异(P<0.05);转染以MRP为靶标的siRNA后,耐药肿瘤细胞对化疗药物敏感性明显增强,IC50明显降低,细胞凋亡率明显增加,与单纯化疗组相比有统计学差异(P<0.05)。结论:以MRP基因为靶标的siRNA,通过降低靶基因mRNA和蛋白表达,明显增加耐药肿瘤细胞药物敏感性和凋亡率。Aim：To study the regulation of drug resistance and the apoptosis ratio of drug resistance carcinoma cells after transfer siRNA targeting MRP. Methods： We transfected siRNA, targeting MRP （100 μmol/L）, by lipofectamine into SW1575/R20（pretreated by DNR at 0. 8,1.6,3. 2,6.4,12. 8 μg/ mL）, K562/ADM（pretreated by ADM at 1.6,3. 2,6. 4,12. 8,25.6 μg/mL）, and CNE2/DDP（ pretreated by DDP at 0. 125,0. 25,0. 5,1.0,2.0,4.0 μmoL/mL）. The negative control group is cell without any treatment, the positive control group is cell only treated by chemicaltherapy. After transfected for 6 hours, we added 20% FBS serum OPTI-MEM. The cell＇s viability was detected at 24, 48 and 72 h by MTT method. After transfecting for 24 hours, we collected the cells to detect the mRNA level of MRP by RT-PCR. After transfecting for 48 hours, we collected the cells to detect the transfect ratio, the apoptosis ratio and the expression ratio of MRP by FCM. Results： After transfected siRNA targeting MRP and bcl- 2, the level of mRNA and MRP and bcl-2 decrease significantly. Meanwhile the IC50 decreased significantly and apoptosis ratio increased significantly compared with ADM group （ P 〈 0. 05 ）. For the group transfected with siRNA targeting MRP combining with bcl-2, the IC50 is the lowest and the apoptosis ratio is the highest among all groups（P 〈0. 05）. Conclusion：As we transfect siRNA targeting MRP and bcl- 2, the drug sensitivity and apoptosis increased with level of MRP and bcl-2 decreasing.

关 键 词：BCL-2基因 SIRNA 多药耐药相关蛋白基因 

分 类 号：R733.3[医药卫生—肿瘤]