论文部分内容阅读
目的:研究以MRP为靶标的siRNA抑制相应基因表达后,对鼻咽癌细胞株CNE2和耐药CNE2/DDP细胞放疗敏感性的增加。方法:通过脂质体将以MRP基因为靶标的siRNA(终浓度100μmol/L)处理CNE2和CNE2/DDP细胞,6 h后分别给予照射剂量1、2、4和6 Gy,另设相同剂量的单纯照射组,以未作任何处理的对数生长期细胞为空白对照。于照射后24、48和72 h,用MTT法检测各组细胞生长抑制率。倒置显微镜下观察细胞生长状况,胰酶消化后透射电镜下检测细胞超微结构。RT-PCR检测各组细胞MRP基因mRNA表达水平,荧光显微镜及流式细胞仪检测细胞MRP蛋白表达率和细胞凋亡率。结果:细胞分别转染以MRP为靶标的siRNA后,MRP基因的mRNA及蛋白表达明显减低,细胞对放疗敏感性明显增高,差异有统计学意义,P<0.01;细胞凋亡率也显著升高,P<0.05;倒置显微镜和透射电子显微镜下见细胞出现凋亡改变。结论:MRP与肿瘤细胞对放疗敏感性密切相关,以其为靶标的siRNA,通过降低靶基因蛋白和mRNA表达,明显增加耐药和亲本肿瘤细胞对放疗的敏感性。
OBJECTIVE: To investigate the radiosensitivity of nasopharyngeal carcinoma cell line CNE2 and drug-resistant CNE2 / DDP cells induced by MRP-targeted siRNA to inhibit the corresponding gene expression. METHODS: CNE2 and CNE2 / DDP cells were treated with siRNA targeting MRP gene (final concentration of 100 μmol / L) by liposomes. After 6 h, irradiation doses of 1, 2, 4 and 6 Gy were given separately and the same dose Simple irradiation group, untreated logarithmic growth phase cells as a blank control. At 24, 48 and 72 h after irradiation, the cell growth inhibition rate of each group was detected by MTT assay. The cell growth was observed under an inverted microscope. The ultrastructure of the cells was observed under transmission electron microscope after trypsin digestion. The expression of MRP mRNA was detected by RT-PCR. The expression of MRP protein and apoptosis rate were detected by fluorescence microscopy and flow cytometry. Results: After transfection of siRNA targeting MRP, the mRNA and protein expression of MRP gene was significantly decreased, and the sensitivity of cells to radiotherapy was significantly increased, the difference was statistically significant, P <0.01; the apoptosis rate was also significantly increased , P <0.05; see inverted microscope and transmission electron microscope under the cell apoptosis. CONCLUSION: MRP is closely related to the radiosensitivity of tumor cells. The siRNA targeted by it can significantly increase the sensitivity of drug-resistant and parental tumor cells to radiotherapy by reducing the expression of target genes and proteins.