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目的探讨超声辐照微泡联合脂质体介导双自杀基因(CD/TK)对MCF-7细胞的体外杀伤作用。方法将培养的MCF-7细胞分为5组:裸质粒组、脂质体组、超声辐照微泡组、超声辐照脂质体组、超声辐照微泡联合脂质体组。转染pEGFP-KDRp-CD/TK质粒于MCF-7细胞,用荧光显微镜及流式细胞仪检测转染效率。转染后,再分为空白对照组、未转染组、转染组;未转染组和转染组又各分为3个亚组,给予前药丙氧鸟苷(GCV)、5-氟胞嘧啶(5-Fc)、GCV+5-Fc。四甲基偶氮唑盐(MTT)法检测双自杀基因对MCF-7细胞的体外杀伤作用。结果超声辐照微泡联合脂质体组绿色荧光蛋白(GFP)表达最多、最强。超声辐照微泡联合脂质体组转染率(39.59%±1.19%)最高(P<0.05)。MTT检测细胞抑制率结果显示,转染组细胞抑制率明显高于未转染组(P<0.01),转染组中联合用药组细胞抑制率明显高于单一用药组(90.77%±2.68%vs 64.75%±2.27%、67.81%±2.43%;P<0.05)。转染组各前药组细胞抑制率均显著高于超声辐照微泡联合脂质体对MCF-7细胞的转染率(P<0.05)。结论超声辐照微泡联合脂质体能明显提高基因转染效率,是较理想的乳腺癌基因治疗策略。
Objective To investigate the killing effect of ultrasound irradiation microbubbles combined with liposome-mediated double suicide genes (CD / TK) on MCF-7 cells in vitro. Methods The cultured MCF-7 cells were divided into five groups: naked plasmid group, liposome group, ultrasound irradiation microbubble group, ultrasound irradiation liposome group, ultrasound irradiation microbubbles combined with liposome group. Transfection of pEGFP-KDRp-CD / TK plasmids in MCF-7 cells, transfection efficiency was detected by fluorescence microscopy and flow cytometry. After transfection, the cells were divided into blank control group, untransfected group and transfected group. The untransfected group and transfected group were divided into three subgroups: gavage (GCV), 5- Flucytosine (5-Fc), GCV + 5-Fc. In vitro killing effect of double suicide genes on MCF-7 cells by MTT assay. Results Ultrasound irradiated microbubbles combined with liposome group, the highest expression of green fluorescent protein (GFP), the strongest. The transfection efficiency of ultrasound irradiation microbubbles combined with liposome group (39.59% ± 1.19%) was the highest (P <0.05). The results of MTT assay showed that the inhibitory rate of transfected cells was significantly higher than that of untransfected cells (P <0.01). The inhibitory rate of transfected cells in combination group was significantly higher than that in single drug-treated group (90.77% ± 2.68% vs 64.75% ± 2.27%, 67.81% ± 2.43%; P <0.05). The transfected group of the prodrug group of cell inhibition rate was significantly higher than the ultrasound irradiation microbubbles combined liposomes on MCF-7 cells transfected rate (P <0.05). Conclusion Ultrasound irradiated microbubbles combined with liposomes can significantly improve gene transfection efficiency, which is an ideal strategy for breast cancer gene therapy.