摘要
目的探讨短发夹RNA(shRNA)靶向沉默DLL4基因对MCF-7细胞凋亡的诱导及对化疗药物多西他赛的增敏作用。方法针对DLL4基因的序列设计具有特异性的shRNA,经脂质体转染MCF-7细胞,作为实验组,空脂质体转染的MCF-7细胞为脂质体组,未做任何处理的MCF-7细胞为对照组。采用免疫细胞化学方法检测3组细胞DLL4蛋白的表达情况;采用流式细胞仪检测3组细胞凋亡率及细胞周期改变;采用噻唑蓝(methyl-thiazoyl-tetrazolium bromide,MTT)法测定3组细胞的增殖情况及对多西他赛的敏感性。结果实验组平均光密度值及阳性面积率均低于对照组和脂质体组(P<0.01);实验组细胞在24 h、48 h及72 h时间点A值均低于对照组和脂质体组(P<0.01);转染后24 h、48 h、72 h及96 h,实验组细胞凋亡率高于对照组和脂质体组(P<0.05);RNA干扰(RNAi)沉默DLL4基因后,实验组G2/M期细胞比例高于对照组和脂质体组(P<0.05);实验组的IC50值较对照组和脂质体组低(P<0.05)。结论 RNAi技术抑制DLL4基因的表达能够抑制MCF-7细胞的增殖、诱导细胞凋亡及增强细胞对多西他赛作用的敏感性。DLL4可能会成为乳腺癌治疗的重要靶点。
Objective To explore effect of DLL4 gene in MCF-7 cells of human breast cancer which was inhibitted by short hair in RNA（shRNA） on inducing apoptosis and chemosensitivity to docetaxel.Methods Specific shRNA was designed in accordance with DLL4 gene and transfected into MCF-7 cells of human breast cancer with liposomal（Lip-shRNA group）,MCF-7 cells transfected with only liposomal as Lip group,and control group without any treatment.Expressions of DLL4 protein in 3 groups were detected by immunohistochemical method,apoptosis and cell cycle distribution were examined by flow cytometry（FCM）.Proliferations and sensitivity of MCF-7 cells to docetaxel in 3 groups were determined by methylthiazoyl-tetrazolium bromide（MTT）.Results The averages optical density value and rate of positive area of Lip-shRNA group were significantly lower than that of other 2 groups（P0.01）.The levels of A value at 24 h,48 h,and 72 h in Lip-shRNA group were significantly lower than that of other 2 groups（P0.01）.Rates of cell apoptosis at 24 h,48 h,72 h and 96 h in Lip-shRNA group were significantly higher than that of other 2 goups（P0.05）, and ratio of G2/M was higher too（P0.05）.IC50 of Lip-shRNA group to docetaxel was significantly lower than that of other 2 groups（P0.05）.Conclusions The RNA interference technology can effectively block the expression of DLL4 gene.Inhibition of DLL4/Notch signaling pathway can lead to proliferation inhibition of cancer cell and a concomitant increase in cells undergoing apoptosis,and can enhance the cell sensitivity to docetaxel.DLL4 may be an important target for therapeutic approach of breast cancer.
出处
《中国普外基础与临床杂志》
CAS
2013年第1期54-59,共6页
Chinese Journal of Bases and Clinics In General Surgery
