肝癌SMMC7721细胞株P27RF-Rho mRNA基因沉默对5-Fu药物敏感性的影响

发布时间：2018-03-07 15:10

本文选题：肝肿瘤　切入点：慢病毒　出处：《吉林大学学报(医学版)》2017年02期 　论文类型：期刊论文

【摘要】：目的:探讨肝癌SMMC7721细胞株P27RF-Rho mRNA基因沉默对5-氟脲嘧啶(5-Fu)药物敏感性的影响,为临床晚期肝癌治疗提供理论依据。方法:构建P27RF-Rho RNAi载体,P27RF-Rho基因沉默慢病毒感染SMMC7721肝癌细胞,Western blotting法检测基因沉默效果。SMMC7721细胞分为Scramble-siRNA阴性对照组、5-Fu组、P27RF-Rho-siRNA组和P27RF-Rho-siRNA+5-Fu组。荧光显微镜检测细胞转染效果,Western blottting法检测RNAi基因沉默效率,MTT法检测各组细胞生长曲线,划痕实验检测细胞迁移能力,Transwell小室检测细胞侵袭能力,Western blotting法检测各组细胞中肿瘤相关蛋白P27及RhoC表达。结果:成功构建P27RF-Rho RNAi慢病毒载体。Western blotting法,P27RF-Rho-siRNA组细胞中P27RF-Rho蛋白表达水平明显低于5-Fu组和Scramble-siRNA组(P0.05)。与其他3组比较,P27RF-Rho-siRNA+5-Fu组细胞生长速度降低(P0.05);划痕实验中P27RF-Rho-siRNA+5-Fu组细胞迁移能力明显减低(P0.01);P27RF-Rho-siRNA+5-Fu组穿过Transwell小室微孔滤膜的平均细胞数明显少于其他3组(P0.01);Western blotting法检测,P27RF-Rho-siRNA+5-Fu组细胞中癌相关蛋白P27表达水平明显高于其他3组(P0.05),侵袭相关蛋白RhoC表达水平低于其他3组(P0.05)。结论:P27RF-Rho基因沉默能明显增强5-Fu对肝癌SMMC7721细胞的药物敏感性。
[Abstract]:Objective: To investigate the hepatocellular carcinoma cell line SMMC7721 P27RF-Rho mRNA gene silencing on 5- fluorouracil (5-Fu) effect of drug sensitivity, and provide a theoretical basis for clinical treatment of advanced hepatocellular carcinoma. Methods: to construct P27RF-Rho RNAi vector, P27RF-Rho gene silencing lentivirus infected SMMC7721 cell, detect the gene silencing effect of.SMMC7721 cells Western blotting were divided into Scramble-siRNA negative the control group, 5-Fu group, P27RF-Rho-siRNA group and P27RF-Rho-siRNA+5-Fu group. Fluorescence microscope was used to detect the transfection effect, detection of RNAi gene silencing efficiency of Western blottting method, MTT method to detect the cells growth curve, cell migration and wound healing assay, invasion assay of Transwell cells, the expression of tumor cells were detected by blotting Western and RhoC P27 related protein results: to construct P27RF-Rho RNAi lentivirus vector.Western blotting method, P27RF-Rh The level was significantly lower than that of 5-Fu group and Scramble-siRNA group, the expression of P27RF-Rho protein in o-siRNA cells (P0.05). Compared with other 3 groups, P27RF-Rho-siRNA+5-Fu group cell growth rate decreased (P0.05); group P27RF-Rho-siRNA+5-Fu cell migration in a scratch assay was significantly decreased (P0.01); the average number of cells in the P27RF-Rho-siRNA+5-Fu group through the Transwell chamber of microporous membrane was significantly less than the other 3 group (P0.01); Western blotting detection method, the expression level of P27RF-Rho-siRNA+5-Fu cells in cancer associated protein P27 was significantly higher than that of the other 3 groups (P0.05), invasion related protein expression level of RhoC was lower than that of the other 3 groups (P0.05). Conclusion: P27RF-Rho gene silencing can significantly enhance the drug sensitivity of 5-Fu on SMMC7721 cells.

【作者单位】：吉林大学第一医院肝胆胰外科;
【基金】：吉林省卫生厅自然科学基金资助课题(3D515U383428)
【分类号】：R735.7

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