慢病毒介导靶向P27RF-Rho基因沉默对肝癌细胞侵袭性的影响

doi:10.13481/j.1671-587x.20160214

吉林大学学报(医学版) ›› 2016, Vol. 42 ›› Issue (02): 260-265.doi: 10.13481/j.1671-587x.20160214

慢病毒介导靶向P27RF-Rho基因沉默对肝癌细胞侵袭性的影响

马强, 谢淑丽, 王广义, 邢光远, 杨耀群, 吕国悦

吉林大学第一医院肝胆胰外科, 吉林长春 130021

收稿日期:2015-05-14 发布日期:2016-03-31
通讯作者: 吕国悦,教授,硕士研究生导师(Tel:0431-88783331,E-mail:lgy08@sina.com) E-mail:lgy08@sina.com
作者简介:马强(1990-),男,吉林省长春市人,医师,在读医学硕士,主要从事肝胆胰腺疾病外科治疗研究。
基金资助:
吉林省科技厅科研基金资助课题(20130413021GH)

Influence of lentiviral-mediated P27RF-Rho gene silence in invasion of liver cancer cells

MA Qiang, XIE Shuli, WANG Guangyi, XING Guangyuan, YANG Yaoqun, LYU Guoyue

Department of Hepatobiliary and Pancreatic Surgery, First Hospital, Jilin University, Changchun 130021, China

Received:2015-05-14 Published:2016-03-31

摘要/Abstract

摘要：

目的: 探讨慢病毒靶向介导技术沉默P27RF-Rho基因,阐述其对肝癌细胞侵袭性的影响。方法: 构建P27RF-Rho RNAi慢病毒。慢病毒感染肝癌细胞BEL7402。实验分为P27RF-Rho-siRNA实验组、Scramble-siRNA阴性对照组和BEL7402空白对照组。Western bloting法检测P27RF-Rho基因沉默效果及肝癌相关蛋白RhoA、RhoC、VEGF、P53和PTEN表达水平;明胶酶谱分析肿瘤侵袭相关基质金属蛋白酶(MMPs)活性;细胞划痕和体外Transwell小室侵袭实验比对细胞迁移和侵袭能力的改变。结果: Western blotting法检测,P27RF-Rho-siRNA实验组BEL7402中P27RF-Rho、RhoA、RhoC和VEGF蛋白表达水平明显低于2个对照组(P<0.05),P53和PTEN表达水平高于2个对照组(P<0.05)。明胶酶谱,P27RF-Rho-siRNA实验组MMPs活性明显降低(P<0.01)。划痕实验,P27RF-Rho-siRNA实验组细胞迁移距离明显低于2个对照组(P<0.01)。实验组穿过Transwell小室的平均细胞数明显少于2个对照组(P<0.01)。结论: 沉默P27RF-Rho基因可以降低肝癌细胞BEL7402的侵袭和迁移能力。

关键词: P27RF-Rho, 慢病毒, 肝癌细胞, 侵袭和迁移

Abstract:

Objective: To investigate the silencing of P27RF-Rho gene with lenvirus targeting mediated technique, and to clarify its influence in the invasion of liver cancer cells.Methods: The P27RF-Rho RNAi lentivirus was constructed.The liver cancer BEL7402 cells were infected with lentivirus.The experiment was divided into P27RF Rho-siRNA group, scramble-siRNA group and BEL7402 group. The effect of silencing P27RF-Rho gene and the expression levels of hepatocellular carcinoma (HCC) associated proteins RhoA, RhoC, VEGF, P53 and PTEN were detected;the activities of matrix metalloproteinase (MMPs)associated with tumor invasion were analyzed by Gelatin zymography; the variation of transfer ability and invasion abilities were compared by Wound healing assay experiment and Transwell experiment.Results: The Western blotting results showed the expression levels of P27RF-Rho, RhoA, RhoC, and VEGF proteins in the BEL7402 cells in experiment group were significantly lower than those in two control groups(P<0.05), and the expression levels of P53 and PTEN were higher than those in two control groups(P<0.05).The results of Gelatin zymography showed the activities of MMPs in experiment group were significantly lower than those in two control groups(P<0.01);Wound healing assay showed that the migration ability of the BEL7402 cells in experiment group was significantly inhibited (P<0.01);the number of cells passed through the Transwell Chambers in experiment group was significantly less than those in two control groups (P<0.01).Conclusion: Silenceing P27RF-Rho can weaken the invasion ability and migration ability of human HCC BEL7402 cells.

Key words: P27RF-Rho, lentivirus, liver cancer cells, invasion and migration

中图分类号:

R735.7

马强, 谢淑丽, 王广义, 邢光远, 杨耀群, 吕国悦. 慢病毒介导靶向P27RF-Rho基因沉默对肝癌细胞侵袭性的影响[J]. 吉林大学学报(医学版), 2016, 42(02): 260-265.

MA Qiang, XIE Shuli, WANG Guangyi, XING Guangyuan, YANG Yaoqun, LYU Guoyue. Influence of lentiviral-mediated P27RF-Rho gene silence in invasion of liver cancer cells[J]. Journal of Jilin University Medicine Edition, 2016, 42(02): 260-265.

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慢病毒介导靶向P27RF-Rho基因沉默对肝癌细胞侵袭性的影响

Influence of lentiviral-mediated P27RF-Rho gene silence in invasion of liver cancer cells

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