Effects of siRNA silencing Rictor gene expression on proliferation, migration and invasion of hepatocellular carcinoma cells

Abstract

Objective To investigate the effect of siRNA targeted silencing Rictor expression on the proliferation, migration and proliferation in hepatocellular carcinoma cells.

Methods LipofectamineTM2000 was used to transfect SMMC-7721 and Hep3B cells to construct siRNA vector fragments (silenced group) or sense-free siRNA fragments (control group) targeting Rictor expression. After transfection for 48 h, Rictor mRNA level was detected by QPCR and Rictor, p-Akt, cyclin and EMT-related proteins were detected by Western blotting. MTT assay, EdUincorporation assay, Transwell assay and Boyden assay were used to detect the proliferation, migration and proliferation of HCC cells, respectively.

Results After transfection for 48 h, most of the Rictor mRNA level of the silenced group was lower than that of the control group. The ability of cell proliferation, migration and invasion in the silenced group was weakened when compared to that in the control group ( P<0.01). In MTT assay and EdU incorporation assay, knockdown of Rictor expression inhibited proliferation in two hepatocellular carcinoma cell lines, while in Transwell assay and Boyden assay, knockdown of Rictor expression inhibited migration and invasion in both cell lines ( P<0.01).

Conclusion Rictor gene plays an oncogene role in hepatocellular carcinoma cells. Silencing Rictor expression inhibits the proliferation, migration and invasion of hepatocellular carcinoma cells, which may be related to down-regulating the expression of p-Akt, CCND1 and N-cadherin, ZEB1 and Vimentin, and up-regulating the expression of E-cadherin, P21 and P27.

Abstract

摘要： 目的 探讨靶向沉默 Rictor 表达对肝癌细胞增殖、迁移和增殖的影响。 方法 采用 Lipofedamine TM2000 向 SMMC-7721 和 Hep3B 细胞转染成功构建靶向沉默 Rictor 表达的 siRNA 载体片段(沉默组)或无义 siRNA 片段(对照组) , 转染 48 h 后采用 QPCR 检测 RktormRNA 水平, 以 Western blotting 检测 Rktor、p-Akt、细胞周期蛋白和EMT相关蛋白的 表达情况。MTT 实验、EdU 增殖实验、Transwell 实验和 Boyden 实验分别检测肝癌细胞的增殖、迁移和增殖能力。 结果 转染 48 h 后, 沉默组细胞的 RictormRNA 蛋白水平大部分都低于对照组细胞；沉默组细胞的增殖、迁移、侵袭能 力均低于对照组细胞, 差异有统计学意义 ( P<0.01)。MTT 实验及 EdU 增殖实验表明, 两组肝癌细胞在沉默 Rictor 表达 后增殖减慢、S期细胞比例明显减少 ( P 均<0.01); Transwell 实验和 Boyden 实验表明, 两组肝癌细胞在沉默 Rictor 表达后 穿膜细胞数显著减少 ( P 均<0.01)。 结论 Rktor 基因在肝癌细胞中起到癌基因的作用, 沉默 Rictor 的表达抑制肝癌细 胞的增殖、迁移和侵袭的过程, 可能与下调 p-Akt、CCND1 和 N-cadherin、ZEB1、Vimentin 蛋白表达, 上调 E-cadherin, P21 和 P27 蛋白表达有关。

Journal

Journal ID (publisher-id): CTM

Title: China Tropical Medicine

Publisher: China Tropical Medicine (China )

ISSN (Electronic): 1009-9727

Publication date (Print): 1 March 2020

Publication date (Electronic): 1 April 2020

Volume: 20

Issue: 3

Pages: 212-218

Affiliations

¹Department of Oncology, Haikou Municipal People’s Hospital and Central South University Xiangya Medical College Affiliated Hospital, Haikou, Hainan 570208, China

²Cancer Center, Integrated Hospital of Traditional Chinese Medicine, Southern Medical University, Guangzhou, Guangdong 410000, China

Author notes

Corresponding author: LUO Rongcheng, E-mail: luorc02@ 123456vip.163.com

Article

Publisher ID: j.cnki.46-1064/r.2020.03.04

DOI: 10.13604/j.cnki.46-1064/r.2020.03.04

Copyright statement: © 2020 Editorial Department of China Tropical Medicine

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Subject: Journal Article