沉默RohA基因对舌鳞状细胞癌细胞增殖的影响  

Effects of RhoA silencing on proliferation of tongue squamous cancer cells

作　　者：严国鑫[1] 樊兵[1] 邹荣海[1] 张健[1] 孙晓峰[1] 童磊[2] 王奇民[2] 韩金宏[2] 鲁旭飞 王莹[4] 周元[4] 何宗轩 廖奕翔[2] 李宁[5] 曹蕾[5] 陈正岗[2,6] 

机构地区：[1]无锡市第二人民医院口腔科,无锡214002 [2]青岛大学医学院附属青岛市市立医院口腔医学中心,青岛266071 [3]即墨市普东卫生院口腔科,青岛266234 [4]潍坊医学院口腔医学院,潍坊261021 [5]大连医科大学研究生院,大连116044 [6]上海交通大学医学院附属第九人民医院口腔颌面外科,上海200011

出　　处：《华西口腔医学杂志》2016年第6期620-625,共6页West China Journal of Stomatology

基　　金：国家自然科学基金(81372908);南京医科大学科技发展基金重点项目(2012NJMU248);青岛市卫计委计划项目(2014-WJZD009;2013-WSZD011)~~

摘　　要：目的通过RNA干扰技术沉默RhoA基因从而探讨RhoA对舌癌细胞增殖和生长的影响及其作用机制。方法体外培养舌鳞状细胞癌SCC-4细胞,以小分子干扰RNA转染沉默RhoA基因的表达。实验分为3组:实验组(又分为实验1组和实验2组,脂质体分别转染对应序列1的RhoA-siRNA和序列2的RhoA-siRNA)、阴性对照组(脂质体转染NC-siRNA)和空白对照组(不转染siRNA)。采用实时定量聚合酶链反应技术检测SCC-4细胞转染后RhoA m RNA的表达,Western blot检测RhoA、Cyclin D1、p21和p27蛋白的表达,四唑盐比色法检测舌癌细胞生长水平和倍增时间。结果与阴性对照组和空白对照组相比,实验组舌癌细胞的RhoA基因及蛋白表达降低,p21、p27蛋白表达升高,Cyclin D1蛋白表达降低,细胞倍增时间延长,增殖能力降低(P<0.05)。结论沉默RhoA基因可以抑制舌癌细胞的增殖和生长,RhoA基因通过调控细胞周期信号转导途径影响舌癌细胞的增殖,RhoA基因可以成为舌癌基因治疗的靶点。Objective This study investigated the effect of RhoA silencing through RNA interference on proliferation and growth of tongue cancer cells, as well as explored the possible mechanisms of this effect. Methods SSC-4 tongue cancer cells were cultured in vitro and then transfected with small interfering RNA to knock down RhoA expression. The tested cells were divided into three groups： experimental group （experimental group 1： transfected with RhoA-siRNA- 1 ; experimental group 2： transfected with RhoA-siRNA-2）, negative control group （transfected by random sequence NC-siRNA）, and blank control group （transfected with Lipofectamine）.The expression levels of RhoA mRNA were respectively measured by quantitative real-time polymerase chain reaction and western blot assay. Moreover, the expression levels of cyclin D 1, p21, and p27 and RhoA protein were evaluated by Westem blot assay. Proliferation and growth potentiality were analyzed through evaluation of doubling times and methyl thiazolyl tetra-zolium assessment. Results The expression levels of RhoA gene and protein of experimental groups significantly decreased following siRNA transfection compared with those in the negative and blank control groups. The expression of cyclin D 1 decreased significantly and that of p21 and p27 increased significantly. The doubling time was extended and the growth potentiality decreased. Conclusion The results indicated that RhoA silencing can inhibit proliferation of tongue cancer cells, whereas RhoA affects cell proliferation by regulating the cell cycle pathway. Thus, RhoA is a potential target in gene therapy for tongue cancer.

关 键 词：RHOA 舌癌 细胞增殖 RNA干扰 

分 类 号：R739.86[医药卫生—肿瘤]