摘要
目的通过RNA干扰技术沉默Rce1基因,探讨Rce1对舌癌细胞侵袭和迁移能力的影响。方法体外培养舌鳞状细胞癌Cal-27和SCC-4细胞,设计合成Rce1基因的小干扰RNA(si RNA),采用脂质体载体瞬时转染沉默Rce1基因。根据转染的si RNA不同,将实验组分为Rce1-si RNA-1组、Rce1-si RNA-2组、Rce1-si RNA-3组,脂质体载体分别转染相对应序列的si RNA。脂质体转染si CON作为阴性对照组,不转染si RNA作为空白对照组。采用实时定量聚合酶链反应检测各组Rce1、Rho A以及K-Ras基因的表达,蛋白质印迹法(Western blot)检测Rce1、Rho A、K-Ras、金属基质蛋白酶(MMP)-2及MMP-9的表达,采用体外侵袭实验检测Cal-27和SCC-4的侵袭能力,细胞划痕实验检测细胞迁移能力。结果实时定量聚合酶链反应及Western blot检测结果显示,与阴性对照组和空白对照组相比,实验组的Rce1基因及蛋白表达均下调(P<0.05),Rho A、K-Ras基因及蛋白表达无统计学差异(P>0.05),MMP-2、MMP-9表达下降(P<0.05)。体外侵袭实验表明,与对照组相比,实验组在聚碳酯膜上的细胞总数明显减少(P<0.05)。细胞划痕实验表明,实验组划痕处细胞闭合时间明显长于对照组(P<0.05)。结论体外沉默Rce1可以有效下调其在舌癌细胞Cal-27和SCC-4中的表达水平,降低迁移和侵袭能力。
Objective This study aimed to explore the influence of Rce1 on invasion and migration of tongue squamous cell carcinoma cells by silencing the Rce1 gene with RNA interference. Methods The tongue squamous cell carcinoma Cal-27 and SCC-4 cells were cultured in vitro. The small interfering RNA (siRNA) of the Rce1 gene was designed, and the Rcel gene expression was silenced vialiposome transfection. According to the siRNA transfected by liposome, the experimental group was divided into three groups, namely, Rce1-siRNA-1, Rce1-siRNA-2, and Rce1-siRNA-3 groups. Negative control group was transfected by siCON, and the blank control group was untransfected by siRNA. The Rce1, RhoA, and K-Ras gene expression levels in each group were analyzed by real-time quantitative polymerase chain reaction. The Rce1, RhoA, K-Ras, MMP-2, and MMP-9 protein expression levels were analyzed by Western blot. The invasiveness of tongue cancer cell Cal-27 and SCC-4 were determined by Transwell invasion assay, and cell migration assay was performed by cell scratch assay. Results Real-time quantitative polymerase chain reaction and Western blot results showed that compared with the negative and blank control groups, the Rce1 gene and protein expression levels in three experimental groups decreased (P<0.05). The RhoA, K-Ras gene and protein expression levels were insignificantly different among groups (P>0.05). Meanwhile, the MMP-2 and MMP-9 expression levels decreased (P<0.05). Transwell invasion assay results showed that the total number of cells in the PET film of the experimental groups was significantly decreased compared with the control group (P<0.05). The cell scratch test showed that the cell closure time of the scratch in the interference group was significantly longer than those in the control and blank groups (P<0.05). Conclusion Silencing Rce1 in vitro can effectively downregulate its expression in tongue squamous cell carcinoma cells Cal-27 and SCC-4 and reduce the migration and invasion abilities of these cells.
作者
孙君君
陶匀雅
周元
何宗轩
盛善桂
王奇民
童磊
赵开
王少如
陈正岗
Junjun Sun;Yunya Tao;Yuan Zhou;Zongxuan He;Shangui Sheng;Qimin Wang;Lei Tong;Kai Zhao;Shaoru Wang;Zhenggang Chen(Dept. of Stomatology, Qingdao Chengyang District Hospital, Qingdao 266109, China;Medical Center of Stomatology, Affiliated Qingdao Municipal Hospital, Qingdao University, Qingdao 266071, China;Dept. of Oral and Maxillafacial Surgery, The Affiliated Hospital of Qingdao University, Qingdao 266005, China;Stomatology College, Dalian Medical University, Dalian 116044, China)
出处
《华西口腔医学杂志》
CAS
CSCD
北大核心
2019年第2期143-148,共6页
West China Journal of Stomatology
基金
国家自然科学基金(81372908)~~
关键词
Rce1
鳞状细胞癌
侵袭
迁移
Rce1
squamous cell carcinoma
invasion
migration
