摘要
目的:研究RNA结合基序单链相互作用蛋白3(RNA-binding motif,single-stranded-interacting protein 3,RBMS3)对胃癌细胞侵袭的影响,并探讨其可能的分子作用机制。方法:首先采用实时荧光定量PCR和蛋白质印迹法检测人正常胃黏膜上皮细胞GES-1以及胃癌细胞MKN-28、MKN-45、NCI-N87和SGC-7901中RBMS3的表达水平。然后采用脂质体转染法将RBMS3过表达载体[RBMS3-pcDNA3.1(+)]转染至胃癌MKN-45和SGC-7901细胞中,同时设置空白对照组和空载体转染的阴性对照组。采用实时荧光定量PCR和蛋白质印迹法验证RBMS3过表达效果后,Transwell小室法检测细胞侵袭能力的变化,实时荧光定量PCR和蛋白质印迹法检测细胞中上皮-间质转化相关蛋白以及Wnt信号通路中β-连环蛋白(β-catenin)的表达水平变化。最后,用Wnt通路激动剂氯化锂(lithium chloride,LiCl)处理RBMS3过表达的胃癌MKN-45和SGC-7901细胞后,再次采用Transwell小室法检测细胞侵袭能力的变化。结果:与正常胃黏膜细胞相比,4种胃癌细胞中RBMS3 mRNA和蛋白的表达水平均明显降低(P值均<0.05)。RBMS3过表达质粒转染后,胃癌MKN-45和SGC-7901细胞中RBMS 3基因过表达,2种细胞的侵袭能力随之明显降低(P值均<0.05)。在RBMS3过表达的胃癌MKN-45和SGC-7901细胞中,N-钙黏蛋白(N-cadherin)和β-catenin表达水平均明显降低(P值均<0.05),而E-钙黏蛋白(E-cadherin)表达水平明显升高(P值均<0.05)。用LiCl处理RBMS3过表达的MKN-45和SGC-7901细胞后,RBMS3对2种细胞侵袭的抑制作用均被部分抵消(P值均<0.05)。结论:RBMS3可能通过阻滞Wnt/β-catenin信号通路,抑制人胃癌细胞的侵袭和上皮-间质转化。
Objective： To investigate the effect of RNA-binding motif, single-stranded-interacting protein 3 （RBMS3） on the invasion of gastric cancer cells, and to explore its possible action mechanism. Methods： The expression levels of RBMS3 mRNA and protein in human normal gastric mucosa epithelial cells （GES-1） and gastric cancer cells （MKN-28, MKN-45, NCI-N87 and SGC-7901） were detected by real-time fluorescent quantitative PCR （RFQ-PCR） and Western blotting, respectively. The RBMS3 overexpression vectors [RBMS3-pcDNA3.1（＋）] were transfected into gastric cancer MKN-45 and SGC-7901 cells by using LipofectAMINE 2000, while the corresponding empty vector-transfection group and untransfection group were arranged as the negative control and blank control, respectively. The efficiency of RBMS3 overexpression was verified by RFQ-PCR and Western blotting. The cell invasion ability was detected by Transwell assay. The expression levels of epithelial-mesenchymal transition-related proteins and Wnt signal pathway protein J3-catenin were detected by RFQ-PCR and Western blotting. After RBMS3 overexpressed MKN-45 and SGC-7901 cells were treated with Wnt pathway agonist lithium chloride （LiCI）, the change of cell invasion ability was retested by Transwell assay. Results： The mRNA and protein levels of RBMS3 in 4 gastric cancer cell lines were obviously lower than those in normal gastric mucosa epithelial cells （all P 〈 0.05）. After transfection of RBMS3 overexpression vectors, RBMS3 gene was overexpressed in gastric cancer MKN-45 and SGC-7901 cells （both P 〈 0.05）, the invasion abilities of these two cells were significantly declined （both P 〈 0.05）. In RBMS3 gene-overexpressed MKN-45 and SGC-7901 cells, the expressions of N-cadherin and 13-catenin were significantly decreased （all P 〈 0.05）, while the expression of E-cadherin was significantly increased （both P 〈 0.05）. After LiCI treatment, the inhibitory effect of RBMS3 on invasion of gastric cancer MKN-45 and SGC-7901 c
出处
《肿瘤》
CAS
CSCD
北大核心
2017年第10期1032-1040,共9页
Tumor
关键词
胃肿瘤
基因表达调控
肿瘤侵润
上皮-间质转化
WNT信号通路
RNA结合基序单链相互作用蛋白3
Stomach neoplasms
Gene expression regulation
Neoplasm invasiveness
Epithelial-mesenchymal transition
Wnt signaling pathway
RNA-binding motif, single- stranded-interacting protein 3
