沉默RhoA基因对腺样囊性癌细胞迁移和侵袭的影响  

Effects of RhoA silencing on the migration and invasion of salivary adenoid cystic carcinoma

作　　者：迟增鹏 周建华[2] 李文健 王莹 徐晓妹[5] 陈正岗[2] CHI Zeng-peng;ZHOU Jianhua;LI Wenjian;WANG Ying;XU Xiaomei;CHEN Zhenggang(College of Stomatology,Wei-fang Medical University,Weifang 261021,China;Department of Stomatology,Qingdao Municipal Hospital,Qingd-ao University,Qingdao 266071,China;School of Stomatology,Dalian Medical University,Dalian 116044,China;Department of Stomatology,Fourth People􀆳s Hospital of Jinan,Jinan 250031,China;Department of Oncology,Qingdao Municipal Hospital,Qingdao University,Qingdao 266071,China)

机构地区：[1]潍坊医学院口腔医学院,山东潍坊261021 [2]青岛大学附属青岛市市立医院口腔医学中心,山东青岛266071 [3]大连医科大学口腔医学院,辽宁大连116044 [4]济南市第四人民医院口腔科,山东济南250031 [5]青岛大学附属青岛市市立医院肿瘤科,山东青岛266071

出　　处：《口腔疾病防治》2022年第4期237-244,共8页Journal of Prevention and Treatment for Stomatological Diseases

基　　金：国家自然科学基金项目(81372908)。

摘　　要：目的探讨RhoA基因对唾液腺腺样囊性癌(salivary adenoidcysticcarcinoma,SACC)细胞的迁移和侵袭能力的影响。方法将液氮冻存的20例SACC及正常癌旁组织研磨匀浆,分别提取总RNA和总蛋白检测RhoA的表达情况。同时构建RhoA⁃siRNA转染2个细胞株SACC⁃LM和SACC⁃83细胞进行细胞学实验,实验分为实验组(转染RhoA⁃siRNA基因),阴性对照组(转染siRNA⁃NC基因)和空白对照组。通过qRT⁃PCR检测各组细胞RhoA的mRNA表达并确定转染效率;Western blot分析各组细胞RhoA及上皮-间充质转换(epithelial⁃mesenchymal transition,EMT)标志分子(E⁃cadherin、N⁃cadherin、Vimentin)的蛋白表达;Transwell实验及划痕实验分析各组细胞侵袭和迁移能力的变化。结果与正常癌旁组织相比,RhoA mRNA和蛋白相对表达量在SACC组织中增高(P<0.01);实验组与对照组相比,RhoA mRNA的相对表达量和蛋白的相对表达量降低,E⁃cadherin蛋白的相对表达量增加,N⁃cadherin、Vimentin蛋白的相对表达量降低(P<0.01);实验组细胞侵袭和迁移能力降低(P<0.01)。结论RhoA在SACC组织中表达增高;体外沉默RhoA基因可有效抑制SACC⁃LM和SACC⁃83细胞迁移和侵袭能力,其可能通过作用EMT影响SACC细胞迁移和侵袭能力。Objective To explore whether RhoA plays a role in the migration and invasion of the salivary adenoid cystic carcinoma cell lines SACC⁃LM and SACC⁃83.Methods Total RNA and total protein were extracted from 20 sal⁃ivary adenoid cystic carcinoma(SACC)and normal adjacent tissues frozen in liquid nitrogen to detect RhoA expression.RhoA⁃siRNA was constructed to transfect two cell lines(SACC⁃LM and SACC⁃83)for cytological experiments.The re⁃search included an experimental group(RhoA⁃siRNA transfection),negative control group(siRNA⁃NC transfection)and blank group by transient transfection with liposomes.Expression of RhoA mRNA and protein as well as the protein ex⁃pression of biomarkers of epithelial⁃mesenchymal transition(EMT)were analyzed,including E⁃cadherin,N⁃cadherin,and Vimentin.Furthermore,the changes in invasion and migration of cells in each group were analyzed by comparing the number of transmembrane cells in the Transwell assay and the results of the scratch test.Results Compared with normal adjacent tissues,RhoA protein and mRNA levels increased in SACC tissues.Compared with the control group,the relative expression levels of RhoA mRNA and protein decreased(P<0.01),the relative expression levels of E⁃cad⁃herin protein increased,and the relative expression levels of N⁃cadherin and vimentin protein increased in the experi⁃mental group(P<0.01).Additionally,the trial results revealed that RhoA knockdown restrained cell migration and in⁃vasion(P<0.01).Conclusion RhoA expression increased in SACC tissue.Silencing RhoA in vitro could effectively restrain cell migration and invasion in SACC⁃LM and SACC⁃83 cells through the regulation of EMT signaling pathways.

关 键 词：小干扰RNA RHOA 腺样囊性癌 上皮⁃间充质转换 迁移 侵袭 E⁃钙黏蛋白 N⁃钙黏蛋白 波形蛋白 

分 类 号：R78[医药卫生—口腔医学]