摘要
目的探讨DDX3表达上调在人宫颈癌细胞增殖中的作用。方法采用免疫组织化学方法检测2012年4月至2013年3月河南省人民医院59例宫颈癌组织及癌旁非肿瘤组织标本中的DDX3表达情况。同时,以宫颈癌细胞HeLa为研究对象,通过慢病毒介导的宫颈癌细胞过度表达来检测DDX3的功能。采用细胞增殖与活性检测试剂盒评估细胞存活率,Boyden室进行迁移和侵袭测定,即时荧光定量聚合酶链反应检测DDX3 mRNA表达水平,Western blot检测细胞中上皮间质转化(EMT)和PI3K/Akt信号通路相关蛋白表达。结果 DDX3过表达与国际妇产科联盟(FIGO)分期、宫颈浸润深度和淋巴结转移呈正相关(P<0.05)。Kaplan-Meier分析显示,DDX3高表达的宫颈癌患者具有较差的总体生存率(P<0.05)。与慢病毒载体对照(pLVX-Con)组相比,在shRNA-DDX3慢病毒(pLVX-DDX3)转染组中检测到DDX3的蛋白表达和mRNA表达均明显增加(均P<0.01)。在pLVX-DDX3转染第72小时的HeLa细胞存活数目显著高于pLVX-Con转染细胞(P<0.05)。与pLVX-Con转染的细胞相比,DDX3过表达显著促进了HeLa细胞的迁移和侵袭(P<0.05),并且显著上调了HeLa细胞的N-Cadherin、波形蛋白和Snail的表达(P<0.05)。在pLVX-DDX3组中,Akt及其下游靶基因p-GSK3β的磷酸化水平和β-catenin表达较pLVX-Con组显著升高(P<0.05),而当向pLVX-DDX3组加入PI3K抑制剂LY294002后,p-Akt、p-GSK3β和β-catenin明显降低(P<0.05),并且N-Cadherin、波形蛋白和Snail的水平表达下调(P<0.05)。结论 DDX3过表达促进宫颈癌细胞的增殖、迁移和侵袭,并能诱导EMT,其作用机制可能与激活PI3K/Akt信号通路有关。
Objective To investigate the role of DDX3 up-regulation in the proliferation of human cervical cancer cells and its correlation with clinical prognosis.Methods Expression levels of DDX3 in the 59 specimens of cervical cancer and adjacent non-neoplastic tissue collected at Henan Provincial People′s Hospital from April 2012 to March 2013 were detected using immunohistochemistry.A lentivirus-mediated DDX3-over-expression cell line was constructed based on HeLa cells of cervical cancer.CCK-8 assay was used to evaluate cell survival rate.Boyden chamber was used to measure the cell migration and invasion.Real-time fluorescence quantitative PCR was used to detect DDX3 expression level and Western blot was used to detect the expression of EMT and PI3K/Akt signal pathway-related proteins.Results DDX3 overexpression was associated with FIGO stage,depth of cervical invasion and lymph node metastasis(P<0.05).Kaplan-Meier analysis revealed that cervical cancer patients with high expression of DDX3 had a poor overall survival(P<0.05).Compared with the cells transfected with pLVX-Con vector,the expression of DDX3 protein and mRNA was significantly increased in the cells transfected with pLVX-DDX3(all P<0.01).Cell proliferation was significantly increased following transfection with pLVX-DDX3 for 72 h in HeLa cells compared with that transfected with pLVX-Con(P<0.05).Compared with the controls,DDX3 overexpression significantly promoted the migration and invasion of HeLa cells(P<0.05),and increased the expression of N-Cadherin,vimentin and Snail in HeLa cells(P<0.05).In pLVX-DDX3 group,the expression levels ofβ-catenin,phosphorylated Akt,and pAkt′s downstream target p-GSK3βwere significantly higher than those of pLVX-Con group(P<0.05).The expression levels of p-Akt,p-GSK3βandβ-catenin were decreased when the PI3K/Akt pathway was blocked using the PI3K inhibitor LY294002(P<0.05),and the expression levels of N-Cadherin,vimentin and Snail were also significantly decreased(P<0.05).Conclusions DDX3 overexpression promotes prol
作者
郭芳芳
赵瑞皎
李杜娟
徐紫光
孔令非
Guo Fangfang;Zhao Ruijiao;Li Dujuan;Xu Ziguang;Kong Lingfei(Department of Pathology,Henan Provincial People′s Hospital,Zhengzhou 450000,China)
出处
《中华病理学杂志》
CAS
CSCD
北大核心
2021年第2期119-124,共6页
Chinese Journal of Pathology
基金
河南省医学科技攻关计划联合共建项目(2018020401)。
