敲低E2F1抑制人舌鳞癌细胞增殖及迁移的机制研究  

The mechanism of knockdown of E2F1 inhibiting the proliferation and migration of human tongue squamous cell carcinoma cells

作　　者：杜明珠 张斌 Du Mingzhu;Zhang Bin(Dept of Oral and Maxillofacial Surgery,The First Affiliated Hospital of Jinzhou Medical University,Jinzhou 121000)

机构地区：[1]锦州医科大学附属第一医院口腔颌面外科,锦州121000

出　　处：《安徽医科大学学报》2024年第12期2127-2134,共8页Acta Universitatis Medicinalis Anhui

基　　金：辽宁省教育厅科学技术研究项目(编号:JYTJCZR2020062)。

摘　　要：目的在人舌鳞癌细胞中敲低E2F转录因子1(E2F1),探究其对舌鳞癌细胞增殖、迁移和侵袭的影响并探索其可能的作用机制。方法RT-PCR检测人舌鳞癌组织及癌旁组织样本中E2F1表达。RT-PCR检测人口腔上皮细胞HOEC和人舌鳞癌细胞SCC-9、CAL-27、TSCCa和SCC-25细胞中E2F1基因的本底表达,筛选E2F1显著高表达的细胞株进行后续实验。在舌鳞癌候选细胞中敲低E2F1基因后利用CCK-8、EdU、克隆形成、细胞划痕、Transwell、细胞凋亡和细胞周期实验检测敲低E2F1对人舌鳞癌细胞增殖、迁移、侵袭及凋亡的影响。Western blot检测上皮间充质转化(EMT)标志物钙粘蛋白E(E-cadherin)、钙粘蛋白N(N-cadherin)和蜗牛家族转录抑制因子1(Snail)蛋白及WNT信号通路标志物WNT家族成员3A(WNT3A)、β-连环蛋白(β-catenin)和细胞周期蛋白D1(CCND1)表达变化。结果RT-PCR结果显示,相比癌旁组织,E2F1在癌组织中表达明显升高(P<0.001);同时相比HOEC细胞,E2F1在SCC-9、CAL-27、TSCCa和SCC-25细胞中表达均明显升高(P<0.05),而在SCC-25细胞中高表达最为明显,因此,后续实验使用SCC-25作为实验细胞株。SCC-25细胞中敲低E2F1后,CCK-8细胞存活能力明显被抑制(P<0.001);EdU阳性细胞数、细胞克隆成球数明显减少(P<0.001);细胞总凋亡比例明显升高(P<0.001);细胞处于G_(1)期细胞比例增多(P<0.01),相反处于G_(2)期细胞比例减少(P<0.01);细胞迁移和侵袭能力均明显被抑制(P<0.001);WNT信号通路蛋白WNT3A、β-catenin和CCND1蛋白表达降低(P<0.001),同时EMT相关蛋白N-cadherin和Snail蛋白表达降低(P<0.001),而E-cadherin蛋白表达升高(P<0.001)。结论敲低E2F1抑制人舌鳞癌SCC-25细胞的增殖、迁移、侵袭和EMT过程,同时促进细胞凋亡,这种抑癌作用可能是通过阻断WNT信号通路的激活而实现。Objective To investigate the effects of knockdown of E2F transcription factor 1(E2F1)in human tongue squamous cell carcinoma cells on proliferation,migration,and invasion of tongue squamous cell carcinoma cells,and to explore its possible mechanisms.Methods The expression of E2F1 in human tongue squamous cell carcinoma was detected by RT-PCR,and the background expression of E2F1 gene in HOEC,SCC-9,CAL-27,TSCCa and SCC-25 cells was detected by RT-PCR,and the cell lines with significantly high expression of E2F1 were screened for subsequent experiments.CCK-8,EdU,colony formation,cell scratch,Transwell,apoptosis and cell cycle experiments were used to detect the effects of knockdown of E2F1 on proliferation,migration,invasion and apoptosis of human tongue squamous cell carcinoma cells after knockdown of E2F1 gene in tongue squamous cell carcinoma candidate cells.Western blot was used to detect the expression of epithelial mesenchymal transition markers E-cadherin,N-cadherin and snail family transcription inhibitor 1(Snail)protein and WNT signaling pathway markers WNT family members 3A(WNT3A),β-catenin and cyclin D1(CCND1).Results RT-PCR results showed that the expression of E2F1 in cancer tissues was significantly higher than that in adjacent tissues(P<0.001).At the same time,compared with HOEC cells,the expression of E2F1 in SCC-9,CAL-27,TSCCa and SCC-25 cells significantly increased(P<0.05),and the high expression in SCC-25 cells was the most obvious.SCC-25 would be used as an experimental cell line in subsequent experiments.After knocking down E2F1 in SCC-25 cells,the viability of CCK-8 cells was significantly inhibited(P<0.001).The number of EdU positive cells decreased significantly(P<0.001).The number of cell clones was significantly reduced(P<0.001).The proportion of total apoptosis significantly increased(P<0.001).The proportion of cells in G 1 phase increased(P<0.01).The proportion of cells in G 2 phase decreased(P<0.01).The cell migration and invasion ability were significantly inhibited(P<0.001).Western

关 键 词：E2F1 舌鳞癌 增殖 迁移 侵袭 凋亡 EMT WNT 

分 类 号：R782[医药卫生—口腔医学]