宫颈腺癌细胞中HPV18-E7与上皮间质转化的关系研究

Study on the relationship between HPV18-E7 and epithelial-mesenchymal transition in cervical adenocarcinoma cells

目的:探究宫颈腺癌细胞中HPV18-E7与上皮间质化(EMT)之间的关系。方法:针对HPV18-E7的si RNA转染入人宫颈腺癌He La细胞,观察细胞形态变化,并运用Real-time PCR技术、免疫印迹实验和细胞免疫荧光染色实验在m RNA水平和蛋白水平检测E7、EMT相关的标记因子的表达及定位变化;应用细胞划痕愈合实验及Transwell细胞体外侵袭实验检测细胞迁移和侵袭能力。结果:特异性沉默HPV18-E7表达后He La-SIE7细胞间连接变得疏松,细胞中E7与间质性标记物的m RNA及蛋白表达水平较对照组细胞显著下降,上皮性标记因子较对照组细胞明显增加(P<0.01);随着He La细胞E7表达的下调,Ecadherin在细胞膜上的表达增加,间质性标记因子在细胞质内的表达随之下降。He La-SIE7细胞迁移及侵袭能力明显低于对照组(P<0.01);沉默HPV18-E7表达的He La细胞其增殖能力也明显减弱。结论:宫颈腺癌细胞中HPV18-E7能够引起EMT,促进肿瘤发生侵袭转移。

Objective： To determine the role of HPV18-E7 oncogene in the EMT-like process. Methods： HPV18-E7 siRNAs were transfected in human cervical cancer HeLa cells. Real-time PCR, western blot and immunofluorescence technique were performed to examine the localization and expression of E7 andE MT markers, respectively. Furthermore, the Wound healing assay and Matrigel invasion assay were used to evaluate the invasion and migration ability. Results： On the mRNA and protein levels, E7 depletion significantly induced the expression of E-caderin. Furthermore, N-caderin, Vimetin, and fibronectin expression were decreased in HeLa-siE7 cells. E-cadherin was up-regulated and mainly observed in contact areas between cells in HeLa/siE7 cells, while the mesenchymal markers such as N-caderin, and fibronectin were reduced and detected mainly in the cytoplasm. Down-regulation of HPV18-E7 by transient transfection of HPV18-E7 siRNAs in HeLa cells caused significant inhibition of cell invasion and migration compared with the siNC transfected cells. Conclusion： In HeLa cells HPV18-E7 can enhance invasion and migration by inducing EMT.