摘要
目的观察miR-128、ZEB1对子宫内膜癌(EC)细胞迁移侵袭和上皮间质转化的影响,并验证其二者之间的靶向关系。方法采用qRT-PCR和Western blotting法分别检测3种EC细胞系(HEC-1A、Ishikawa、KLE)及1种子宫内膜上皮细胞系hEEC中miR-128和ZEB1。以miR-128表达水平最低的Ishikawa细胞系作为后续研究对象,将Ishikawa细胞分为miR-128过表达组和阴性对照组,分别转染miR-128 mimics和miRNA-NC,48 h后收集细胞进行后续实验。细胞划痕实验和Transwell实验检测转染后各组细胞迁移和侵袭能力,Western blotting法检测转染后各组细胞ZEB1、E-cadherin、Fibronectin、Vimentin等蛋白。双荧光素酶报告基因实验验证miR-128与ZEB1的靶向调控关系。结果与hEEC相比,EC细胞系HEC-1A、Ishikawa、KLE中miR-128表达降低(P均<0.05),ZEB1蛋白表达升高(P均<0.05)。转染后,Ishikawa细胞中miR-128的表达明显升高(P<0.05),穿膜细胞数目及划痕愈合率均降低(P均<0.05),ZEB1、Fibronectin、Vimentin蛋白的相对表达量均降低(P均<0.05),E-cadherin蛋白表达升高(P<0.05)。双荧光素酶报告基因实验结果显示,miR-128能够靶向调控ZEB1基因(P<0.05)。结论miR-128可以抑制Ishikawa细胞的迁移、侵袭及上皮间质转化,ZEB1可促进Ishikawa细胞的迁移、侵袭及上皮间质转化,miR-128与ZEB1具有靶向调控关系。
Objective To observe the effects of miR-128 and ZEB1 on the migration,invasion and epithelial mesenchymal transition(EMT)of endometrial carcinoma(EC)cells and to verify the targeted relationship between them.Methods The qRT-PCR and Western blotting were used to detect the relative expression levels of miR-128 and ZEB1 in three EC cell lines(HEC-1 A,Ishikawa,and KLE)and one endometrial epithelial cell line hEEC,respectively.Ishikawa cell line with the lowest expression level of miR-128 was taken as the follow-up study object.Ishikawa cells were divided into the negative control group and miR-128 overexpression group,and were transfected with miRNA-NC and miR-128 mimics,respectively.After 48 h,the cells were collected for follow-up experiments.Cell Scratch test and Transwell test were used to detect the migration and invasion abilities of cells in each group after transfection.Western blotting was used to detect ZEB1,E-cadherin,Fibronectin and Vimentin protein of cells in each group after transfection.Double luciferase reporter gene experiment was used to verify the targeted regulatory relationship between miR-128 and ZEB1.Results Compared with hEEC,the expression of miR-128 in EC cell lines HEC-1 A,Ishikawa and KLE decreased(all P<0.05),and the expression of ZEB1 protein increased(P<0.05).After transfection,the expression of miR-128 increased significantly in Ishikawa cells(P<0.05),the number of transmembrane cells and scratch healing rate decreased(all P<0.05),the relative expression levels of ZEB1,Fibronectin and Vimentin decreased(all P<0.05),and the expression of E-cadherin protein increased(P<0.05).The results of double luciferase reporter gene experiment showed that miR-128 could target and regulate ZEB1 gene(P<0.05).Conclusions MiR-128 can inhibit the migration,invasion and EMT of Ishikawa cells.ZEB1 can promote the migration,invasion and EMT of Ishikawa cells.MiR-128 and ZEB1 have a targeted regulatory relationship.
作者
李品
蔡智慧
李晶晶
王琰
陈静
杜晓欣
王兰
闫丽伟
LI Pin;CAI Zhihui;LI Jingjing;WANG Yan;CHEN Jing;DU Xiaoxin;WANG Lan;YAN Liwei(Department of Gynecology,Affiliated Hospital of Hebei University,Baoding 071000,China)
出处
《山东医药》
CAS
2022年第27期1-5,共5页
Shandong Medical Journal
基金
河北省保定市科技计划项目(2041ZF309)。
作者简介
第一作者:李品(1986-),女,主治医师,主要研究方向为妇科相关恶性肿瘤。E-mail:dr_lpfck@yeah.net;通信作者:闫丽伟(1988-),女,主治医师,主要研究方向为妇科疾病的诊疗。E-mail:vfvhr9@163.com。
