miR-128-3p通过调控Notch1表达影响子宫内膜癌细胞的增殖和凋亡  

miR-128-3p affects proliferation and apoptosis of endometrial cancer cells by regulating Notch1 expression

作　　者：顾晓荔[1] 任晓爽 廖予妹[1] GU Xiao-Li;REN Xiao-Shuang;LIAO Yu-Mei(Department of Obstetrics and Gynecology,the Second Affiliated Hos-pital of Zhengzhou University,Zhengzhou 450014,China)

机构地区：[1]郑州大学第二附属医院妇产科,郑州450014 [2]济南市妇幼保健院妇科,济南250001

出　　处：《中国免疫学杂志》2021年第16期1972-1978,共7页Chinese Journal of Immunology

基　　金：河南省基础与前沿技术研究计划项目(No.122300410409)。

摘　　要：目的:探讨微小RNA-128-3p(miR-128-3p)对子宫内膜癌(EC)细胞增殖和凋亡的影响及作用机制。方法:培养人子宫内膜血管内皮细胞hEEC和EC细胞系HEC-1A、Ishikawa和AN3CA,RT-qPCR检测细胞中miR-128-3p和Notch1mRNA表达水平,Western blot法检测细胞中Notch1蛋白水平。转染miR-128-3p模拟物或Notch1小干扰RNA至Ishikawa细胞,构建过表达miR-128-3p或低表达Notch1的Ishikawa细胞,四甲基噻唑蓝染色法(MTT)、流式细胞术、Western blot分别检测过表达miR-128-3p或低表达Notch1对Ishikawa细胞增殖、凋亡及细胞周期蛋白D1(cyclinD1)、活化的半胱天冬酶-3(cleaved-caspase-3)和β-连接素(β-catenin)蛋白表达的影响。双荧光素酶报告基因实验验证Ishikawa细胞中miR-128-3p与Notch1调控关系。结果:与hEEC细胞比较,EC细胞系HEC-1A、Ishikawa和AN3CA中miR-128-3p表达水平降低(P<0.05),Notch1 mRNA和蛋白表达水平升高(P<0.05)。过表达miR-128-3p或低表达Notch1后,Ishikawa细胞存活率和cyclinD1蛋白水平降低(P<0.05),细胞凋亡率和cleaved-caspase-3蛋白水平升高(P<0.05)。miR-128-3p在Ishikawa细胞中靶向负调控Notch1表达。过表达Notch1可逆转过表达miR-128-3p对Ishikawa细胞存活率、凋亡率及cyclinD1和cleaved-caspase-3蛋白表达的影响。过表达miR-128-3p可降低Ishikawa细胞中β-catenin蛋白表达,而过表达Notch1可逆转过表达miR-128-3p对Ishikawa细胞β-catenin蛋白表达的影响。结论:过表达miR-128-3p可抑制EC细胞增殖,并促进细胞凋亡,其可能通过抑制Notch1表达和Wnt/β-catenin信号通路发挥作用。Objective:To investigate the effects and mechanism of miR-128-3 p on proliferation and apoptosis of endometrial cancer(EC)cells.Methods:Human endometrial vascular endothelial cells hEEC and EC cell lines HEC-1 A,Ishikawa and AN3 CA were cultured. RT-qPCR detected the expression levels of miR-128-3 p and Notch1 mRNA in cells,and Western blot assay detected the expression level of Notch1 protein. To construct Ishikawa cells overexpressing miR-128-3 p or low expressing Notch1,miR-128-3 p mimic or Notch1 small interfering RNA was transfected into Ishikawa cells. MTT,flow cytometry,Western blot detected the effects of overexpressing miR-128-3 p or low expressing Notch1 on Ishikawa cell proliferation,apoptosis and the expression of cyclinD1,cleaved-caspase-3 and β-catenin protein. Dual luciferase reporter gene assay verified the relationship between miR-128-3 p and Notch1 in Ishikawa cells.Results:Compared with hEEC cells,the expression levels of miR-128-3 p in endometrial cancer cell lines HEC-1 A,Ishikawa and AN3 CA were decreased(P<0.05),and the expression levels of Notch1 mRNA and protein were increased(P<0.05). After overexpression of miR-128-3 p or low expression of Notch1,Ishikawa cell survival rate and cyclinD1 protein level were decreased(P<0.05),apoptosis rate and cleaved-caspase-3 protein level were increased(P<0.05). miR-128-3 p targets negative regulation of Notchl expression in Ishikawa cells. Overexpression of Notch1 reversed the effects of overexpression of miR-128-3 p on Ishikawa cell viability,apoptotic rate,and the expression of cyclinD1 and cleaved-caspase-3 protein expression. Overexpression of miR-128-3 p reduced the expression of β-catenin protein in Ishikawa cells,while overexpression of Notch1 reversed the effect of overexpression of miR-128-3 p on the expression of β-catenin protein in Ishikawa cells.Conclusion:Overexpression of miR-128-3 p can inhibit the proliferation of endometrial cancer cells and promote apoptosis,which may play a role in inhibiting Notch1 expression and Wnt/β-catenin

关 键 词：子宫内膜癌 miR-128-3p NOTCH1 细胞增殖 凋亡 

分 类 号：R737.33[医药卫生—肿瘤]