摘要
目的探讨miR-129-3p靶向LPAR3调控肝癌细胞增殖、迁移和侵袭的分子机制。方法qRT-PCR和Western blotting检测正常肝细胞HL-7702和3种肝癌细胞SMMC-7721、HepG2和BEL-7402中miR-129-3p和LPAR3的表达情况。构建过表达miR-129-3p的SMMC-7721细胞株,MTT法检测细胞增殖活力,Transwell法检测细胞迁移和侵袭能力,Western blotting检测LPAR3、Cyclin D1、MMP-2、PI3K和AKT蛋白的表达。采用双荧光素酶报告基因法和Western blotting验证miR-129-3p和LPAR3的靶向关系。结果与正常肝细胞相比,肝癌细胞中miR-129-3p的表达显著降低,LPAR3的表达显著升高。过表达miR-129-3p可抑制SMMC-7721的增殖、迁移和侵袭。LPAR3是miR-129-3p的靶基因,miR-129-3p可负性调控LPAR3的表达。过表达LPAR3可部分逆转miR-129-3p对SMMC-7721细胞的增殖、迁移和侵袭的抑制作用。miR-129-3p通过调控LPAR3抑制PI3K和AKT蛋白的表达。结论miR-129-3p通过靶向下调LPAR3抑制PI3K/AKT信号通路活化,进而抑制肝癌细胞的增殖、迁移和侵袭。
Objective To investigate the molecular mechanism of miR-129-3p targeting LPAR3 on regulation of the proliferation,migration and invasion of hepatoma cells.Methods The expressions of miR-129-3p and LPAR3 in normal liver cell line HL-7702 and three kinds of hepatoma cells SMMC-7721,HepG2 and BEL-7402 were detected by qRT-PCR and Western blotting.The SMMC-7721 cell line with over-expression of miR-129-3p was constructed.The cell proliferation activity was detected by MTT assay.The cell migration ability and invasion ability were detected by Transwell.The expressions of LPAR3,Cyclin D1,MMP-2,PI3K and AKT proteins were detected by Western blotting.The targeting relationship between miR-129-3p and LPAR3 was verified by dual luciferase reporter gene assay and Western blotting.Results Compared with normal hepatocytes,the expression of miR-129-3p was significantly decreased,while the expression of LPAR3 was significantly increased in hepatoma cells.The over-expression of miR-129-3p inhibited the proliferation,migration and invasion of SMMC-7721 cells.LPAR3 was a target gene of miR-129-3p,and miR-129-3p negatively regulated the expression of LPAR3.The over-expression of LPAR3 partially reversed the inhibitory effect of miR-129-3p on proliferation,migration and invasion of SMMC-7721 cells.miR-129-3p inhibited the expressions of PI3K and AKT proteins by modulating LPAR3.Conclusion miR-129-3p inhibits the activation of PI3K/AKT signaling pathway by down-regulating LPAR3,thereby inhibits the proliferation,migration and invasion of hepatoma cells.
作者
白璐
马英杰
王郁杰
BAI Lu;MA Yingjie;WANG Yujie(Department of Gastroenterology,People’s Hospital of Zhengzhou,Zhengzhou 450000,China)
出处
《胃肠病学和肝病学杂志》
CAS
2020年第1期7-14,共8页
Chinese Journal of Gastroenterology and Hepatology
