LncRNA NNT-AS1通过miR-582-5p/MCL1对肝癌细胞侵袭和转移的影响

Effects of LncRNA NT-AS1 on invasion and metastasis of liver cells via miR-582-5p/MCL1

目的探讨LncRNA NNT-AS1通过miR-582-5p/MCL1轴对肝癌的侵袭和转移的影响。方法培养正常肝L-02细胞和肝癌细胞MHCC97H、LM3、SMCC7721、Huh7、Hep3B,选取肝癌Huh7细胞系,将细胞进行分组与转染;采用qRT-PCR检测各组细胞中NNT-AS1、miR-582-5p表达,WB检测各组细胞MCL1蛋白表达,RIP检测LncRNA NNT-AS1和miR-582-5p的结合情况,双荧光素酶报告基因实验验证miR-582-5p和MCL1靶向关系,Transwell实验检测细胞侵袭和转移能力。结果与正常肝细胞L-02比较,NNT-AS1在肝癌细胞系中高表达,miR-582-5p在肝癌细胞系中低表达;NNT-AS1和miR-582-5p沉默后,细胞中NNT-AS1和miR-582-5p表达显著降低,细胞侵袭和转移能力也明显减弱;LncRNA NNT-AS1可结合吸附miR-582-5p,MCL1是miR-582-5p的下游靶基因、且MCL1在肝癌细胞系中高表达;干扰LncRNA NNT-AS1或过表达miR-582-5p表达可抑制肝癌细胞侵袭和迁移;同时沉默NNT-AS1和过表达miR-582-5p,可显著降低细胞侵袭和迁移能力;干扰MCL1也可抑制肝癌细胞侵袭和迁移能力增强的现象。结论沉默LncRNA NNT-AS1可通过miR-582-5p作用,抑制MCL1表达从而抑制肝癌细胞侵袭和转移。

Objective To investigate the effect of LncRNA NT-AS1 on invasion and metastasis of liver cancer through miR-582-5p/MCL1 axis.Methods Normal liver cells L-02 cells and liver cancer cells MHCC97H,LM3,SMCC7721,Huh7,and Hep3B were cultured.The Huh7 cell line of liver cancer was selected,and such cells were grouped and transfected.The expressions of NNT-AS1 and miR-582-5p in each group were detected by qRT-PCR.The expression of MCL1 protein in each group was detected by WB.The binding of LncRNA NNT-AS1 and miR-582-5p was detected by RIP.The targeting relationship between miR-582-5p and MCL1 was verified by dual luciferase reporter gene assay.Transwell assay was employed to detect cell invasion and metastasis.Results Compared with normal liver cell L-02,NNT-AS1 was highly expressed in liver cancer cell lines,and miR-582-5p was lowly expressed in liver cancer cell lines.After the silence of NNT-AS1 and miR-582-5p,the expression of NNT-AS1 and miR-582-5p were significantly decreased,and the cell invasion and metastasis ability was also significantly weakened.LncRNA NNT-AS1 could bind and adsorb miR-582-5p.MCL1 was a downstream target gene of miR-582-5p,and MCL1 was highly expressed in liver cancer cell lines.Interfering with LncRNA NNT-AS1 or overexpressing of miR-582-5p could inhibit the invasion and migration of liver cancer cells.Simultaneous silencing of NNT-AS1 and overexpressing of miR-582-5p could significantly reduce cell invasion and migration.Interfering with MCL1 could also inhibit the invasion and migration of liver cancer cells.Conclusion Silencing LncRNA NNT-AS1 can inhibit the expression of MCL1 through miR-582-5p,thereby inhibiting the invasion and metastasis of liver cancer cells.