Hsa_circ_0003998调控miR-218-5p/Wnt/β-catenin通路对肝癌细胞增殖、迁移和侵袭的影响

Effects of Hsa_circ_0003998 regulating miR-218-5p/Wnt/β-catenin pathway on proliferation, migration, and invasion of liver cancer cells

目的:探讨circ_0003998调控肝癌细胞增殖、迁移和侵袭的分子作用机制。方法:培养人肝上皮细胞THLE-2和肝癌细胞Huh7、MHCC97、Hep3B、PLC/PRF/5,采用qRT-PCR方法检测细胞中circ_0003998和miR-218-5p表达,在Huh7细胞中敲减circ_0003998和miR-218-5p的表达,采用CCK8实验检测细胞增殖活性,Transwell实验检测细胞迁移和侵袭能力,双荧光素酶实验验证circ_0003998和miR-218-5p的靶向关系,Western blot实验检测细胞中β-catenin、N-cadherin、Vimentin和E-cadherin蛋白表达。结果:circ_0003998在肝癌细胞中高表达,miR-218-5p在肝癌细胞中低表达;敲减circ_0003998能抑制肝癌Huh7细胞增殖、迁移和侵袭;在Huh7细胞中敲减circ_0003998可上调miR-218-5p的表达;抑制miR-218-5p能够逆转敲减circ_0003998对Huh7细胞增殖、迁移和侵袭的抑制作用;敲减circ_0003998通过上调miR-218-5p抑制Huh7细胞中Wnt/β-catenin通路活性和EMT。结论:敲除circ_0003998能抑制Huh7细胞增殖、迁移和侵袭,其作用机制之一是上调miR-218-5p,抑制Wnt/β-catenin通路活性和EMT。

Objective:To investigate the molecular mechanism of circ_0003998 regulating the proliferation,migration,and invasion of liver cancer cells.Methods:Human liver epithelial cells ThLE2 and liver cancer cells Huh7,MHCC97,Hep3B,and PLC/PRF/5 were cultured.The expressions of circ_0003998 and miR-218-5p were detected by qRT-PCR assay,and the expressions of circ_0003998 and miR-218-5p were knocked out in Huh7 cells.The proliferation activity of cells was detected by CCK8 assay,and the migration and invasion ability of cells was detected by Transwell assay.The targeted relationship between circ_0003998 and miR-218-5p was verified by double luciferase assay,and the protein expressions ofβ-catenin,N-cadherin,Vimentin,and E-cadherin in cells were detected by Western blot.Results:The circ_0003998 was highly expressed in liver cancer cells,while miR-218-5p was low in liver cancer cells.Knockout circ_0003998 inhibits the proliferation,migration,and invasion of Huh7 cells.Knockdown circ_0003998 upregulated the expression of miR-218-5p in Huh7 cells.Inhibition of miR-218-5p can reverse the inhibition of knockdown circ_0003998 on proliferation,migration,and invasion of Huh7 cells.Knockdown circ_0003998 inhibited Wnt/β-catenin pathway activity and EMT in Huh7 cells by upregulating miR-218-5p.Conclusion:Circ_0003998 inhibited the proliferation,migration,and invasion of Huh7 cells by up-regulating miR-218-5p and inhibiting the activity of Wnt/β-catenin signaling pathway and EMT.