The miR-758/astrocyte elevated gene-1 signaling pathway regulates metastasis of human hepatocellular carcinoma HepG2 cells / 中华肝胆外科杂志

ABSTRACT

Objective To investigate the mechanism of miR-758 in hepatocellular carcinoma cell HepG2,and to investigate the regulatory role of miR-758 on astrocyte elevated gene-1 (AEG-1).Methods Transient transfection of miR-758 into HepG2 cells was performed to study the effect of miR-758 on tumor cell metastasis by transwell migration and invasion experiments.CCK8 assay was used to detect the cell proliferation activity.The cell cycle was analyzed by flow cytometry.The effect of miR-758 on epidermal mesenchymal transition (EMT) was determined by the expression of EMT markers.Transient transfection of miR-758 into human umbilical vein endothelial cells (HUVECs) was performed to explore the effect of miR-758 on luminal formation.AEG-1 3'UTR containing the binding site of miR-758 was constructed into luciferase expression vector.The miR-758 and the vector was co-transfected into HepG2 cells.And then the change in expression level of AEG-1 protein was detected through Western Blot.Results The overexpression of miR-758 inhibited HepG2 cell migration and invasion,as well as the cell proliferation and the cell cycle.The miR-758 was also found to inhibit EMT of HepG2 cells and the lumen formation of HUVEC cells.After the co-transfection of miR-758 with the plasmid containing AEG-1 gene 3'UTR into HepG2 cells,the luciferase expression was decreased.The luciferase expression was restored when the binding site of miR-758 in the 3'UTR was mutated.Further evidence by Western Blot showed the protein level of AEG-1 in HepG2 cells was significantly decreased after transfection of miR-758.Conclusions The miR-758 negatively regulates multiple steps during cancer metastasis,including cell migration,invasion,cell proliferation,EMT,as well as angiogenesis.And AEG-1 has been identified as a downstream target of miR-758.