HBx promotes liver tumor in mice by inhibiting microRNA-145a-5p

Abstract

Objective To investigate the changes of miRNA-145a-5p in liver progenitor cells and mice infected with hepatitis B virus X protein (HBx) at different time points to preliminarily explore the mechanism of HBx causing liver tumor. Methods HBx gene was transfected into 14-19 liver precursor cells by lentivirus vector to construct HBx-EGFP-14-19 cell line, and then the suspension of obtained cells was injected into KM mice through hepatic portal vein to establish an HBx model with a complete immune system. The liver tissues were collected on days 30, 90, 180, and 360. The expression of HBx in HBx-EGFP-14-19 cells and HBx mice was detected by qRT-PCR and Western blotting. After miR-145a-5p mimic and inhibitor was transfected into the HBx-EGFP-14-19 cells and HBx mice having been fed for 180 d, respectively, the RNA expression of miR-145a-5p, c-Myc, Kras, apoptosis-related and cell cycle-related factors was detected by qRT-PCR, and the protein expression of them were detected by Western blotting. The cell proliferation was detected by CCK8 assay. Results The cell model capable of stably expressing HBx and HBx mouse model were established successfully. Overexpression of HBx induced the down-regulation of miR-145a-5p in HBx-EGFP-14-19 cells and HBx mice, while enhanced the expression of c-Myc and Kras. MiR-145a-5p mimic/agomir inhibited cell proliferation and down-regulated the expression of c-Myc, Kras, Bcl-2, CDK4, CyclinE, CyclinD1, and up-regulated the expression of Bad and Bax, while miR-145a-5p inhibitor/antagomir led to the opposite Results from the mimic. Conclusion Overexpression of HBx leads to the occurrence of liver tumor in mice, and its mechanism may be related to its regulation of miR-145a-5p and its target genes c-Myc and Kras and inhibition for cell apoptosis.

Keywords

• hepatitis b virus x
• hepatocellular carcinoma
• mir-145a-5p
• c-myc
• kras