MicroRNA-325-3p Inhibits Cell Proliferation and Induces Apoptosis in Hepatitis B Virus-related Hepatocellular Carcinoma by Down-regulation of Aquaporin 5

Background: Hepatitis B virus (HBV) infection is acknowledged as the main cause of hepatocellular carcinoma (HCC). Moreover, previous studies have revealed that microRNAs (miRNAs) widely participate in regulation of various cellular processes, such as viral replication. Hence, the purpose of this study was to investigate the roles of aquaporin 5 (AQP5) and miR-325-3p in the proliferation and apoptosis of HBV-related HCC cells.

Methods: AQP5 and miR-325-3p expression in both normal and HBV-HCC tissues or cells (both Huh7-1.3 and HepG2.2.15) was detected using qRT-PCR. AQP5 expression was knocked down in HBV-related Huh7-1.3 and HepG2.2.15 cells using small interfering RNA (siRNA) technology. Down-regulation was confirmed using real-time PCR and Western blot analysis. Effects of AQP5 down-regulation on the proliferation and apoptosis were assessed. Dual luciferase reporter gene assay, Western blot and qRT-PCR were employed to evaluate the effect of miR-325-3p on the luciferase activity and expression of AQP5. Moreover, miR-325-3p mimic-induced changes in cellular proliferation and apoptosis were detected through CCK-8 assay, BrdU assay, flow cytometry analysis and ELISA.

Results: In this study, the expression of AQP5 was up-regulated in human HBV-HCC tissue, Huh7-1.3 and HepG2.2.15 cells. Knockdown of AQP5 significantly inhibited the proliferation and promoted apoptosis of HBV-HCC cells. Next, miR-325-3p was obviously down-regulated in HBV-HCC. In concordance with this, MiR-325-3p directly targeted AQP5, and reduced both mRNA and protein levels of AQP5, which promoted cell proliferation and suppressed cell apoptosis in HCC cells. Overexpression of miR-325-3p dramatically inhibited cell proliferation and induced cell apoptosis.

Conclusions: Our findings clearly demonstrated that introduction of miR-325-3p inhibited proliferation and induced apoptosis of Huh7-1.3 and HepG2.2.15 cells by directly decreasing AQP5 expression, and that silencing AQP5 expression was essential for the pro-apoptotic effect of miR-325-3p overexpression on Huh7-1.3 and HepG2.2.15 cells. It is beneficial to gain insight into the mechanism of HBV infection and pathophysiology of HBV-related HCC.