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CN 22-1108/R
Volume 39 Issue 8
Aug.  2023
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Article Navigation >  Journal of Clinical Hepatology  > 2023  >  39(8): 1895-1902

Expression of miR-409-3p in hepatoma HepG2 cells and its mechanism in cell proliferation

DOI: 10.3969/j.issn.1001-5256.2023.08.019
  • 1.

    Department of Gastroenterology, Yancheng First People's Hospital, Yancheng, Jiangsu 224008, China

  • 2.

    Department of Gastroenterology, Nanjing Pukou People's Hospital, Nanjing 211899, China

  • 3.

    Medical Center for Digestive Diseases, The Second Affiliated Hospital of Nanjing Medical University, Nanjing 210011, China

More Information
  • Corresponding author: LIU Zheng, liuzheng117@126.com (ORCID: 0000-0002-2195-4538)
  • Received Date: 2022-11-05
  • Accepted Date: 2023-02-27
  • Published Date: 2023-08-20
    Abstract
  •   Objective  To investigate the expression and significance of miR-409-3p in hepatoma carcinoma cell lines and possible molecular mechanism.  Methods  Quantitative real-time PCR was used to measure the expression of miR-409-3p in normal LO2 hepatocytes and four hepatoma cell line (HepG2, BEL-7402, SMMC-7721, and MHCC-97H). Hepatoma HepG2 cells were transiently transfected with miR-409-3p mimics and microRNA mimics control using the cationic liposome method, and then CCK8, plate colony formation assay, and flow cytometry were used to observe the effect of miR-409-3p on the proliferation, cell cycle, and apoptosis of hepatoma cells in vitro. Western blot was used to measure the change in the expression of c-Met protein in HepG2 cells with overexpression of miR-409-3p, and luciferase reporter gene assay was used to identify targeting relationship. The independent-samples t test was used for comparison between two groups, and a one-way analysis of variance was used for comparison between multiple groups, followed by the SNK test.  Results  Based on the TCGA microRNA expression profile data of liver cancer, the expression level of miR-409-3p in liver cancer tissue was significantly lower than that in adjacent tissue (t=7.752, P < 0.05). Compared with the LO2 cells, the HepG2, SMMC-7721, MHCC-97H, and BEL-7402 cells had a significant reduction in the expression level of miR-409-3p (F=31.043, P < 0.05). Compared with the miR-con group, the HepG2 cells transfected with miR-409-3p mimics had a significant increase in the expression level of miR-409-3p (t=-8.836, P < 0.05), suggesting that the interference was effective. CCK8 assay showed that compared with the miR-con group, the HepG2 cells transfected with miR-409-3p mimics had a significant reduction in proliferative capacity at 48, 72, and 96 hours (t=2.876, 3.359, and 3.707, all P < 0.05). Plate colony formation assay showed that the miR-409-3p mimics group had a significantly lower plating efficiency than the miR-con group (t=2.846, P=0.047). Flow cytometry showed that compared with the miR-con group, overexpression of miR-409-3p resulted in the increased number of HepG2 cells in G2 phase (t=-3.763, P < 0.05), while there was no significant difference in apoptosis rate (t=0.714, P=0.515). Luciferase reporter gene assay showed that c-Met was a target gene of miR-409-3p (t=4.970, P=0.007). Compared with the miR-con group, the HepG2 cells transfected with miR-409-3p mimics had a significant reduction in the expression of c-Met protein (t=-8.509, P=0.001).  Conclusion  By inhibiting the protein expression of c-Met, miR-409-3p regulates downstream signaling pathways to induce cell cycle arrest in G2 phase and thus inhibits the proliferation of hepatoma HepG2 cells.

     

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