20(S)-Ginsenoside Rg3 Partially Induces Maturation of HepG2 Cells via the AMPK/HNF4A Pathway.

Background: We aimed to investigate whether 20(S)-ginsenoside Rg3 (Rg3) reduced heterogeneity by inducing the maturation of HepG2 cells via the AMP-activated protein kinase (AMPK)/hepatocyte nuclear factor 4A (HNF4A) pathway.

Methods: This in vitro cell research study was conducted under the guidance of Hebei Medical University, Hebei, China, from Sep 2022 to Dec 2023. HepG2 cells were treated with varying concentrations of Rg3 in a low glucose microenvironment. The mRNA expression of ALBUMIN (ALB, a marker for hepatocyte function) and HNF4A (a marker for differentiation of HCC cell), and AMPK protein levels were measured after significant changes in cell morphology were observed. Additionally, 5-Aminoimidazole-4-carboxamide1-β-D-ribofuranoside (AICAR) (an AMPK agonist) and Compound C (an AMPK inhibitor) were used to explore further the underlying mechanism.

Results: Under treatment of 5 μM, 10 μM, and 20 μM Rg3, some cells became flattened and larger, and there was an increase in the mRNA expression of ALB and HNF4A (P<0.05). However, there was a decreasing trend in AMPK protein content with 8 μM Rg3 (P<0.05). Compared to the control group, some cells exposed to 8 μM Rg3 exhibited pronounced morphological changes, along with upregulated expression of ALB and HNF4A mRNA. However, no such changes were observed when 8 μM Rg3 was combined with 1.6 mM AICAR. Compared to the control group, 10 μM Compound C or 8 μM Rg3 treatments led to similar changes in cell morphology and showed an increasing trend in HNF4A mRNA expression. Additionally, after treatment with Compound C, pHNF4A was mainly in the nucleus, while after Rg3 treatment, it was mostly in the cytoplasm.

Conclusion: Rg3 partially induced the maturation of HepG2 cells through the AMPK/HNF4A pathway.