Glycyrrhizin induces autophagic cell death in MHCC97-H cell line in vitro

Objective To investigate the inhibitory effect of Glycyrrhizin in MHCC97-H cell line in vitro and explore the relevant mechanism. Methods MHCC97-H cells were cultured in vitro and treated with Glycyrrhizin in different concentrations and then cell viability was assayed at different time points. The concentration and time were selected with 50% cell viability. MHCC97-H cell plate clone formation assay and invasion-migration experiment were also performed to study the tumor-suppressor efficacy of Glycyrrhizin. Acridine orange staining was used to evaluate the formation of autophagic vacuoles. Meanwhile, 3-MA and Atg7-siRNA were both employed to avoid the autophagy activation in MHCC97-H cells and cell viability was reassessed. Western-blot was carried out to study the expression of autophagic proteins of LC3B, p-mTOR and p-ERK1/2. Results It showed Glycyrrhizin significantly inhibited MHCC97-H cell viability and the concentration and time at 50% cell viability were 2 mmol/L and 48 h respectively. Clone number in Glycyrrhizin group was significantly smaller than that in the control group (176.7±14.5 vs. 410.0±32.1). Invasion-migration rate was also lower in Glycyrrhizin group compared with the control group (41.0%±3.8% vs. 100%). Autophagic vacuoles was increased in MHCC97-H cells when treated with Glycyrrhizin and expression of LC3B-Ⅱ was enhanced and LC3B-Ⅱ/Ⅰ Ratio was increased, at the same time degradation of P62 was accelerated. Reduced p-mTOR in concurrence with upregulated p-ERK1/2 could be observed in MHCC97-H cells administered with Glycyrrhi-zin. Cell groups additionally treated with 3-MA or Atg7-siRNA exerted higher cell viability (64.3% vs. 45.9% and 67.7% vs. 47.1%, respectively). Conclusion Glycyrrhizin can induce excessive autophagy in hepatocellular carcinoma cells to cause autophagic cell death and exhibit great potential in clinical application. Key words: Hepatocellular carcinoma; Glycyrrhizin; Autophagy; MHCC97-H; Signaling pathway, mTOR, ERK1/2