Abstract: Objective To investigate the combined effect and mechanism of metformin (Met) and 2-deoxy-D-glucose (2DG) on cell proliferation and apoptosis in liver cancer cells HepG2 and Hep3B. Methods Wst-1 reagent was used to determine the antiproliferation effects after treatments with Met and 2DG alone or combined in HepG2 and Hep3B cells. Microscopy was used to observe cell morphological changes after treatments with Met and 2DG alone or combined in HepG2 and Hep3B cells. Cell apoptosis was observed by flow cytometry after treatment of different kinds of drugs. Western blotting was used to analyze the protein expressions of Caspase3, PARP, Mcl-1 of HepG2. Results The survival rate of HepG2 cells in the combination group was (22.48±0.51)%, and compared with the control group (100.00±5.05)%, Met group (80.68±5.10)% and 2DG group (72.56±4.34)%, the differences were statistically significant (P＜0.001; P＜0.001; P=0.001). The survival rate of Hep3B cells in the combination group was (29.16±1.34)%, and compared with the control group (100.00±1.23)%, Met group (59.58±1.92)% and 2DG group (33.87±1.95)%, the differences were statistically significant (P＜0.001; P＜0.001; P=0.001). Microscopy observation showed that combined treatment of Met and 2DG caused less viable adherent cells of HepG2, but more floating dead cells. While the combination group also caused a decrease in the density of Hep3B cells, but did not significantly increase the shedding of cells. The apoptosis of HepG2 cells in the combination group was (39.63±0.21)%, and compared with the control group (7.12±0.14)%, Met group (12.56±0.35)% and 2DG group (15.16±1.93)%, the differences were statistically significant (P＜0.001; P＜0.001; P=0.001). The apoptosis of Hep3B cells in the combination group was (12.58±1.03)%, and compared with the control group (2.82±0.51)% and Met group (8.98±0.86)%, the differences were statistically significant (P＜0.001; P=0.007), but compared with the 2DG group (12.40±1.78)%, the difference was not statistically significant (P=1.000). Furthermore, Western blotting demonstrated that the combined treatment induced evident Caspase-3 activation and poly ADP-ribose polymerase （PARP） cleavages, and decreased expression of Mcl-1. Conclusion The combination of Met and 2DG can effectively inhibit cell proliferation of HepG2 and Hep3B, and induce apoptosis of HepG2 cells. The mechanism may be involved with Caspase-3 activation, cutting PARP substrate and decreasing Mcl-1 protein.

Key words: Liver neoplasms, Apoptosis, Metformin, 2-deoxy-D-glucose