The biological role of autophagy in regulating and controlling the proliferation of liver cancer cells induced by bufalin.

Autophagy is an important mechanism which regulates the processes of cell growth and death. The biological role of autophagy in regulating and controlling the proliferation of liver cancer cells by bufalin remains unknown. In the present study we investigated the effect of bufalin on autophagy of liver cancer cells. The growth inhibition and autophagy of liver cancer cells were detected using acridine orange fluorescence staining, flow cytometry and transmission electron microscopy. Combined with autophagy inhibitors (3‑MA and CQ), CCK8 staining and western blot analysis were used to detect the effect of bufalin on the proliferation and autophagy‑related protein expression in HCC‑LM3 cells at the indicated time‑points. Results indicated that combined with autophagy inhibitors 3‑MA and CQ, the inhibitive effect of bufalin on the proliferation of HCC‑LM3 cells was significantly enhanced. When combined with autophagy inhibitors 3‑MA or CQ, bufalin significantly reduced the autophagosome and acidic vesicles in HCC‑LM3 cells. When combined with autophagy inhibitors 3‑MA and/or CQ for 12 h, bufalin significantly inhibited the expression of LC3‑I and Beclin‑1 in HCC‑LM3 cells, and upregulated the expression of LC3‑II and P62 in HCC‑LM3 cells. When combined with autophagy inhibitors 3‑MA and/or CQ for 24 h, bufalin significantly inhibited the LC3‑II expression in HCC‑LM3 cells, and upregulated the LC3‑I, P62 and Beclin‑1 expression in HCC‑LM3 cells. When combined with autophagy inhibitors 3‑MA and/or CQ for 48 h, bufalin significantly inhibited the expression of LC3‑II and Beclin‑1 in HCC‑LM3 cells, and upregulated the expression of LC3‑I and P62 in HCC‑LM3 cells. These findings indicated that bufalin induced cell autophagy and inhibited proliferation of liver cancer cells by influencing the expression of autophagy related proteins including LC3‑I, LC3‑II, P62 and Beclin‑1 in liver cancer cells. The autophagic state of liver cancer cells affected the inhibitory effect of bufalin on the proliferation of liver cancer cells.