Zoledronic Acid Regulates Autophagy and Induces Apoptosis in Colon Cancer Cell Line CT26

Abstract

Zoledronic acid (ZOL) is the third generation of bisphosphonates, which can inhibit many tumors growth, especially to inhibit the growth of colon cancer. However, the molecular mechanism is still very mysterious. In this study, we observed that ZOL could regulate CT26 colon cancer cells autophagy, promote CT26 cells apoptosis, and inhibit CT26 cells proliferation. Western blotting analysis showed that proapoptosis protein caspase-3 was basically unchanged, whereas the expression of the activated caspase-3 was significantly increased, after CT26 cells were treated with different doses of zoledronic acid. Western blot also showed that ZOL could significantly affect the expression of p-p53 and autophagy-related proteins beclin-1 and p62. In conclusion, the antitumor effect of ZOL on CT26 colon cancer cells in vitro is achieved by apoptosis induction and autophagy regulation, resulting in inhibition of cell proliferation.

Figure 1

CT26 cells were exposed to the increasing ZOL concentrations (0–250 μM) for 24 h and were then processed for MTS assay (n = 5). The bars represent the means ± SD from two independent experiments. ^∗∗p < 0.01 versus the untreated control cells.

Figure 2

CT26 cells were exposed to the increasing ZOL concentrations for 24 h and were then processed for FCM (n = 5). The bars represent the means ± SD from two independent experiments. ^∗∗p < 0.01 versus the untreated control cells.

Figure 3

The cells were exposed to the 200 μMZOL and at 12 h, 24 h, 36 h, and 48 h of exposure, the cell fractions were prepared and analyzed by 15% SDS-PAGE followed by Western blotting. (a and c) The level of c-caspase-3 in CT26 cells after being exposed to ZOL. (b and d) The level of p-p53 in CT26 cells after being exposed to ZOL. The data shown in C and D are the mean ± SD of the results of three independent experiments, respectively (^∗p < 0.05 represents significant differences between the experimental and untreated control values).

Figure 4

The cells were exposed to the 200 μMZOL and at 12 h, 24 h, 36 h, and 48 h of exposure, the cell fractions were prepared and analyzed by 15% SDS-PAGE followed by Western blotting. (a and c) The level of P62 in CT26 cells after being exposed to ZOL. (b and d) The level of beclin-1 in CT26 cells after being exposed to ZOL. The data shown in C and D are the mean ± SD of the results of three independent experiments, respectively (^∗p < 0.05 represents significant differences between the experimental and untreated control values).