Deguelin, a natural rotenoid, inhibits mouse myeloma cell growth in vitro via induction of apoptosis

Abstract

Deguelin is a naturally occurring rotenoid with strong cancer chemopreventive and antitumor activities. In the present study, we investigated the antitumor activity of deguelin against MPC-11 murine myeloma cells and the possible mechanism of action in vitro. Our results revealed that deguelin inhibited the proliferation of MPC-11 cells in a concentration- and time-dependent manner and caused the apoptotic death of MPC-11 cells. Following exposure to deguelin, the phosphorylation of Akt was decreased. The inhibition of cell growth may be associated with decreased levels of phosphorylated Akt. Deguelin-induced apoptosis was characterized by the upregulation of Bax, downregulation of Bcl-2 and activation of caspase-3. In conclusion, deguelin inhibits murine myeloma cell proliferation by inducing apoptosis via regulation of the Bax/Bcl-2 ratio and by inhibition of the activation of Akt. Its potential as an anticancer agent against multiple myeloma warrants further investigation.

Figure 1.

Chemical structure of deguelin.

Figure 2.

Proliferation inhibitory effect of deguelin on MPC-11 cells. Concentration- and time-dependent inhibition of the proliferation of MPC-11 cells by deguelin revealed by MTT assay. Cells were seeded in 96-well-plates and treated with various concentrations of deguelin for 24, 48 or 72 h. Data are expressed as mean ± SD for at least 5 independent experiments. The asterisk indicates a significant difference between control and deguelin-treated groups (^*P<0.05 vs. control group, ^**P<0.01 vs. control group).

Figure 3.

Morphological changes induced by deguelin. (A and B) Fluorescence and (C and D) bright-field microscope images of PI-stained nuclei of MPC-11 cells following incubation with deguelin for 48 h. (A and C) 0 ng/ml deguelin; (B and D) 100 ng/ml deguelin. Original magnification, ×400. Apoptotic cells containing condensed and fragmented fluorescent nuclei are visible in deguelin-treated cells, but not in untreated cells. PI, propidium iodide.

Figure 4.

Patterns of DNA following treatment with deguelin. Following exposure to deguelin for 48 h, agarose gel electrophoresis patterns of DNA isolated from MPC-11 cells treated with (A) 0, (B) 50 and (C) 100 ng/ml deguelin. Ladder-like patterns of DNA fragments are visible in deguelin-treated cells, but not in untreated cells. M, marker.

Figure 5.

Deguelin inhibits phosphorylation of Akt. MPC-11 cells were incubated with 0–100 ng/ml deguelin for 48 h. Cells were lysed and analyzed by immunoblotting using antibodies specific for Akt. The phosphorylation of Akt was markedly inhibited by deguelin, whereas the level of total Akt was not influenced.

Figure 6.

Effects of deguelin on Bcl-2 and Bax expression and activation of caspase-3. MPC-11 cells were treated with deguelin at various concentrations (0–100 ng/ml) for 48 h and the expression levels of Bcl-2/Bax and cleaved caspase-3 were analyzed by western blotting. Total protein extracts were subjected to 15% SDS-PAGE for western blot analysis. Deguelin inhibited Bcl-2 expression in a concentration-dependent manner and simultaneously increased Bax expression. The level of cleaved caspase-3 was increased by deguelin.