Figure 4.

Bufalin and MK2206 surpassed IL-6-mediated cell growth and induced cooperative apoptosis of MM cells in the absence of BMSCs. (a and b) H929 and U266 cells were stimulated with IL-6 (10 ng/ml) for 1 h and subsequently cultured with the indicated reagents for 48 h. Cell viability was measured by the CCK8 assay. Cell lysates were subjected to western blot analysis using anti-p-AKT, anti-AKT, and β-actin Abs. β-actin was used as a loading control. The values under the bands were representative of the mean quantitation ratios compared with the control groups. (c) U266 cells were treated with 12 nM of bufalin in the absence and/or presence of 6 μM of MK2206 for 48 h and subsequently centrifuged to obtain the supernatant. The samples were subjected to an IL-6 ELISA. (d and f) H929 and U266 cells were co-cultured with or without BMSCs for 48 h with 12 nM of bufalin in the presence and/or absence of 6 μM of MK2206, and the apoptotic rates were measured by flow cytometry. Subsequently, the cell lysates were subjected to western blot using anti-p-AKT, anti-AKT and anti-β-actin Abs. β-actin was used as a loading control. The values under the bands were representative of the mean quantitation ratios compared with the control groups. Each bar represented the mean±S.E. of triplicate experiments (*P<0.05; **P<0.01)