Most of the
chemotherapy treatments for
bladder cancer aim to kill the
cancer cells, but a high recurrence rate after medical treatments is still occurred.
Bufalin from the skin and parotid
venom glands of toad has been shown to induce apoptotic cell death in many types of
cancer cell lines. However, there is no report addressing that
bufalin induced cell death in human
bladder cancer cells. The purpose of this study was investigated the mechanisms of
bufalin-induced apoptosis in a human
bladder cancer cell line (T24). We demonstrated the effects of
bufalin on the cell growth and apoptosis in T24 cells by using
DAPI/TUNEL double staining, a PI exclusion and flow cytometric analysis. The effects of
bufalin on the production of
reactive oxygen species (ROS), the level of mitochondrial membrane potential (ΔΨ(m)), and
DNA content including sub-G1 (apoptosis) in T24 cells were also determined by flow cytometry. Western blot analysis was used to examine the expression of G(0)/G(1) phase-regulated and apoptosis-associated
protein levels in
bufalin-treated T24 cells. The results indicated that
bufalin significantly decreased the percentage of viability, induced the G(0)/G(1) phase arrest and triggered apoptosis in T24 cells. The down-regulation of the
protein levels for
cyclin D, CDK4,
cyclin E, CDK2, phospho-Rb, phospho-AKT and Bcl-2 with the simultaneous up-regulation of the
cytochrome c, Apaf-1, AIF,
caspase-3, -7 and -9 and
Bax protein expressions and
caspase activities were observed in T24 cells after
bufalin treatment. Based on our results,
bufalin induces apoptotic cell death in T24 cells through suppressing AKT activity and anti-apoptotic Bcl-2
protein as well as inducing pro-apoptotic
Bax protein. The levels of
caspase-3, -7 and -9 are also mediated apoptosis in
bufalin-treated T24 cells. Therefore,
bufalin might be used as a therapeutic agent for the treatment of human
bladder cancer in the future.