Investigation of ouabain-induced anticancer effect in human androgen-independent prostate cancer PC-3 cells

To determine the therapeutic potential of cardiac glycosides in androgen-independent prostate cancer, we examined ouabain-induced cytotoxic effect as well as the signaling pathways in PC-3 cells. Ouabain induced a time- and concentration-dependent cytotoxicity using mitochondrial MTT reduction assays, and the effective threshold concentration was in nanomolar level. At the concentrations less than 10 nM, ouabain induced a decrease of mitochondrial activity until a 7-hr exposure was performed, while it induced a rapid drop of mitochondrial function as early as a 2-hr treatment of cells with high concentrations of ouabain suggesting the involvement of two distinct mechanisms to ouabain action. After functional examinations, the data showed that both low and high concentrations of ouabain induced an inhibition of Na+-K+ ATPase and a subsequent Ca-45(2+) influx into PC-3 cells. High concentrations of ouabain induced a significant and time-dependent loss of mitochondrial membrane potential (A,,,), a sustained production of reactive oxygen species (ROS), and severe apoptotic reaction. Ouabain also induced an increase of Par-4 (prostate apoptosis response 4) expression. Furthermore, an antisense, but not nonsense, oligomer against Par-4 expression significantly inhibited the cytotoxicity induced by low concentrations of ouabain. It is suggested that ouabain induces two modes of cytotoxic effect in human hormone-independent prostate cancer PC-3 cells. Low concentrations of ouabain induce the increase of Par-4 expression and sensitize the cytotoxicity; while high concentrations of ouabain induce a loss of DeltaPsi(m), a sustained ROS production and a severe apoptosis in PC-3 cells. (C) 2003 Elsevier Inc. All rights reserved.