Proliferation and apoptosis of HeLa cells induced by in vitro stimulation with digitalis

In the HeLa tumor cell line, we studied the characteristics of the dual effect of digitalis compounds on cell growth (proliferation and death). In addition, we explored whether both effects occur by means of the same mechanism. HeLa cell cultures were exposed to increasing concentrations (0.01 nM-10 mu M) of ouabain, strophantidin, digoxin, and digoxigenin at 24-96h intervals. Cell growth in treated cultures was compared with cell growth under nontreated conditions. Additionally, we studied changes in nuclear morphology, as well as in genomic DNA degradation, cytochrome c release, and caspase-9 and -3 presence and processing induced by toxic concentrations of digitalis. Digitalis compounds increased HeLa cell number when exposed to concentrations < 10 nM during a 48h period. Ethacrynic acid (a nonsteroid inhibitor for Na+/K+-ATPase) did not induce cell growth at these concentrations. Digitalis concentrations > 10nM induced cell death in a concentration- and exposure period-dependent fashion. Changes in nuclear morphology, DNA fragmentation, mitochondrial cytochrome c release, and proteolytic processing of caspases-9 and -3, suggest apoptotic cell death. The IC50 for the inducing effect of apoptosis by ouabain at 96h was 18nM and corresponds with the IC50 for the Na+/K+-ATPase inhibition in HeLa cells. In conclusion, the dual effect of digitalis compounds on HeLa cells growth is concentration and time-dependent. The apoptosis-inducing effect correlates with inhibition of Na+/K+-ATPase. Prolifieration does not appear to be mediated through this pathway. The apoptosis-induction pathway is possibly cytochrorne c-dependent. (c) 2006 Elsevier B.V All rights reserved.