Influence of Apoptosis on the Enhancement of Radiotoxocity by Ouabain

BACKGROUND: The Na+, K(+)-ATPase inhibitor ouabain enhances the toxicity of irradiation and we have previously demonstrated that the drug suppresses repair capacity. The influence of ouabain on apoptosis is not known and is examined in this study. MATERIALS AND METHODS: Seven human cell lines of defined TP53 status were irradiated with 60Co-gamma irradiation in the presence and absence of 10(-7) M ouabain. Cell survival was determined by the clonogenic assay, apoptosis by acridine orange staining and cell cycle delays by flow cytometry. RESULTS: The ouabain-induced enhancement of radiotoxicity, expressed as the ratio of SF2's, is independent of TP53 status and ranges from 1.1 to 2.8 depending upon cell line. Ouabain prolongs the irradiation-induced G2 delay in TP53 mutant tumor cell lines by a factor greater than 2, but not in the normal lung fibroblast L132, where the cell recovery is not altered in the presence of ouabain. Twenty hours post irradiation, ouabain enhances apoptosis induced by irradiation by factors of 1.3 to 1.7 depending on the cell line. CONCLUSION: Ouabain preferentially enhances the radiotoxicity in tumor cells irrespective of TP53 status. In the pattern of DNA damage responses which are influenced by ouabain we show that the G2 cell cycle delay is prolonged and that early apoptosis events are upregulated in TP53 wild type and TP53 mutant cells. It is concluded that apoptosis plays a significant role in the enhancement of radiotoxicity by ouabain.