Lack of Effects of Ciprofloxacin and the Topoisomerase II Inhibitors,m-AMSA and Nalidixic Acid, on DNA Repair in Cultured Rat Liver Cells

Abstract

Several quinolone antibiotics, including ciprofloxacin, have been reported to elicit autoradiographic unscheduled DNA synthesis (UDS) in cultured rat hepatocytes. In the present investigation, ciprofloxacin (CF), at 250–1500 μM, produced autoradiographic UDS in cultured rat hepatocytes, whereas neither the quinolone nalidixic acid norm-AMSA, both topoisomerase II inhibitors, produced autoradiographic UDS. CF also reduced cytoplasmic [3H]thymidine levels ([3H]TdR) relative to control at 250–1500 μMand concomitantly increased nuclear grain counts accounting for most of the net increase yielding positive UDS values. To obtain definitive information on whether the positive UDS observed with CF was due to DNA repair, DNA repair synthesis was measured in parental DNA separated from newly replicated DNA using a bromodeoxyuridine incorporation density gradient method. This method was used to measure DNA repair synthesis in parental DNA of both replicating rat liver epithelial cells (ARL-18) and nonproliferating rat hepatocytes in primary culture. Primary hepatocytes exposed to CF from 250 to 1500 μMdid not express DNA repair synthesis in parental DNA isolated by density gradient centrifugation but rather exhibited a concentration-related decrease in the level of [3H]TdR associated with DNA. In rat liver epithelial (ARL-18) cells, CF from 250 to 500 μMlikewise did not elicit DNA repair synthesis and also caused a concentration-related decrease in the level of [3H]TdR associated with parental DNA. In contrast, in both cell types a substantial level of repair synthesis occurred in parental DNA as a result of exposure to 2-acetylaminofluorene, a DNA-reactive carcinogen, and in hepatocytes a similar finding was made for the drug hydralazine. Also, after induction of DNA repair in hepatocytes by ultraviolet light, the DNA polymerase α inhibitor aphidicolin almost completely abolished repair synthesis, whereas CF had a negligible effect on the inhibition of repair relative to control. These results indicate that CF did not elicit authentic DNA repair and also did not inhibit DNA repair synthesis. The fact that CF elicited autoradiographic UDS and that the topoisomerase II inhibitorsm-AMSA and nalidixic acid did not indicates that effects on topoisomerase II are not the basis for the positive UDS result with CF as has been hypothesized in the past.