Partial attenuation of hydroxyurea-induced embryotoxicity by deoxyribonucleotides in mouse and rat embryos treated in vitro

Abstract

Hydroxyurea (HU) is a well known developmental toxicant in all animal species tested. It inhibits DNA synthesis, and addition of deoxycytidine monophosphate (dCMP) has been shown previously to attenuate the developmental toxicant effects of HU in vivo. The purpose of the present investigation was to determine whether addition of deoxyadenosine monophosphate (dAMP) or dCMP would attenuate HU-induced embryotoxicity using a rodent whole embryo culture system. Rat embryos were removed on the morning of day 10 of gestation, and mouse embryos were removed on day 8 of gestation (day 0 = the day a vaginal plug was found). Embryos were treated with various concentrations of HU (up to 500 μg/ml) for 1 hr at 37 °C before being washed and cultured for 43 hr in rat serum containing dAMP or dCMP. At the end of the culture period, six endpoints were evaluated for each viable embryo: morphological score; number of somite pairs; crown-rump and head lengths, DNA and protein contents. HU (300 μg/ml) significantly decreased values for all endpoints in embryos from both mice and rats; however, mouse embryos appeared to be more sensitive to the effects of the drug. dAMP and dCMP alone produced some embryotoxicity at high concentrations. The combination of HU + dAMP had no consistent beneficial effect in rat embryos and no effect on mouse embryos. The combination of HU + dCMP improved growth and development slightly. As determined by HPLC analysis, HU treatment (300 μg/ml for 1 hr) decreased all nucleotide pools. Subsequent treatment with dAMP increased all pool levels, although these levels remained below those of control embryos. These results suggest that the developmental toxicant effects of HU are not due solely to alterations in deoxyribonucleotide pool levels.