D-MANNITOL, A SPECIFIC HYDROXYL FREE-RADICAL SCAVENGER, REDUCES THE DEVELOPMENTAL TOXICITY OF HYDROXYUREA IN RABBITS

Hydroxyurea (HU) is a potent mammalian teratogen. Within 2-4 hours aft er maternal injection, HU causes 1) a rapid episode of embryonic cell death and 2) profound inhibition of embryonic DNA synthesis. A variety of antioxidants delays the onset of embryonic cell death and reduces the incidence of birth defects. Antioxidants do not block the inhibiti on of DNA synthesis, indicating that early embryonic cell death is not caused by inhibited DNA synthesis. We have suggested that some HU mol ecules may react within the embryo to produce H2O2 and subsequent free radicals, including the very reactive hydroxyl free radical. The free radicals could cause the early cell death; antioxidants are believed to terminate the aberrant free radical reactions resulting in lessened developmental toxicity. To investigate whether hydroxyl free radicals cause the early episode of cell death, pregnant New Zealand white rab bits were injected subcutaneously on gestational day 12 with a teratog enic dose of HU (650 mg/kg) in the presence or absence of 550 mg/kg of D-mannitol (Man), a specific scavenger of hydroxyl free radicals. Osm otic control rabbits received HU plus 550 mg/kg of xylose (Xyl, a nona ctive aldose). At term, the teratologic effects of HU were ameliorated by Man as evidenced by decreased incidences of the expected limb malf ormations. Xyl exerted no de monstrable effect on HU teratogenesis. Hi stological examination of limb buds at 3-8 hours after maternal inject ion, showed that Man delayed the onset of HU-induced cell death by as much as 4 hours. Xyl had no effect. That Man acts within the embryo wa s shown by performing intracoelomic injections on alternate implantati on sites with Man, Xyl, or saline followed by subcutaneous injection o f the pregnant doe with HU. Embryos were harvested 3-8 hours later. Li mb buds from saline- and Xyl-injected embryos exhibited the typical pa ttern of widespread HU-induced cell death at 3-4 hours, whereas Man-in jected embryos did not exhibit cell death until 5-8 hours. These resul ts are consistent with those reported for antioxidant-mediated amelior ation of HU-induced developmental toxicity and with the hypothesis tha t hydroxyl free radicals are the proximate reactive species in HU-indu ced early embryonic cell death. (C) 1994 Wiley-Liss, Inc.