Generation of reactive dysmorphogenic intermediates by rat embryos in culture: Effects of cytochrome P-450 inducers

Abstract

We have investigated the capacity of cultured whole rat embryos to convert 2-acetylaminofluorene (AAF) to reactive metabolites capable of eliciting dysmorphogenic effects in the same embryos. Cultured embryos (Sprague-Dawley) were exposed to AAF for periods of 2 or 24 hr, after which metabolites were isolated from the culture medium and identified with HPLC. Embryotoxic effects were evaluated in the same embryos. Day 10 embryos preexposed in utero to pregnenolone-16α-carbonitrile (PCN) exhibited marked increases in capacity to convert AAF to a variety of hydroxylated metabolites. 3-Methylcholanthrene (3MC) was also a very effective inducer in utero but Aroclor 1254 (PCB), and isosafrole (ISF) evoked only minimal induction while phenobarbital (PB) was not demonstrably effective. Exogenously added hepatic postmitochondrial supernatant (S9) fractions from adult male rats pretreated with PCB, 3MC, or ISF exhibited induced monooxygenase activities as well as increased capacity to convert AAF to dysmorphogenic intermediates in the culture system. PB and PCN displayed much lesser effects. PCN was a very effective inducer of hepatic monooxygenases of pregnant rats but, when this tissue was utilized as an enzyme source, no significant increase in malformations was observed. Embryos with relatively high monooxygenase activities also displayed a high incidence of embryonic abnormalities when cocultured with AAF. Malformation incidence was strongly correlated with hydroxy metabolite generation, suggesting that induction in utero of P-450-dependent, embryonic monooxygenases resulted in the production of embryotoxic metabolites by the embryos own enzymes. The data also indicated that endogenous bioactivation (within the conceptus) was considerably more effective than bioactivation effected by an exogenous (hepatic) enzyme source.