Kinetic evidence for heterogeneous responsiveness of mixed function oxidase isozymes to inhibition and induction by allylisopropylacetamide in chick embryo liver
Mixed Function Oxygenases
Changes in hepatic mixed function oxidase kinetics after administration of allylisopropylacetamide (AIA) to chick embryos indicate that the activities of different cytochrome P-450 isozymes, including those participating in the metabolism of the same substrates, can be simultaneously increased and inhibited by a single xenobiotic. Up to 4 h after administration in ovo, or in vitro, AIA exclusively inhibited mixed function oxidases. At 24 h after administration in ovo, AIA simultaneously decreased the Vmax of the isozymes active in 7-ethoxycoumarin deethylation and in biphenyl and antipyrine hydroxylations in control liver and caused new isozymes with higher Km and Vmax values to appear. At the same time, AIA increased the Vmax values for isozymes active in aminopyrine demethylation and decreased the Vmax for benzo(a)pyrene hydroxylation (EC 188.8.131.52). As an inhibitor, AIA did not exhibit substrate selectivity but tended to inhibit isozymes with higher substrate affinity noncompetitively and lower affinity isozymes competitively. Competitive mechanisms and generalized P-450 breakdown could only partially account for the inhibition of mixed function oxidases by AIA. The inhibition at low doses of AIA (0.1 to 0.3 mg/egg) occurred without any decrease in P-450 and at higher doses it exceeded and was more persistent than the decrease in P-450. The data indicate that in addition to the known mechanisms for mixed function oxidase inhibition by AIA there is another noncompetitive mechanism independent of P-450 breakdown. As an inducer, AIA, like phenobarbital rather than beta-naphthoflavone increased the metabolism of aminopyrine and the concentration of Mr = 50,000 and 51,000 proteins preferentially. However, unlike either, AIA selectively induced new high Km and Vmax isozymes active toward 7-ethoxycoumarin, biphenyl, and antipyrine and increased the concentration of a Mr = 53,000 protein. These actions distinguish AIA from either the phenobarbital or polycyclic hydrocarbon class of inducers. The simultaneous inhibition by AIA of higher affinity isozymes with selective induction of low affinity isozymes produced a "crossover effect" in which after AIA administration the rates of 7-ethoxycoumarin deethylase and biphenyl and antipyrine hydroxylases were decreased at low and increased at high substrate concentrations. The findings demonstrate the complexity and selectivity of AIA's actions as a mixed function oxidase inhibitor and inducer and illustrate the potential heterogeneity of responses that can occur in the mixed function oxidase system after exposure of an organism to a xenobiotic.