Cytochrome P-450-dependent formation of reactive oxygen radicals: isozyme-specific inhibition of P-450-mediated reduction of oxygen and carbon tetrachloride

Abstract

1. Ethanol-inducible P450 IIE1 exhibits a high rate of oxygen consumption and oxidase activity. The enzyme is selectively distributed in the liver centrilobular area, the acinar region specifically destroyed after treatment with P450 IIE1 substrates/inducers such as ethanol, carbon tetrachloride, chloroform, N-nitrosodimethylamine and paracetamol. 2. Twenty substrates and ligands for cytochrome P450 IIB4 and P450 IIE1 were evaluated for their ability to inhibit microsomal and reconstituted NADPH-dependent oxidase activity, and the P450 IIE1-catalysed reduction of carbon tetrachloride to chloroform. Type I ligands and substrates did not inhibit the processes whereas nitrogen-containing compounds such as octylamine, cimetidine, imidazole and tryptamine inhibited NADPH oxidation and H2O2 formation in microsomes from starved and acetone-treated rats by around 50%. 3. Tryptamine, octylamine, isoniazid and p-chloroamphetamine inhibited reconstituted P450 IIE1-dependent oxidase activity with half maximal effects at 14-170 microM. 4. Isoniazid, cimetidine and tryptamine inhibited the P450 IIE1-dependent reduction of carbon tetrachloride, whereas acetone was without effect. 5. The oxygen dependency of microsomal oxidase activity exhibited high-affinity and low-affinity phases, with partial saturation at 20 microM of O2. 6. It is concluded that microsomal oxidase activity takes place at physiological concentrations of O2 and that isozyme-specific type II ligands compete with oxygen or carbon tetrachloride for reduction by P-450 haem.