Beta-naphthoflavone induction of a cytochrome P-450 arachidonic acid epoxygenase in chick embryo liver distinct from the aryl hydrocarbon hydroxylase and from phenobarbital-induced arachidonate epoxygenase.
Cytochrome P-450-mediated arachidonic acid metabolism in chick embryo liver microsomes was increased by both Ah receptor-dependent (2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and beta-naphthoflavone) and independent (phenobarbital) P-450 inducers. Arachidonic acid epoxides and monohydroxyeicosatetraenoic acids were increased 9-12-fold. omega-1-OH arachidonic acid was also significantly increased by TCDD and beta-naphthoflavone while omega-OH arachidonic acid, the main metabolite in uninduced livers, was decreased by all three agents. The P-450s catalyzing the enhanced arachidonate metabolism in beta-naphthoflavone- and phenobarbital-treated liver were investigated in reconstituted systems containing wholly or partially purified P-450s. beta-Naphthoflavone induced formation of a 55-kDa P-450 selective for arachidonate metabolism and for epoxygenation in particular. This P-450 was purified (beta NFAA). It was found to be distinct from a 54.5-kDa beta-naphthoflavone-induced P-450 catalyzing aryl hydrocarbon hydroxylase and 7-ethoxyresorufin deethylase (designated NF1). Mean turnover numbers for arachidonate epoxygenase, aryl hydrocarbon hydroxylase, and 7-ethoxyresorufin deethylase were 11.2, 0.56, and 0.04, respectively, for reconstituted beta NFAA and 0.33, 11.8, and 2.4 for NF1. beta NFAA and NF1 also differed in chromatography elution characteristics and N-terminal amino acid sequences. Both were low spin, with carbon monoxide binding peaks at 448 nm. The phenobarbital-induced arachidonate epoxygenation was catalyzed by P-450 fractions containing the main 48- and 49-kDa phenobarbital-induced P-450s; fractions in which the 49-kDa P-450 predominated were the most active. Turnover numbers for arachidonic acid epoxygenation were not correlated with those for aminopyrine demethylation or 7-ethoxycoumarin deethylation for P-450s from phenobarbital-treated livers or with aryl hydrocarbon hydroxylase, 7-ethoxyresorufin deethylase, or 7-ethoxycoumarin deethylase for P-450s from beta-naphthoflavone-treated livers. Also, different P-450s catalyzed the epoxygenation and the omega-hydroxylation of arachidonic acid in both beta-naphthoflavone- and phenobarbital-treated livers. The findings support a physiologic role for P-450-induced arachidonate metabolism and provide a basis for a possible link between TCDD's induction of P-450 and alterations of cellular homeostasis.
Nakai
K
K
Department of Pharmacology, Cornell University Medical College, New York, New York 10021.
Ward
A M
AM
Gannon
M
M
Rifkind
A B
AB
D000595:Amino Acid Sequence; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D001189:Aryl Hydrocarbon Hydroxylases / Q000378:metabolism; D001571:Benzoflavones / Q000494:pharmacology*; D002642:Chick Embryo; D000090762:Cytochrome P-450 CYP2J2; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis*; D015786:Cytochromes b5 / Q000378:metabolism; D004790:Enzyme Induction / Q000187:drug effects; D008099:Liver / Q000201:enzymology; D008862:Microsomes, Liver / Q000201:enzymology; D006899:Mixed Function Oxygenases / Q000378:metabolism; D008969:Molecular Sequence Data; D010105:Oxygenases / Q000096:biosynthesis*; D010634:Phenobarbital / Q000494:pharmacology; D000072317:Polychlorinated Dibenzodioxins / Q000494:pharmacology; D019324:beta-Naphthoflavone
D001571:Benzoflavones; D000072317:Polychlorinated Dibenzodioxins; D016718:Arachidonic Acid; D019324:beta-Naphthoflavone; D015786:Cytochromes b5; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D010105:Oxygenases; D001189:Aryl Hydrocarbon Hydroxylases; D000090762:Cytochrome P-450 CYP2J2; D010634:Phenobarbital