HEPATIC CATECHOLESTROGEN SYNTHASES - DIFFERENTIAL EFFECT OF SEX, INDUCERS OF CYTOCHROMES-P-450 AND OF ANTIBODY TO THE GLUCOCORTICOID INDUCIBLE CYTOCHROME-P-450 ON NADPH-DEPENDENT ESTROGEN-2-HYDROXYLASE AND ON ORGANIC HYDROPEROXIDE-DEPENDENT ESTROGEN-2/4-HYDROXYLASE ACTIVITY OF RAT HEPATIC MICROSOMES

Formation of catecholestrogens (CE) by rat hepatic microsomes was re-examined because as recently shown; (1) CE formation can be catalyzed by an NADPH-dependent estrogen-4-hydroxylase (E-4-H(NADPH)) and by peroxidatic, organic hydroperoxide-dependent estrogen-2/4-hydroxylase (E-2/4-H(OHP)), in addition to the established NADPH-dependent estrogen 2-hydroxylase (E-2-H(NADPH)); and (2) the indirect radiometric and the COMT-coupled radioenzymatic assays, used in many previous studies, may fail to provide an accurate measure, in particular, of 4-OH-CE. Using a direct product isolation assay, hepatic microsomes of both male and female rats were shown to express E-2/4-H(OHP)) activity with properties similar to those of peroxidatic activity in other tisses. The activities of E-2/4-H(OHP) and E-2-H(NADPH) were affected differently by 5 out of 7 inducers of cytochromes P -450 administered in vivo. Phenobarbital and dexamethasone caused a 4- and 2-3-fold increase in E-2-H(NADPH) activity, respectively, but only a 38 and 20% increase in E-2/4-H(OHP) activity. Ketoconazol and beta-napthoflavone caused a modest increase in E-2-H(NADPH) activity but a decrease in OHP-dependent activity. Clofibrate decreased peroxidatic activity by 50% and NADPH-dependent activity by approximately 20%. Both activities were increased by ethanol but decreased by isoniazide, an agent which induces the same form of cytochromes P -450 as ethanol. Polyclonal antibody against P -450p, a form of P -450 induced by glucocorticoids, inhibited E-2-H(NADPH) but not E-2/4-H(OHP) activity of untreated and of dexamethasone- and phenobarbital-treated rats. This study establishes that CE formation may occur in liver via the peroxidatic pathway and indicates that this pathway depends on forms of P -450 different from those mediating E-2-H(NADPH) activity. It also confirms and extends previous observations of the involvement of multiple, constitutive and induced forms of cytochrome P -450 in NADPH-dependent 2-hydroxylation in liver.