Dehydroepiandrosterone 7α- and 7β-hydroxylation in mouse brain microsomes.: Effects of cytochrome P450 inhibitors and structure-specific inhibition by steroid hormones

Presently, several works question the effects of dehydroepiandrosterone (DHEA) reported in vivo and designate its 7-hydroxylated metabolites as native antiglucocorticoids and potent mediators in the triggering of immune response. Among mouse tissues and organs, and second to liver, the largest production of 7 alpha- and 7 beta-hydroxylated derivatives of DHEA takes place in brain microsomes. To contribute to identification of cytochromes P450 (CYPs) responsible for 7 alpha- and 7 beta-hydroxy-DHEA production, effects of CYP inhibitors and of several steroid hormones on DHEA 7-hydroxylation were examined. Using mouse brain microsomes as a source of enzyme, we report now that strong and smaller inhibitions of DHEA 7 alpha-hydroxylation were obtained with ketoconazole and alpha-naphthoflavone, respectively, and that neither changed DHEA 7 beta-hydroxylation. Metyrapone and antipyrine also inhibited 7 alpha-hydroxylation, but by contrast, significantly increased 7 beta-hydroxylation of DHEA, This indicated that at least, two different CYPs were responsible for 7 alpha- and 7 beta-hydroxylation of DHEA. Steroids sharing a 3 beta-hydroxylated structure with DHEA, namely pregnenolone, 5-androstene-3 beta,17 beta-diol and 3 beta-hydroxy-5 alpha-androstan-17-one, were strong inhibitors of DHEA 7 alpha-hydroxylation (non-competitive inhibition with pregnenolone, K-i=2.0+/-0.3 mu M). In contrast, 7 beta-hydroxylation yields were not decreased by the 3 beta-hydroxysteroids tested. Moderate inhibition of 7 alpha- and 7 beta-hydroxylation was obtained with 3-oxosteroids, namely testosterone, progesterone, corticosterone and 4-androsten-3,17-dione. Taken together, these data indicate specific inhibition patterns of DHEA 7 alpha- and 7 beta-hydroxylation by CYP inhibitors and steroid hormones in mouse brain microsomes and may be used as criteria necessary for identification of the responsible CYP species.