FUNCTIONAL AROMATASE EXPRESSION IN PORCINE ADRENAL-GLAND AND TESTIS

The expression of aromatase cytochrome P450 (P450(arom)) in the adrena l glands, testes, and placentas of fetal and newborn pigs was investig ated. Western immunoblot analysis detected a single 48-50-kDa protein band in these tissues as well as in other porcine tissues known to exp ress P450(arom) including Day 12 tubular conceptuses, theca interna, a nd granulosa. Slight differences in migration suggested that the P450 protein expressed in the testis was larger than that in the adrenal gl and, which was, in turn, larger than that in placenta, theca, and gran ulosa. Consistent with P450(arom) expression in these tissues, a cDNA encoding porcine P450(arom) hybridized to a 2.3-kb transcript in North ern analyses of porcine blastocysts, placentas, and fetal and newborn adrenal glands and testes, as well as in theca and granulosa tissues f rom preovulatory follicles. No differences in transcript size were det ectable among tissues. The identity of P450(arom) transcripts was conf irmed by sequence analysis of partial cDNA clones amplified from porci ne fetal adrenal glands, testes, and placentas according to the RACE p rocedure. The sequences of the adrenal and testis clones were identica l but differed from the placental sequence, which represented the firs t 85 amino acids of porcine P450(arom). Specifically, the adrenal and testis clones expressed transcripts that resembled the ovarian isoform of porcine P450(arom) rather than the porcine placental isoform, pred icting a two-amino acid deletion and 12 predicted amino acid substitut ions. P450(arom) activity was examined to further define expression in these tissues. Activity in adrenal, testis, and placental homogenates was inhibited by 4-hydroxyandrostenedione (4OH-A4) whereas inhibition by etomidate was demonstrated in the adrenal and testis homogenates b ut not in the placental homogenates. The sensitivity of activity in th e newborn porcine adrenal glands and testes to inhibition by etomidate was similar to that of ovarian P450(arom) activity. The level of P450 (arom) activity was highest in the placenta and lowest in the adrenal gland, and no effect of fetal sex was noted in either tissue. Immunocy tochemical studies localized the expression of P450(arom) in the adren al gland of newborns to cells at the corticomedullary junction and, wi th greater intensity, to cells around the developing medullary lobules . The same cells expressed cytochrome P450 17 alpha-hydroxylase, the e xpression of which also extended throughout the zona fasciculata. The interstitial cells were the site of P450(arom) expression in the testi s, but no expression could be detected in any cells within the spermat ic tubules. These data demonstrate that fetal and newborn porcine adre nal glands and testes express an active P450(arom) that resembles the isoform expressed in the ovary. The localization of adrenal expression suggests a possible role in medullary maturation and function in the fetal and newborn pig.