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A J Conley, W E Rainey, and J I Mason


This study examined fetal steroidogenic enzyme expression and function during pregnancy in the pig. Northern and Western analyses were performed to detect the cytochrome P450 enzyme 17α-hydroxylase/17–20 lyase (P450c17) and that for cholesterol side-chain cleavage (P450scc), as well as 3β-hydroxysteroid dehydrogenase (3β-HSD) expression in several porcine fetal tissues. The data demonstrate higher steroidogenic enzyme expression in the fetal adrenal glands and testes than in the placenta at all stages of development examined. Although steroidogenic enzyme expression was maintained throughout gestation in both the fetal adrenals and the testes, adrenal P450c17 expression was higher in the early and late stages when compared with the intermediate stages of fetal development. The stimulation of fetal adrenal steroidogenic enzyme expression in the later stage fetuses was accompanied by increased expression of P450c17 in both the fetal testes and placenta. The expression of 3β-HSD by porcine fetal testes was low compared with that of the fetal adrenal gland at all stages of development. Adrenal explants and cultured cells secreted cortisol and androstenedione but much lower amounts of corticosterone, dehydroepiandrosterone and aldosterone. Secretion of cortisol and androstenedione by adrenal explants was maintained by ACTH for 5 days of culture but declined in controls. In cultured porcine fetal adrenal cells, ACTH and angiotensin II stimulated the secretion of multiple steroids. Porcine fetal testis explants and cultured cells secreted testosterone, dehydroepiandrosterone and androstenedione, but were only moderately responsive to trophic stimulation by LH. In general, the data suggest that the fetal adrenal glands and the fetal testes have the potential to contribute significantly to the production of steroids during pregnancy in pigs.

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D L Johnson, A J Conley, and L L Martin

The direct electrochemistry of human, bovine and porcine cytochrome P450c17 (CYP17) has been examined on an edge-oriented pyrolytic graphite electrode. The recombinant protein was immobilized on an electrode modified with a surfactant to simulate the environment of a biological membrane, and hence physiological electron-transfer conditions. The P450 enzymes all retained ‘electron-transfer’ activity while immobilized at the electrode surface as assessed by the presence of catalytic signals under aerobic conditions. The redox potentials for porcine P450c17 were more positive (anodic) than both the human and bovine forms, perhaps reflecting the differences in substrate specificity for these species. In addition, these enzymes were all influenced by pH, consistent with a single proton associated with the single electron-transfer event. Ionic strength of the buffer medium also shifted the redox potentials towards positive, suggesting that electrostatic forces contribute to the protein environment required for the electron-transfer process. The effect of substrate on the redox potential for each P450c17 was measured in the presence of pregnenolone, progesterone, 17α-hydroxypregnenolone and 17α-hydroxyprogesterone. However, no influence on the redox parameters was observed.