TPA inhibits the synthesis of androgens and cortisol and enhances the synthesis non-17 alpha-hydroxylated steroids in cultured human adrenocortical cells

Previously, we showed that 12-O-tetradecanoyl phorbol 13-acetate (TPA), an activator of protein kinase C, is a mitogen for fetal human definitive zone adrenocortical cells in culture. In the present experiments, TPA inhibited forskolin-stimulated cortisol, dehydroepiandrosterone (DHEA), and dehydroepiandrosterone sulfate (DHEAS) synthesis, and enhanced forskolin-stimulated progesterone and corticosterone synthesis. These changes in the pattern of steroidogenesis were shown to result from changes in enzyme activities after forskolin treatment. TPA increased forskolin-stimulated 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) 2-fold, while depressing forskolin-stimulated 17 c-hydroxylase to basal values DHEA sulfotransferase increased 3-fold on transfer of human adrenocortical cells from serum-containing to defined, serum-free medium; TPA inhibited this increase. Experiments in which TPA was added at various times during the time course of forskolin stimulation of 17 alpha-hydroxylase showed that TPA prevents the increase in the level of 17 alpha-hydroxylase, and does not have a direct inhibitory effect on 17 alpha-hydroxylase activity. TPA also inhibited stimulation of 17 alpha-hydroxylase by cAMP analogs, indicating that the inhibition of 17 alpha-hydroxylase by TPA is not due to an effect on adenylate cyclase. Previous experiments have shown that stimulation of intracellular cAMP is sufficient for androgen synthesis by the human adrenocortical cell, under defined, serum-free conditions, and that its high rate of androgen synthesis likely results from the relative levels of 3 beta-HSD, 17 alpha-hydroxylase, and DHEA sulfotransferase in the cell. Enzyme induction by cAMP results in increased production of both androgens and glucocorticoids, whereas activation of protein kinase C changes the balance of enzymes, resulting in increased non-17 alpha-hydroxylated steroid synthesis and decreased androgen and cortisol biosynthesis.