Effects of ACTH on steroidogenesis in bovine adrenocortical cells in primary culture—increased secretion of 17α-hydroxylated steroids associated with a refractoriness in total steroid output

Abstract

The long-term effects of ACTH on steroidogenesis in bovine adrenocortical cells maintained in primary culture have been investigated. Cells in monolayer culture were incubated in the presence or absence of ACTH for up to 72 h, and the steroid content of the incubation medium was assayed at 12 h intervals. During the first 12 h, adrenocortical cells incubated in the presence of ACTH (10 −9 M and 10 −6 M) produced substantially more cortisol and corticosterone than did cells incubated in the absence of ACTH. The production of steroidogenic intermediates such as pregnenolone, progesterone, and 17α-hydroxypregnenolone, as well as 17α-hydroxyprogesterone, 11-deoxycortisol, and 11-deoxycorticosterone also was increased by short-term (12 h) treatment with ACTH. Thereafter, corticosteroid production by cells incubated in the continued presence of ACTH decreased in a time and concentration dependent fashion. The maximal rate of cortisol production by cells incubated in the presence of ACTH (10 −9 M and 10 −6 M) for 72 h was only one third that of cells incubated in the presence of ACTH for 12 h. More dramatically, by 36 h, corticosterone secretion by cells incubated in the presence of ACTH (10 −6 M) declined to less than 20% of that of nontreated cells, and the production of 11-deoxycorticosterone was no longer detectable. ACTH also induced refractoriness in the production of other C 21-steroids (pregnenolone, progesterone, 17α-hydroxypregnenolone, 17α-hydroxyprogesterone, and 11-deoxycortisol) as well as of C 19-steroids (dehydroepiandrosterone, androstenedione, and 11β-hydroxyandrostenedione). The ACTH-induced refractoriness in the production of C 21-steroids lacking a 17α-hydroxyl group occurred earlier than that of 17-hydroxylated C 21-steroids. Despite the decline in total corticosteroid production, the long term effect of ACTH was to enhance the relative secretion of 17α-hydroxylated steroids and C 19-steroids. Adrenocortical cells incubated for 72 h in the presence of ACTH continued to secrete cortisol, 17α-hydroxyprogesterone, 11-deoxycortisol, and 11β-hydroxyan drostenedione in increased amounts. In fact, 11-deoxycortisol became a major secretory product of the ACTH-refractory adrenocortical cell. These results are indicative that ACTH acts in diverse manners on the bovine adrenocortical cell to affect corticosteroid secretion. The initial stimulation of corticosteroid production appears to be reflective of an increase in overall substrate (cholesterol) utilization and probably is mediated, in part, by an increase in cholesterol side chain cleavage activity. The secretion of 17α-hydroxysteroids and C 19-steroids is enhanced further by an action of ACTH to increase 17α-hydroxylase activity and possibly also 17,20-lyase activity. The ACTH-induced refractoriness in corticosteroid production, on the other hand, appears to result primarily from a decline in precursor (cholesterol) utilization.