THE ZONA-RETICULARIS IS THE SITE OF BIOSYNTHESIS OF DEHYDROEPIANDROSTERONE AND DEHYDROEPIANDROSTERONE-SULFATE IN THE ADULT HUMAN ADRENAL-CORTEX RESULTING FROM ITS LOW EXPRESSION OF 3-BETA-HYDROXYSTEROID DEHYDROGENASE

Based on indirect evidence, it has often been assumed that the zona re ticularis of the adult human adrenal cortex is the source of the adren al androgens, dehydroepiandrosterone (DHEA) and DHEA sulfate (DHEAS), but direct tests of this concept have been few. Using the techniques o f cell culture, Northern blotting, and RIA, we compared the properties of separated adult zonal cells to those of fetal zone cells, a cell t ype well known to secrete large amounts of DHEA(S) due to its low expr ession of 3 beta-hydroxysteroid dehydrogenase (3 beta HSD). In nine gl ands from donors of a wide age range, the zona fasciculata and zona re ticularis were separated and dissociated, and the cells were placed in culture. After 5 days, serum was removed by a 24-h period in serum-fr ee defined medium followed by a 24-h exposure to cAMP analogs, with th e optional addition of insulin, also in serum-free medium. The separat ed fasciculata and reticularis cells showed large differences in the D HEA(S)cortisol (F) production ratios from added pregnenolone precursor , consistent with the synthesis of only F and essentially no DHEA(S) b y fasciculata cells and with the synthesis of mostly DHEA(S) with litt le or no F by both reticularis cells and fetal zone cells. The differe nt patterns of steroidogenesis were accompanied by a much lower level of expression of type II 3 beta HSD in reticularis cells, similar to t hat in fetal zone cells. In contrast, other genes were similarly regul ated in the two adult zones and in the fetal zone by both cAMP and ins ulin. The levels of messenger ribonucleic acids for 17 alpha-hydroxyla se, cholesterol side-chain cleavage enzyme, 21-hydroxylase, and 11 bet a-hydroxylase responded to cAMP and insulin in both reticularis cells and fetal zone cells in the same pattern as that previously establishe d in fasciculata cells. The central role of the limited expression of 3 beta HSD in the DHEA(S)-synthesizing property of reticularis cells w as established by inhibition of 3 beta HSD in fasciculata cells with t rilostane, which caused them to increase their DHEA/F production ratio to a level exceeding even that in fetal zone cells. There did not app ear to any age-related changes in gene expression that could account f or the large age-related decline in DHEA(S) biosynthesis in humans in either reticularis or fasciculata cells. Thus, the most likely cause o f the age-related decline in adrenal androgen biosynthesis is an age-r elated decline in the number of functional reticularis cells, without a major change in the differentiated properties of the zonal cells as a function of age.