Steroid 5 alpha-reductase activity in endothelial cells from human umbilical cord vessels.

The metabolism of radiolabeled progesterone and androstenedione was evaluated in endothelial cells from human umbilical cord vein and arteries maintained in culture. The predominant metabolite of progesterone was 5 alpha-pregnane-3,20-dione and that of androstenedione was 5 alpha-androstane-3,17-dione. Thus, the major pathway of progesterone and androstenedione metabolism within these cells is via steroid 5 alpha-reductase. The rate of formation of 5 alpha-pregnane-3,20-dione from progesterone by venous endothelial cells was linear with incubation time up to 4 h and with cell number up to 1.6 X 10(6) cells/ml. The apparent Km of 5 alpha-reductase for progesterone was 0.4 microM; and, the Vmax was 55 pmol 5 alpha-pregnane-3,20-dione formed/mg protein X h. The rate of 5 alpha-androstane-3,17-dione formation from androstenedione also was linear with incubation time up to 4 h. In addition to 5 alpha-androstane-3,17-dione, the metabolism of androstenedione by either venous or arterial cells resulted in the formation of various minor metabolites, including testosterone and 5 alpha-reduced steroids, viz. 5 alpha-dihydrotestosterone, androsterone, isoandrosterone, 5 alpha-androstane-3 alpha, 17 beta-diol, and 5 alpha-androstane-3 beta, 17 beta-diol. Estrogens (i.e. estradiol-17 beta and estrone) were not detected as products of androstenedione metabolism. The formation of these metabolites are indicative that the steroid-metabolizing enzymes present in endothelial cells are: 5 alpha-reductase, 17 beta-hydroxysteroid oxidoreductase, 3 alpha-hydroxysteroid oxidoreductase, and 3 beta-hydroxysteroid oxidoreductase.

Androstenedione metabolism in human lung fibroblasts.

Human lung fibroblasts in culture metabolized [3H]androstenedione to a number of different compounds, including testosterone, 5 alpha-androstanedione, androsterone, 5 alpha-dihydrotestosterone, isoandrosterone, and 5 alpha-androstane-3 alpha,-17 beta-diol. The major products were 5 alpha-androstanedione and testosterone. Estrone, estradiol-17 beta and 5 beta-reduced steroids were not formed. The production rates of testosterone and 5 alpha-androstanedione from [3H]androstenedione by lung fibroblasts were studied both as a function of incubation time and substrate concentration. The rates of formation of testosterone and 5 alpha-androstanedione remained linear with time up to 4 h. The apparent Km of human lung fibroblast 5 alpha-reductase was 1 microM, and that of 17 beta-hydroxysteroid oxidoreductase was 11 microM. The findings of this study suggest that mesenchyma may contribute to the metabolism of androstenedione in human lung tissue.

Epidermal keratinocytes: a source of 5 alpha-dihydrotestosterone production in human skin.

The major products of testosterone, androstenedione, and progesterone metabolism by human epidermal keratinocytes are 5 alpha-reduced steroids, viz. 5 alpha-dihydrotestosterone, 5 alpha-androstanedione, and 5 alpha-dihydroprogesterone, respectively. The rates of metabolite formation by these cells were linear with incubation time up to 3 h. The apparent Km of keratinocyte 5 alpha-reductase was 1.3 microM for androstenedione and 1.5 microM for progesterone. 5 alpha-Reductase activity was found only in particulate subcellular fractions of a homogenate of epidermal keratinocytes when assayed with tritium-labeled progesterone as the substrate and NADPH as the cofactor. In addition to 5 alpha-reductase activity, other enzymatic activities found in epidermal keratinocytes were 17 beta-hydroxysteroid oxidoreductase and 3 beta-hydroxysteroid oxidoreductase. These enzymes were expressed in the formation of androstenedione from testosterone, testosterone from androstenedione, isoandrosterone from androstenedione, and 3 beta-hydroxy-5 alpha-pregnan-20-one from progesterone. The apparent Km of 17 beta-hydroxysteroid oxidoreductase for androstenedione in epidermal keratinocytes was 10 microM. When measured at weekly intervals, the rates of product formation from testosterone, androstenedione, or progesterone by cultured epidermal keratinocytes increased several-fold with advancing time in culture up to 3 weeks. The results of these studies suggest that epidermal keratinocytes are a major site of synthesis of biologically potent androgens in human skin, viz. testosterone from androstenedione and 5 alpha-dihydrotestosterone from testosterone. Skin is a target organ for 5 alpha-dihydrotestosterone action, and thus, the local formation of 5 alpha-dihydrotestosterone may play an important role in the regulation of proliferation and differentiation of keratinocytes.

Androstenedione metabolism in human alveolar macrophages.

The metabolism of [3H]androstenedione by human alveolar macrophages was investigated. Alveolar macrophages were obtained by bronchopulmonary lavage by use of a heparinized saline solution devoid of Ca++ and Mg++. After purification, the macrophages were incubated at 37 C in RPMI-1640 medium that contained glucose and [1,2,6,7-3H]androstenedione under various experimental conditions. Control incubations were conducted without macrophages. After incubation, 14C-labeled steroids that corresponded to the metabolites were added as internal recovery standards. The metabolites were characterized by chromatography and crystallization to constant 3H to 14C ratios. Human alveolar macrophages convert [3H]androstenedione to 5 alpha-androstane-3,17-dione, testosterone, 5 alpha-dihydrotestosterone, androsterone, and isoandrosterone. Unidentified polar metabolites also were formed. Therefore, the following enzymes are present in these cells: 5 alpha-reductase, 17 beta-hydroxysteroid dehydrogenase, 3 alpha-hydroxysteroid dehydrogenase, 3 beta-hydroxysteroid dehydrogenase, and unknown hydroxylase(s). The rates of formation of the principal metabolites, 5 alpha-androstanedione and testosterone, remained linear up to 4 h of incubation and with macrophage number up to 1.5 X 10(7) cells/ml. These findings suggest that alveolar macrophages may be involved in the peripheral metabolism of androstenedione to potent androgens in man. It is possible that androgens, formed from blood-borne androstenedione within alveolar macrophages, may modulate phagocytic and other activities in these cells.