In-vitro testosterone metabolism in the mouse preputial gland: intercellular co-operation and changes with cell maturation

Abstract

ABSTRACT In-vitro [14C]testosterone metabolism was investigated in isolated cells of adult male mouse preputial sebaceous glands. Labelled steroids were extracted and chromatographed after a 2-h incubation, and were identified as 5α-dihydrotestosterone, androstenedione, 5α-androstane-3,17-dione, 5α-androstane-3α,17β-diol and 5α-androstane-3β,17β-diol, androsterone and 3-epiandrosterone. In cells separated according to state of maturity (lipid content) by isopycnic centrifugation in a metrizamide gradient, maximal testosterone metabolism occurred in large, nearly mature cells. In this population, mean hydroxysteroid 5α-reductase and 17β-hydroxysteroid dehydrogenase activities were 3·8 and 2·3 nmol/106 cells per 2 h respectively, more than 100-fold greater than in the densest population, comprised of undifferentiated and early differentiating cells. It was also found that the profile of testosterone metabolites was dependent on the proportion of the label metabolized. The metabolite index (MI), i.e. the average number of enzymatic steps undergone per molecule of metabolite, increased with increasing substrate utilization. Metrizamide showed reversible, non-specific inhibition of testosterone metabolism and reduction of the MI. Thus, it was postulated that testosterone is metabolized sequentially by different cells, with metrizamide inhibiting cellular uptake and intercellular substrate transport. This suggested that most of the metabolites would be found in the medium, rather than in the cellular compartment. Further, in incubations run without cell disaggregation, efficient substrate cycling among cells should result in a high MI, independent of metrizamide concentration and substrate utilization. These predictions were all confirmed, providing strong evidence that testosterone metabolism is a cooperative effort among several cells in this tissue. J. Endocr. (1984) 100, 377–388