Metabolism of Testosterone by Principal Cells and Basal Cells Isolated from the Rat Epididymal Epithelium1

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Abstract
The ability of epididymal tissue to convert testosterone to dihydrotestosterone and the functional dependence of the epididymal epithelium on dihydrotestosterone are well established. However, the distribution of Δ4-3-ketosteroid-5α-oxidoreductase (5α-reductase) among the several cell types found in the epididymal epithelium was unknown. Therefore, we determined if principal cells or basal cells from the epithelium of the caput epididymidis metabolize testosterone to dihydrotestosterone. Epididymal tissue was enzymatically dissociated and principal cell and basal cell fractions were isolated by elutriation. About 1.2 × 107 cells were incubated for 4 h at 34°C in 3 ml of culture medium containing 103 nM [3H]-testosterone. After incubation, [14C]-dihydrotestosterone and organic solvent were added and the samples were stored for 2–6 weeks. Steroids were extracted and chromatographed on Sephadex LH-20; peak fractions were analyzed by thin layer chromatography and, in selected cases, the isolated steroids were recrystallized. In six incubations of principal cells, all of the [3H]-testosterone was metabolized to 5α-reduced compounds, whereas in each incubation of basal cells or of cell-free medium virtually all of the 3H was recovered as [3H]-testosterone. Per 106 cells in the principal-cell fraction, 0.2 ± 0.02 pmoles of testosterone remained after 4 h and 17.3 ± 0.8 pmoles of 5α-reduced steroids (63 ± 4% dihydrotestoster-one) had been produced. For incubations of basal cell fractions, 15.1 ± 3 pmoles of testosterone and 1.7 ± 0.2 pmoles of 5α-reduced steroids per 106 cells were present after 4 h. The slight conversion of testosterone to 5α-reduced steroids during incubations of basal cell fractions was attributed to principal cells (7% of all cells) contaminating these fractions rather than to enzymatic activity of basal cells. It was concluded that the 5α-reductase activity of the epithelium in the caput epididymidis is restricted to principal cells.