Characterization of human Sertoli cells in vitro

The Sertoli cell is thought to play a significant role in the hormonal regulation of spermatogenesis within the rat testis. Little, however, is known about Sertoli cell function in man, largely because of the difficulties associated with the isolation of pure cell populations from human tissue. We have now developed a rapid and reproducible technique for establishing a human Sertoli cell monolayer culture. This has involved mechanical separation of the tissue, sequential trypsin and collagenase enzyme digestion, and final disruption of tubules by passage through a wire mesh grid. Using this technique, primary cultures can be maintained for up to 45 days. Ultrastructural studies of these cells have demonstrated the presence of the perinucleolar spheres, cell to cell junctional complexes, abundant lipid droplets, and smooth endoplasmic reticulum, all characteristic of Sertoli cells. Furthermore, biochemical markers of animal Sertoli cells, androgen-binding protein and gamma-glutamyl transpeptidase, have also been identified in these human cells. Concentrated media electrophoresed on nondenaturing gels containing 2 nM [3H]dihydrotestosterone produced a single peak of bound activity which coelectrophoresed with rat androgen-binding protein. This binding activity persisted despite media changes, thus ruling out contamination by serum binding proteins; fresh media lacked demonstrable binding activity. Using a colorimetric assay, these cells were also found to contain significant gamma-glutamyl transpeptidase activity compared to human foreskin fibroblasts and Leydig cells. Enzyme activity increased in a characteristic dose-response fashion in the presence of FSH (0.05-0.5 microgram/ml) and dibutyryl cAMP (0.1-1 microgram/ml), but not with LH or testosterone. These data offer the first demonstration of human Sertoli cells in monolayer culture and their production of a marker specifically regulated by FSH.