Isolation of rat Leydig cells and precursor forms after administration of ethane dimethane sulfonate

The cytotoxic drug ethane dimethane sulfonate (EDS) has been extensively used as a means of studying the regeneration of Leydig cells in the adult rat testis. This study used the EDS-treated rat testis as a source of material for the isolation of regenerating Leydig cells and their precursors and describes the procedures required for the isolation of these cell preparations. As early as 13-15 days after EDS, cells in the precursor fraction can bind low, but detectable, levels of iodinated purified human chorionic gonadotropin. However, no luteinizing hormone (LH) response was detected in terms of steroid production. The precursor fraction of cells isolated from the EDS-treated rat testis 17-19 days after the administration of EDS was heterogeneous in light-microscopic appearance, but identifiable Leydig-like cells were present. The cells in this fraction were the first to exhibit the ability to respond to LH with the production of detectable levels of the reduced androgen, 5 alpha-androstane-3 alpha,17 beta-diol. The amount of androgen produced by both the Leydig cell and precursor fractions had increased by 21 days after EDS and reached the levels produced by immature adultlike Leydig cells, which can be isolated from the 20-day-old rat testes. These studies demonstrate that steroidogenically responsive precursor forms of Leydig cells can be isolated from the EDS-treated testes 17-19 days after depletion of the adult Leydig cell population.