Stem cell kinetics in rat testis after irreversible injury induced by 2,5-hexanedione

Stem cells provide a continuous supply of committed progenitor cells for the process of spermatogenesis. In rodents, stem cells have been identified as single, undifferentiated type A spermatogonia. The rate of stem cell division has not been definitively determined because of difficulty in locating stem cells among a normal compliment of germ cells. The testicular toxicant 2,5-hexanedione (2,5-HD) induces irreversible testicular atrophy with only Sertoli cells and spermatogonia remaining after injury. Stem cell kinetics could be assessed in this toxicant model because of the absence of most mature germ cells. It is also not known if 2,5-HD-exposed rats possess an active stem spermatogonia population. Charles River CD rats were exposed to 1% 2,5-HD in drinking water for 5 wk. At 7 or 35 wk following toxicant exposure, rats were exposed to bromodeoxycytidine continuously via Alzet mini-pumps for 1-28 days. Serial cross sections of testis were used to identify single stem spermatogonia and to determine whether the cells were positive or negative for bromodeoxyuridine incorporation. We obtained a continuous labeling index for stem cells from rats 7 and 35 wk after 2,5-HD exposure and found that stem cells had a cell cycle time of approximately 8-14 days at both time points after toxicant exposure. In conclusion, we have developed a method for the assessment of stem cell kinetics and verified the presence of an actively dividing stem cell population in irreversibly injured testes.