Prepubertal PTU treatment in rat increases Sertoli cell number and sperm production.

The number of Sertoli cells (SCs) ultimately determines the upper limit of sperm production in the testis. Previous studies have shown that thyroid hormones (TH) receptors are abundantly expressed in developing SCs; therefore, it was highly significant to discover that transient neonatal hypothyroidism induced by the goitrogen 6-n-propyl-2-thiouracil (PTU) can extend SCs proliferation beyond the first 2 weeks postnatal and increase testis weight and sperm production. Further studies concluded that treatment must begin before day 8 post birth in rats. Recent studies, however, showed that SCs present in the transition region at the rete testis exhibit a more immature phenotype and have prolonged mitotic activity, which led to the hypothesis that SCs in this region will retain the capacity to respond to PTU treatment over a longer period of time. In the present study, male Wistar rats were treated with PTU from days 21 to 40 and were evaluated at 40 and 160 days of age. Similar to neonatal rat SCs, it was demonstrated that prepubertal SCs in the transition region have a high mitotic activity and are highly sensitive to TH levels. This delayed, transient hypothyroidism resulted in significantly increased testis weight, SCs number and daily sperm production. The results demonstrate for the first time that Sertoli cells showing plasticity in the transition region can be stimulated to increase proliferation and contribute to a late stage surge in testis weight and sperm output.

Horse spermatogonial stem cell cryopreservation: feasible protocols and potential biotechnological applications.

The establishment of proper conditions for spermatogonial stem cells (SSCs) cryopreservation and storage represents an important biotechnological approach for the preservation of the genetic stock of valuable animals. This study demonstrates the effects of different cryopreservation protocols on the survival rates and phenotypic expression of SSCs in horses. The cells were enzymatically isolated from testes of eight adult horses. After enrichment and characterization of germ cells in the suspension, the feasibility of several cryopreservation protocols were evaluated. Three different cryomedia compositions, associated with three different methods of freezing (vitrification, slow-freezing and fast-freezing) were evaluated. Based on the rates of viable SSCs found before and after thawing, as well as the number of recovered cells after cryopreservation, the best results were obtained utilizing the DMSO-based cryomedia associated with the slow-freezing method. In addition, when isolated cells were cultured in vitro, MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay and immunofluorescence analysis indicated that the cryopreserved cells were as metabolically active as the fresh cells and were also expressing typical SSCs proteins (VASA, NANOS2 and GFRA1). Therefore, our results indicate that equine SSCs can be cryopreserved without impairment of structure, function, or colony-forming abilities.