Spermatogonial stem cell survival in ram lambs following busulfan treatment

Abstract

To clarify the effect of busulfan on the depletion of spermatogonial stem cells (SSCs) from shal rams testis, in the first experiment, lambs were treated by intraperitoneal injection with 4 mg/kg busulfan. In the second experiment, different concentrations of busulfan (1, 2 and 4 mg/kg) were injected directly into both sides of the left testis. The testes of 8 lambs were collected by standard castration procedure for histological analysis five weeks after the treatments and the left testis of remaining lambs were collected after eight weeks and a two-time enzymatic digestion process was used to isolate SSCs. The results showed that all rams that had received intraperitoneal injections of busulfan died. But by testicular injecting of same dose of the drug, 40% of the animals died. The testicular injection of rams with 1, 2 and 4 mg/kg of busulfan resulted in a dose dependent decrease in testis size and also spermatocytes population after 5 weeks of treatments. From the results of colony formation 8 weeks after treatment with busulfan, it can be concluded that only in 1 and 2 mg/kg of busulfan, recovery of endogenous germ cells was performed. In conclusion, the results demonstrated that intra-testicular injections of busulfan (2 mg/kg) reduced spermatocytes population in ram testis within 5 weeks of treatments, and this effect was reversible within 8 weeks of injection. However, it was not recommended to inject 4 mg/kg busulfan into the peritoneal cavity or testis of lambs based on its side effects.

Successful in vivo Transplantation of Cultured and Enriched Testicular Germ Cells of Pre-Pubertal Bucks to Busulfan-Treated Homologous Recipients

“…In this study, we demonstrate that the transplantation of cultured and enriched cmGCs cells from donor male goat results in the recolonization of recipient testes previously sterilized by busulfan treatments. However, busulfan treatment can produce systemic toxicity and even lethality resulting from severe bone marrow depression as reported in pigs [Honaramooz et al, 2005], sheep [Rasouli et al, 2020], mice [Kim et al, 1997;Azizi et al, 2021], and rats [Ogawa et al, 1999]. Therefore, to avoid the side effects associated with busulfan treatment, we first optimized the dose and route of busulfan injection for safe preparation of the recipient and then transplanted the enriched population of CD90 + cmGCs with USG guidance.…”

Intratesticular versus intraperitoneal injection of Busulfan for the induction of azoospermia in a rat model

“…No systemic effects were found in rats that had undergone IT Busulfan injection. The least toxic effects of Busulfan in the IT group could be associated with the low circulation of Busulfan in the bloodstream while in the IP injection Busulfan can bio-distribute faster to the remote sites especially kidneys and liver [ 33 , 36 – 38 ]. The most probable cause of mortality in animal modeling following Busulfan admiration is associated with the suppression of leukopoiesis and hematopoiesis.…”