Study On The Mechanism Of Busulfan Poisoning Mice Testis

Busulfan is highly toxic to the spermatogenesis and can cause male sterility.It is an ideal agent for the preparation of transplant recipients of spermatogonial stem cells(SSCs).Orchitis also has an important effect on the spermatogenic function and can even lead to male sterility.However,the mechanism by which busulfan damages the blood-testis barrier(BTB)and affects spermatogenesis is unclear.This is likely due to bu-induced noninfectious testicular inflammation,affecting the secretion of cytokines and impairing fertility.The study was aimed to improve the efficiency and safety of spermatogonial stem cells(SSCs)transplantation recipients so that to decrease male sterility caused by busulfan toxicity,which is helpful in revealing the toxicity mechanism of busulfan on testis.After mice testicles were injected with busulfan,testicular biotin tracing was performed,busulfan levels in epididymal semen and serum inflammatory factor levels were measured,and Immunohistochemistry or immunofluorescence of mouse BTB-associated connexin were detected.The results showed that the highest proportion of busulfan in the epididymal semen reached a summit at 30 min post-treatment then gradually decreased.The earliest observable red fluorescence in the seminiferous tubules traced by biotin was detected at 24 h,suggesting the damage of blood testis barrier(BTB).At 5d,the content of Occludin and connexin43 both began to decrease,and vacuoles appeared in individual seminiferous tubules.As time went by,the content of Occludin and connexin43 became less and less.At21 d,almost all seminiferous tubules were empty,and positive signals basically disappeared.In serum,the concentration of TNF-α,IL-1β and IL-6 progressively increased by busulfan treatment,and at 36 h the amount of the three indexes was significantly higher than that in the control group.The values peaked at 21 d with TNF-α of 1855.51 ± 10.32 pg,IL-1βof 293.59±3.34 pg/m L,and IL-6 of 340.30±12.55 pg/m L,then gradually decreased.(Conculsion)In summary,busulfan could freely cross over BTB and the biotracer could break through BTB within 24 h.With the passage of injection time,the content of tight junction-related protein Occludin and gap junction-related protein connexin43(CX-43)in BTB gradually decreased,indicating that these two connections were destroyed.The serum inflammatory factors TNF-α,IL-1β and IL-6 concentration gradually increased and reached significant levels,reaching the highest level at 21 d,then gradually decreased.It is speculated that Busulfan can freely cross the blood testis barrier,induce inflammation of the testis,damage the testis,and even lead to male sterility.