P-108: Stepwise Pattern of Reactive Oxygen Species during In Vitro Development of Cloned Goat Embryos

Background: Although testicular tissue cryopreservation could be advantageous for fertility preservation in cancerous children, the possibility of apoptosis incidence should not be ignored. Our aim was evaluation of the apoptosis incidence in mouse testicular tissue after vitrification and short-term culture. Materials and Methods: Testes of 7days old NMRI male mice were isolated, divided and distributed into control and vitrification groups. Vitrification was performed in 3 steps with combination of EG and DMSO. Fresh and vitrified-warmed testis pieces were cultured in RPMI and 10% KSR for 20 hours. Just after vitrification (0h) and after 3 and 20 hours of culture real-time PCR, Flow cytometry and light microscopy were performed to study apoptosis gene expression, apoptosis incidence and morphology. Results: Totally, tissue degeneration was increased during culture period. Late apoptosis was higher in vitrification group in 0 and 3 h of culture compared to the control, this rate became similar at the end of the culture period. During culture period, BAX expression was significantly increased, and BCL-2 expression was significantly decreased in vitrification group compared to the control one. Also, P53 expression was lower (p<0.05) in vitrification group at 0 hours of culture compared to the control and it became the same as the control at the end of the culture period. Through whole culture period BAX and P53 had lower expression in 3 hours of culture compared to 0 and 20 hours of culture in both vitrification and control groups. 3 hours post culture, BCL-2 expression was higher compared to 0 and 20 hours of culture in vitrification and control groups. Conclusion: It seems that intrinsic pathway and P53 gene are involved in testicular tissue apoptosis followed by culture and also vitrification.