Effects Of Different Treatments Of Sperm And Sperm Tail On Bovine ICSI Embryo Development And Transplantation

Embryo Transfer(ET)technology can fully exploit the breeding potential of improved cows,shorten the breeding time of breeding bulls,and improve the quality of cattle.Embryos for transplantation can be divided into in vitro fertilized embryos,in vivo fertilized embryos,and intracellular cytoplasmic sperm injection(ICSI)embryos.Currently,Intracytoplasmic Sperm Injection(ICSI)technology has been maturely and successfully used in different species.Nevertheless,the fertilization rate,cleavage rate and blastocyst rate of bovine oocytes after ICSI are still not high.The success of ICSI might be influenced by many factors including sperm status,sperm treatment,oocyte status,and operation approach.In this study,the sperms were pretreated using different methods.Afterwards,ICSI was performed in bovine oocyte by injecting different morphologies of sperm and sperm tail,embryo quality was verified by differential staining and embryo transfer.The aim of this study is to explore the effects of different sperm morphologies and parts on the development of bovine embryos after ICSI.This study is able to provide the theoretical foundation for improving the development rate of bovine embryo after ICSI and optimizing the ICSI operation,but also provide a reference for the solution of human reproductive problems.The results obtained are as follows:Sperms were pretreated by alkali(NaOH)and ultrasound to remove the sperm tails and then used for ICIS.The results showed that the embryo cleavage rate(41%vs 29.4%)and mulberry sac rate(31.7%vs 14.5%)in NaOH treatment group were significantly higher than those in ultrasound treatment group.ICIS was performed by injecting the braked sperm and sperm head treated by NaOH,respectively.The cleavage rate(51.2%vs 38%)in the braked sperm group was significantly higher than that in NaOH treated group.The blastocyst rate(20.9%vs 27.6%)was not significantly different between two groups.ICIS was performed by injecting the sperm tail and only the solution(shame injection),respectively.The parthenogenetic group was used as control.The cleavage rate in the sperm tail treatment group(12%vs 49.7%vs 55.1%)was significantly lower than that in the shame injection group and the parthenogenetic group.The blastocyst rate(66.7%vs 51.2%vs 55.5%)was significantly higher than that in the shame injection group and the parthenogenetic group.There was no significant difference in cleavage rate(49.7%vs 55.1%)and blastocyst rate(51.2%vs 55.5)between the shame injection and parthenogenetic groupsThe number of blastocysts was counted by differential staining.Compared with ICSI blastocysts obtained by immobilized sperm and sperm head treated with NaOH,the total number of blastocysts(87.16vs89.43)and ICM/T(0.21vs0.22)had no significant difference.ICSI blastocysts obtained by braking sperm and by sperm head treated by NaOH were transplanted respectively.Pregnancy rates were 33.33%and 35.71%respectively,with no significant difference.In summary,the efficiency of bovine ICSI can be improved by treating sperms using the NaOH.High embryo development rate can be achieved by using the braked sperm for ICIS.However,there was no significant difference in the quality of developed blastocysts and the pregnancy rate of transplantation compared with the microinjection of sperm treated with NaOH.There is no positive effect on the development of oocyte by injecting the sperm tail for ICIS.During ICSI operation,mechanical damage does not have significant effect on the development of oocyte.