Effect Of Piezo Assisted Hatching On Mouse Embryo Implantation And Development

The zona pellucida(ZP),a glycoprotein layer surrounding the embryo,secretes from oocytes and granulosa cells in the early stage of primary folli-cles.It plays important roles in fertilization and early embryo development.Blastocyst hatching will be inhibited if the zona pellucida became thick or harden.One of the approach for improving blastocyst hatching is the method of assisted hatching(AH).This approach will benefit embryos implantation and pups' delivery.AH involves artificial disruption of the zona pellucida and help the embryo escape from the zona pellucida.Till now,kinds of AH technology have been used in clinical application.In our early studies,we found the hatching ability was varied if different stage of embryo or differ-ent AH technology were used.We also found that methods of zona pellucida drilling with a small hole on morula embryos will obtain the highest rate of blastocyst hatching.However,it is still not known if this method can im-prove the implantation rate and the delivered pups rate.In this study,by em-ploying methods of drilling small hole on morula stage embryo followed by embryo transfer(ET),effect of AH mediated by piezo was evaluated in mice.The final implantation,pups' delivery,pregnancy as well as pups' body weight were monitored.We intend to figure out if this kind of AH approach is suitable for embryo implantation and development.Three groups were set up in our study,including:(1)mating group as control which without AH or ET operation(termed as mating group);(2)ET group which was subjected to ET with in vivo fertilized embryos for observ-ing the effect of ET operation on pregnancy and reproduction(term as ET group);(3)AH+ET group which was subjected to both AH of drilling small hole and ET operation for observing the effect of AH and ET on pregnancy and reproduction(termed as SH group).The results showed that when compared with the mating group,the ET operation could reduce the numbers of embryos implantation,delay the de-liver duration,reduce the litter size of pups and affect the body weight.However,the pregnancy rate,the delivered lived pups rate and the gender ratio were not affected by ET.After ET,the grown pups were mated again.And the deliver duration and the fertility were not affected.SH group significantly improve the blastocyst hatching rate in vitro when compared with the ET group.However,the fetus generated from SH group exhibited delayed development and decreased body weight.AH treatment did not affect the following characters,including the livability of recipient mother,the number of embryo implantation,the implantation rate,the pregnancy rate,deliver duration,the litter size of pups,the delivered pups rate,the delivered live pups rate,the gender ratio,the deliver duration of pups,and the reproductive ability of the SH pups.When compared with the mating group,SH group reduce the implanta-tion rate,delay the deliver duration,reduce the litter size of pups and affect the body weight.However,AH did not affect the pregnancy rate,the deliv-ered live pups rate and the gender after ET.The deliver duration and the fer-tility were not affected in the SH pups.In summary,AH of drilling small hole in zona pellucida by piezo did not improve the implantation rate,the pregnancy rate and the litter size.Conversely,operation of AH and ET could disorder embryo development and the pups body weight.However,there is no affection on the gender and the reproductive ability of the pups.Therefore,we proposed that it should be used cautiously in clinic application for the AH with the method of drilling small hole on zona pellucida.