Fertilization And Embryo Development Of The Female And Male Gametes From Ovarian Graft Heterosexual Recipients In Mice

Ovarian heterosexual transplantation has special value for full utilization of germ cells as a resource in female animals. In order to evaluate the competence of fertilization and embryo development on the female and male gametes developed in the male recipients, ovarian heterosexual transplantation were used to produce the mouse model with both female and male gonads for examining sexual glands of male recipients, sperms and related gene expression. For the first time in mammals, the competence of the female and male gametes produced in the male graft recipients was assessed by performing in vitro fertilization and embryo transfer in this study. The main results were following:1. Ovarian grafts developed well with large antral follicles on day 21 or 28 after transplantation. Morphology of testis and epididymis from the ovary graft recipients was normal, without significant difference compared with that control male mice. The body weight, wet weight of testis and epididymis, and the weight ratio to the body weight of male mice in each group had no statistics difference (P>0.05). It was shown that The seminiferous tubule in testis, epididymis from the ovary graft recipient were normal structure and the lumen of the tubes were filled with sperm by histological section observation, appeared no difference in morphology compared with that control males. The present study demonstrates that ovarian allograft do not appear to greatly affect testis, epididymis structure from the ovary graft recipient2. The sperm abnormal rate in 21 d and 28 d ovarian translantation group was 7.62% and 7.78%, respectively ,without significant difference compared with that control group(6.73%) (P>0.05). The growen ovarian grafts and developed follicles do not appear to significantly affect spermatogenesis in testis and sperm maturation in epididymis of male recipients.3. Reproductive capacity of the ovary graft recipients with developed follicles in ovarian grafts was investigated by breed with normal female mice during 21-60 d after ovarian tansplantation. The litter size in the experimental group was 8.83±2.10, without significant difference compared with that control group(9.85±2.34) (P>0.05). It demonstrates that the ovary graft recipients are fertile, the functional ovarian grafts do not appear to significantly affect spermatogenesis, sperm maturation, and fertilizing ability in vivo of the male recipients.4. Expression of Dazl, Pgk2, Prm2 and Bax mRNA in testis and epididymis were determined by using reverse transcriptase-polymerase chain reaction. The expression of 4 gene mRNAs in 2 experimental groups showed similar trend with the the control group. It indicates that the ovarian transplantation do not appear to significantly affect spermatogenesis in the ovary graft recipients at the molecular level.5. Oocytes and sperms collected from the same ovary graft recipient can fertilize in vitro. The two-cell cleavage rate and blastocyst rate in the experimental group was 60.4% and 33.9% , respectively. This is the first report that both sexual gametes developed in same physical environment in mice with male and female sex glands by heterosexual ovarian transplantation present potential of fertilization in vitro and development to blastocyst. This work has shown that the physical environment of male mice can support the development of oocytes in ovarian allografts and sexual reproduction in animal preserves potential of male and female gametes developing in same body.6. The developmental competence of oocytes and sperms collected from the same ovary graft recipient was assessed by performing in vitro fertilisation and embryo transfer to recipients. Six pups developed normally were born by three surrogate mothers. The pregnancy rate and total birth rate was 25.0% and 4.2%, respectively. This is the first report of live offspring obtained from both sexual gametes developed in same body after heterosexual ovarian transplantation using IVF. The growth of the pups in the experimental group and control group was evaluated by measuring the body weight on day 1, 7, 10, 14 and 20 after birth. The growth of the pups showed similar trend between the experimental group and control group. In conclusion, this work has shown that male and female gametes from recipient mice with male and female sex glands established by heterosexual ovarian transplantation present potential of development and fertilization and that these gametes can produce live offspring. The offspring can develop normally.7. The reproductive competence of offspring produced by in vitro fertilization with both sexual gametes from ovarian grafts and testes of male recipients was examined. An average litter size up to 9.60±3.98 in the normal female mice mated with 2 male offspring was observed in test group and showed no significant difference with control group. The results demonstrate that the offspring produced by both sexual gametes from ovarian grafts and testes of male recipients have normal reproductive capacity, without impact on it.