Intraovarian Transplantation Of Female Germline Stem Cells Rescue Ovarian Function In Chemotherapy-Injured Ovaries

Early menopause and infertility often occur in female cancer patients after chemotherapy(CTx). For these patients, oocyte/embryo cryopreservation or ovarian tissue cryopreservation is the current modality for fertility preservation before chemotherapy. However, the above methods are limited in the long-term protection of ovarian function, especially for fertility preservation. For prepubertal females, oocyte or embryo cryopreservation is impossible because of the immature ovary; For adult unmarried females, embryo cryopreservation needs sperm from donor. Furthermore, both oocyte and embryo cryopreservation require hormone stimulation which will delay the treatment of cancer; Most importantly, hormone stimulation may deteriorate the development of estrogen sensitive cancer because of the elevated level of estrogen. At present, ovarian tissue cryopreservation is still at the experimental stage, and the reimplantation of ovarian tissue may induce the recurrence of cancer. Therefore, considering millions of women have achieved long-term survival after thorough CTx treatment, it is important to rescue their ovarian function and fertility with economic, durable, safe and reliable methods immediately. In 2009, Zou et al. successfully isolated the female germline stem cells(FGSCs) from both neonatal mouse and adult mouse, and confirmed that FGSCs can form eggs in vivo after transplantation, which, for the first time, affirmed the existence of FGSCs in mammal ovary. In 2012, white et al. verified the results of Zou et al. and further successfully isolated and confirmed the existence of human female germline stem cells(hFGSCs). The above experimental results provide fundamental basis for the therapy of infertility and ovarian aging by application of FGSCs. Since so many women, especially young females, who had underwent chemotherapy and suffered from ovarian damage and infertility, we sought to determine whether FGSCs exist in those women and whether stem-cell transplantation could be considered as a therapeutic option to restore ovarian function and infertility caused by CTx using a mouse model of CTx-induced ovarian failure. We successfully isolated FGSCs from mouse of ovarian failure, and identified the characteristic of FGSCs by RT-PCR, immunofluorescence, AP staining, and karyotyping. By transfecting FGSCs by GFP lentiviral and transplanting GFP-labeled FGSCs into the ovaries of infertility mice, after mated with wild-type male mice, we obtained transgenic mice with GFP gene. By detection of GFP gene in the genome DNA from the tail of offspring with the methods of PCR and southern blot, we confirmed that the transgenic mice were derived from GFP-labeled FGSCs. This study first verified the existence of FGSCs in mouse of CTx-induced ovarian failure, and confirmed that the transplantation of FGSCs could restore ovarian function and fertility. Finally, this study provide some theoretical basis and reference value for female infertility patients who want to protect ovarian function and restore the ability of fertility by their own FGSCs, and thus bring new hope of therapeutic method in the future.