BackgroundOvarian tissue cryopreservation (OTC) is the only method of fertility preservation (FP) in prepubertal girls, but the experience remains limited. This study investigates the effectiveness and feasibility of FP of OTC in children facing gonadotoxicity treatment in Chinese first ovarian tissue cryobank.ProcedureOTC and evaluation of 49 children ≤14 years old in the cryobank of Beijing Obstetrics and Gynecology Hospital, Capital Medical University, from July 2017 to May 19, 2022, were analyzed retrospectively. We compared children’s general characteristics, follicle numbers, and hormone levels with and without chemotherapy before OTC.ResultsThe age of 49 children at the time of OTC was 7.55 (1–14) years old. There were 23 cases of hematological non-malignant diseases, eight cases of hematological malignant diseases, four cases of gynecological malignant tumors, one case of neurological malignant tumors, one case of bladder cancer, five cases of sarcoma, three cases of mucopolysaccharidosis, one case of metachromatic leukodystrophy, two cases of dermatomyositis, one case of Turner’s syndrome. The median follicular count per 2-mm biopsy was 705. Age and AMH were not correlated (r = 0.084, P = 0.585). Age and follicle count per 2-mm biopsy was not correlated (r = −0.128, P = 0.403). Log10 (follicle count per 2-mm biopsy) and Log10 (AMH) were not correlated (r = −0.118, P = 0.456). Chemotherapy before OTC decreased AMH levels but had no significant effect on the number of follicles per 2-mm biopsy.ConclusionsOTC is the only method to preserve the fertility of prepubertal girls, and it is safe and effective. Chemotherapy before OTC is not a contraindication to OTC.

Background The massive loss of follicles in the early stage of ovarian tissue transplantation is considered a significant restriction to the efficacy of ovarian tissue cryopreservation (OTC) and transplantation (OT). The use of mesenchymal stem cells (MSCs) before transplantation of ovarian fragments shortened the hypoxic period and boosted neovascularization. Hypoxia-preconditioned MSCs can enhance the potential of angiogenesis. Can hypoxia-preconditioned human umbilical cord mesenchymal stem cell (HucMSCs) and ovarian tissue co-xenotransplantation improve more neovascularization and subsequently more follicle survival in human ovarian tissue? Methods Frozen-thawed cortical pieces from 4 patients were transplanted into the bilateral renal capsule of immune-deficient nude mice without HucMSCs or normoxia/hypoxia-preconditioned HucMSCs. Sixty-four mice were randomly distributed into 4 groups. In each group, the mice were euthanized for blood and/or graft retrieval on post-transplantation days 3 (n = 8) and 7 (n = 8), respectively. Non-grafted frozen-thawed ovarian fragment was taken for non-grafted control. Grafts were histologically processed and analysed for follicle density and atretic follicles by HE, neovascularization by CD34 and CD31 immunohistochemical staining, primordial follicle growth by Ki67 staining, and apoptosis of stromal cell and follicles by immunofluorescence using TUNEL. The ROS and TAC levels of grafted and non-grafted tissue were assessed. We evaluated the protein expression of HIF1α, VEGFA, pAkt, Akt, and GDF9 in grafted and non-grafted ovarian tissue. E2, Prog, AMH, and FSH levels in the plasma of mice were measured after 3 and 7 days of OT. Results Hypoxia-preconditioned HucMSCs positively protect the grafted ovarian tissue by significantly decreasing the apoptosis and increasing higher expression of CD31, CD34, and VEGFA for earlier angiogenesis. They are crucial to preserving the resting primordial follicle pool by modulation of follicle death. Conclusion This is the first study to demonstrate that co-transplantation of hypoxia-preconditioned HucMSC with ovarian tissue improved earlier vascularization of ovarian grafts in the early post-grafting period, which correlates with increased follicle survival and reduced apoptosis. The HIF1α/VEGFA signal pathways may play an important role in elucidating the mechanisms of action of hypoxia-preconditioned HucMSCs with regard to OT and clinical implementation.