Background: Studies have shown that chemotherapy and radiotherapy can cause premature ovarian failure and loss of fertility in female cancer patients. Ovarian cortex cryopreservation is a good choice to preserve female fertility before cancer treatment. Following the remission of the disease, the thawed ovarian tissue could be transplanted back and restore fertility of the patient. However, there is a risk to reintroduce cancer cells in the body and leads to the recurrence of cancer. Since current in vitro culture techniques of primordial follicles have a low success rate in obtaining mature oocytes, an artificial ovary with primordial follicles may be good way to solve this problem.
Methods: In the study, we established an artificial ovary model based on the participation of mesenchymal stem cells (MSCs) to evaluate the effect of MSCs on follicular development and oocyte maturation. P2.5 mouse ovaries were digested into single cell suspensions and mixed with bone marrow derived mesenchymal stem cells (BM-MSCs) at a 1:1 ratio and a reconstituted ovarian model was generated by using phytohemagglutin. The phenotype and mechanism studies were explored by follicle counting, immunohistochemistry, immunofluorescence, in vitro maturation (IVM), in vitro fertilization (IVF), real-time quantitative polymerase chain reaction (RT-qPCR), and Terminal-deoxynucleotidyl transferase mediated nick end labeling(TUNEL) assay.
Results: Our results revealed the addition of BM-MSCs in reconstituted ovary could mesenchymal stem cells at the single-cell level reveals that the addition of stem cells increase the survival rate of oocytes and promote the follicular growth and development. The reconstituted ovaries have normal folliculogenesis and oocyte maturation. After aggregation with ovarian cells, BM-MSCs didn’t pr oliferate and participate in the formation of follicles. On the contrary, their numbers decreased significantly along with the development of reconstituted ovary and the cells were found to be surrounded with growing follicles. When theca cells were labeled with CYP17a1, only individual theca cells showed the overlapped staining with GFP labeled BM-MSCs. The results suggest that BM-MSCs may participate in directing the differentiation of theca layer in the reconstituted ovary.
Conclusions: BM-MSCs promoted ovarian cell survival, follicle formation and development in the artificial ovary. Since it didn’t proliferate in the reconstituted ovary, it may represent a new and safe approach on clinical fertility preservation by improving the success rate of thawed ovaries transplantation.