Investigation of the contributions of adipose-derived mesenchymal stem cells on the in vitro folliculogenesis process

Objective: This proposed study, we aimed to investigate the effects of co-cultures of frozen-thawed ovarian tissue and mesenchymal stem cells isolated from adipose tissue on the efficiency and quality of the in vitro folliculogenesis process. Material and Methods: In this study, AK-MSCs were isolated and characterized from 8-10 weeks old female mice. Ovarian tissues were obtained from 3-week-old female mice, freeze-thawed by vitrification method and agar pads were used. 3D ovary culture was apply for 14 days with or without AK-MSCs. Follicles cultured from the beginning of the culture were photographed under an inverted microscope and their follicle diameters were measured. At the end of the experiment, ovarian tissue pieces belonging to the control and study groups were evaluated histopathologically and mRNA expression levels of Gdf9, Bmp15, Bcl-xl, Bax and Bcl-2 were analyzed by QPCR method. The data obtained as a result of the experiments were evaluated using GraphPad Prism 8.0.2 with one-way analysis of variance (ANOVA). Results: At the end of the experiment, the number of viable follicles decreased in the vitrified group, but when evaluated histologically, the primordial and primary follicle structure was protect. In the virtification+ msc(+) group, in vitro folliculogenesis process was provide and secondary, antral follicle formation was microscopically observed. Follicle diameter increased statistically significantly (P<00.5) until day 6 of the culture process. Secondary and antral follicles were not observed in the vitrification+ msc(-) group. Follicle diameter decreased significantly at day 10 (P<00.1). Expression of vitrification+ msc(+) Gdf9, Bmp15 is higher than vitrification and vitrification+ msc(-). Considering apoptosis markers, it inhibited granulosa cell and stromal cell apoptosis thanks to cytokines secreted from stem cells. The AMH level of the control, virtification, vitrification+ msc(-) groups is less than that of the vitrification+ msc(+) group. Conclusion: In this study, co-culture of AK-MSCs of follicles obtained from frozen-thawed ovarian tissue, which has a low survival rate when cultured, can treat the effects on follicles during the ovarian cryopreservation process and increase survival rates.