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Female germline stem cells (FGSCs) are renewable sources of oocytes that play an indispensable role in re-establishing mammal fertility. Here, we have established FGSCs from neonatal mice, which exhibit characteristics of germline stem cells. We show that compared with monomeric trigonelline and diosgenin, macromolecular compounds Cistanche deserticola polysaccharides (CDPs) in Chinese herbal medicine can enhance the ability of FGSCs to differentiate into oocytes at appropriate concentrations while maintaining self-renewal in vitro . In contrast, trigonelline and diosgenin inhibited the expression of germ cell-specific genes while reducing cell proliferation activity. In summary, CDPs could induce the differentiation and self-renewal of FGSCs in vitro . The comparison of the effects of the active components of different types of Chinese medicine will provide a reference for the development of clinical drugs in the future, and help to elucidate the development process of FGSCs.

Background: In vitro culture of preantral follicles is a promising technology for fertility preservation., Objectives: This study aims to investigate an optimized three-dimensional (3D) fetal bovine serum (FBS)-free preantral follicle culture system having a simple and easy operation., Methods: The isolated follicles from mouse ovaries were randomly divided in an ultra-low attachment 96-well plates supplement with FBS or bovine serum albumin (BSA) culture or encapsulated with an alginate supplement with FBS or BSA culture. Meanwhile, estradiol (E 2 ) concentration was assessed through enzyme-linked immunosorbent assay of culture supernatants. The diameter of follicular growth was measured, and the lumen of the follicle was photographed. Spindle microtubules of oocytes were detected via immunofluorescence. The ability of oocytes to fertilize was assessed using in vitro fertilization., Results: The diameters were larger for the growing secondary follicles cultured in ultra-low attachment 96-well plates than in the alginate gel on days 6, 8, and 10 ( p < 0.05). Meanwhile, the E 2 concentration in the BSA-supplemented medium was significantly higher in the alginate gel than in the other three groups on days 6 and 8 ( p < 0.05), and the oocytes in the FBS-free system could complete meiosis and fertilization in vitro ., Conclusions: The present study furnishes insights into the mature oocytes obtained from the 3D culture of the preantral follicle by using ultra-low attachment 96-well plate with an FBS-free system in vitro and supports the clinical practices to achieve competent, mature oocytes for in vitro fertilization., Competing Interests: The authors declare that the research was conducted without any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 The Korean Society of Veterinary Science.)