This study aimed to recover the ovarian function through in vitro culture of preantral follicles from aged mice. First, we isolated the preantral follicles from ovaries of sixty-seven-week old B6D2F1 mice with decreased fecundity to know how many follicles were present in them, which was 6 preantral follicles including 2 primary, 2 early secondary and late secondary follicles from 8 aged mice. It was confirmed that a few follicles (~2) were present in aged mice through histological analysis compared to adult mice as control. The 9 days of in vitro culture of preantal follicles showed in vitro growth and induced maturation after treatment with hCG (2.5 IU/mL) and EGF (5 ng/mL). Cumulus cells in the cumulus-oocyte complexes (COCs) were removed using hyaluronidase and oocytes at the germinal vesicle (GV) and GV breakdown (GVBD) were obtained from preantral follicle culture of aged mice in vitro. In conclusion, these observations demonstrated that there still were a few preantral follicles in the ovaries of 67 week-old mice, which we were able to culture in vitro and oocytes were obtained from them. This study proposed an in vitro culture system using preantral follicle as a therapeutic strategy for fertility preservation in humans for assisted reproductive medicine.

ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
  Animals and materials
  Isolation of preantral follicles
  In vitro culture of preantral follicles
  In vitro maturation of oocytes in antral follicles
 RESULTS
 DISCUSSION
 CONFLICTS OF INTEREST
 ACKNOWLEDGEMENTS
 REFERENCES

• Jung-Ah Yoon(Fertility Center of CHA Gangnam Medical Center, College of Medicine, CHA University)
• Jung-Kyu Choi(Department of Physiology, School of Medicine, CHA University, Department of Biotechnology, College of Life and Applied Sciences, Yeungnam University) Correspondence