[Establishment of in-vitro mouse preantral follicle culture systems].
OBJECTIVE
To develop mouse preantral follicle culture for toxicology study.


METHODS
The ovaries were from prepubertal mice (C57B1/6JxCBA/Ca) (at the ages of 12-14 days). The ovaries were from prepubertal mice (C57B1/6JxCBA/Ca) (at the ages of 12-14 days). The preantral follicles with a diameter around the range 100-130 microm were mechanically dissected and randomly allocated to 96-well plates. After subsequently cultured individually for 12 days, the follicles were induced ovulation. The in-vitro developmental characteristics of the follicles were observed including hormonogenesis, folliculogenesis and oogenesis 16h later after ovulation. In-vitro growth and maturation of oocytes (IVG) were compared with in-vivo growth and in-vitro maturation of oocytes (IVM) to assess the in-vitro assay.


RESULTS
89.74% (376/419) of follicles were visibly associated with theca cells and possessed a close follicle (granulosa) cell/oocyte apposition. At 12 days of culture, the average diameter of follicles were increased, the survival rate of follicles is 96.81% (364/376), 48.94% (184/376) of the cultured follicles reached the pre-ovulatory stage with a large antrum-like cavity, and 56.38% (212/376) COCs were released by maturation induction in this study. From preantral follicle to oocyte maturation and the formation of first polar body, the morphological observation showed that developmental characteristics of mouse preantral follicles in-vitro were similar to those in-vivo. The patterns of hormone secretion observed in preantral follicle culture were similar to the characteristics of in-vivo follicle hormone secretion. The numbers of oocytes with germinal vesicle (GV) did not differ between IVG and IVM groups, 18.89% oocytes form the first polar body in IVG groups and 53.53% oocytes form the first polar body in IVM groups. No abnormal chromosome segregation found in this study.


CONCLUSIONS
This study successfully developed in-vitro mouse pre-antral follicle culture system, which could be a efficient tool to study folliculogenesis physiology and toxicology system, which could be a efficient tool to study folliculogenesis physiology and toxicology.
Wan
Xuying
X
Department of Health Toxicology, Second Military Medical University, Shanghai 200433, China. wanxuying711@sina.com.cn
Zhu
Yuping
Y
Ma
Xili
X
Zhu
Jiangbo
J
D000818:Animals; D005260:Female; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D009865:Oocytes / Q000166:cytology* / Q000378:metabolism; D006080:Ovarian Follicle / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D010053:Ovary / Q000166:cytology; D010062:Ovulation Induction; D046509:Tissue Culture Techniques / Q000379:methods*