Purpose of the present study was to find the optimal culture conditions for the maturation and fertilization of immature oocytes by the use human body fluids and gonadotropins (Gn) in the mouse model. Cumulus-enclosed mouse immature oocytes were
incubated in the medium containing various human body fluids with or without Gn in vitro, and examined to confirm nuclear maturation (NM) and fertilization. Female ICR mice were stimulated with 7.5IU pregnant mares' serum gonadotropin (PMSG).
Cumulus-enclosed immature oocytes were isolated at 48-52 hr post PMSG injection and cultured in TCM 199 supplemented with various concentrations (20, 50, and 70%) of human body fluids such as fetal cord serum (hCS), follicular fluid (hFF),
peritoneal
fluid (hPF) and amniotic fluid (hAF) in the presence or absence of 10 IU/ml PMSG and 10 IM/ml human chorionic gonadotropin (hCG) for 18 hr. Fetal calf serum (FCS) was used as a control for the supplements. Matured oocytes were fertilized with
sperm
collected from the epididymis of male mice. Fertilization was conducted in T6 medium containing 15 mg/ml bovine serum albumin, and confirmed at 6 hr post-insemination. Evaluation of nucler maturation and fertilization was carried out by rapid
staining
using fuchin. There was no significant difference between the effects of human body fluids and FCS supplements on nuclear maturation of cumulus enclosed mouse immature oocytes. When maturation medium was supplemented with 20% hPF or 20% hAF,
fertilization rates were significantly (P<0.01) lower than that of 20% FCS, hCS and hFF groups. However, higher concentrations of body fluids during IVM were not more beneficial on fertilizability of oocytes. The addition of Gn significantly
increased
the fertilization rates in hPF and hAF groups (hPF without Gn; 51.5%, compared with 85.1% for addition of Gn, and hAF without Gn; 30.1% compared with 85.8% for addition of Gn) at 20% concentration. These results suggest that human body fluids at
20%
concentration and gonadotropins can be used as supplements for the maturation of mouse immature oocytes in vitro. When gonadotropins supplemented with the human body fluids in the maturation medium, fertilizability of mouse immature oocytes was
increased in hPF and hAF groups. These results can be applied to maturation of human immature oocytes in vitro.
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