| Objective:Using 2～6mm bovine follicular oocytes from slaughter house:1 To study whether human follicular fluid can be used as in vitro maturation medium for bovine immature oocytes2 To study the influence of inactivated human follicular fluid, culture temperature , culture volume on bovine oocytes matured in vitro;3 To study the influence of Sodium Ferulate on bovine immature oocytes when Sodium Ferulate was added into in vitro maturation medium used in our laboratory (routine IVM medium).Methods:1 Two hundred and ninty one bovine oocytes were divided into 2 groups. One group used routing IVM medium as IVM medium and the other group used HFE The differences of IVM rates between the two groups were compared.2.1 Seven hundred and twenty four bovine oocytes were divided into 3 groups.The control group used routing IVM medium as IVM medium and the other two groups used inactivating HFF and uninactivating HFF. The differences of IVM rates among the three groups were compared.2.2 Five hundred and fourty two bovine oocytes were used in this experiment and divided into 4 groups. Using HFF as IVM medium, the culture temperature of the four groups were 37.5℃, 38℃, 38.5℃and 39℃respectively. The differences of IVM rates among the four groups were compared.2.3 Four and seventy eight bovine oocytes were used in this experiment and divided into 2 groups. Using HFF as IVM medium, the culture container of the two groups were 4-well plate and micro droplet respectively. The differences of IVM rates among the two groups were compared.3 Eight hundred and fifty five bovine oocytes were used in this experiment and divided into 4 groups. The control group used routine IVM medium and the other three groups were used routine IVM medium added 0.5mg/ml,1 mg/ml or 2mg/ml Sodium Ferulate respectively. The differences of IVM rates among the two groups were compared.Results:1 The IVM rates of HFF group and routine IVM medium group were 58.5% and 63.5% respectively. There were no significant differences between the two groups (P＞0.05).2.1 The IVM rates of routine IVM medium group, inactivating HFF group and uninactivating HFF group were 64.7% , 37.5% and 58.8% respectively. There were significant difference between routine IVM medium group and inactivating HFF group (P＜0.05) but no significant difference between routine IVM medium group and uninactivating HFF group (P＞0.05).2.3 The IVM rates of 37.5℃, 38℃, 38.5℃and 39℃group were 40.7% , 60.7% 58.1% and 54.1% respectively. There were significant difference between 37.5℃group and other three groups (P＜0.05) but no significant difference among the three groups(38℃, 38.5℃and 39℃) (P＞0.05).2.4 The IVM rates of the two groups (using 4-well plate or routine micro droplets) were 62.1% and 65.0% respectively and there were no significant difference between the two groups (P＞0.05).3 The IVM rates of the four groups, routine IVM medium, routine IVM medium with 0.5mg/ml Sodium Ferulate, routine IVM medium lmg/ml Sodium Ferulate and routine IVM medium 2mg/ml Sodium Ferulate were 60.1%,44.2%,53.1% and 41.6% respectively. There were significant difference between the routine IVM medium and routine IVM medium with 0.5mg/ml Sodium Ferulate (P＜0.05). There were significant difference between the routine IVM medium and routine IVM medium with 2mg/ml Sodium Ferulate (P＜0.05). The differences between the routine IVM medium and routine IVM medium with 1mg/ml Sodium Ferulate were not significant (P＞0.05).Conclusions:1 Follicular fluid may be used as a better kind of medium for culturing bovine oocytes.2.1 Follicular fluid which was not inactivated may be used as a kind of medium for culturing oocytes under steady cultural circumstance.2.2 When uninactivating follicular fluid was used be IVM medium, the culture temperature can be set between 38℃.2.3 Four-well plates which volume is larger than conventional culture service can simplify the procedure of in vitro production of bovine embryos and may be used as a kind of medium for culturing oocytes and embryos under steady cultural circumstance.3 Sodium ferulate could not inhibit the bovine nuclear maturation and Sodium ferulate can be used in the further experiment: the influence of Sodium ferulate on the cytoplasm maturation in vitro in bovine oocytes. |
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