化学剂法对兔卵母细胞孤雌激活的效果

Parthenogenetic activation of rabbit oocytes by chemical agents

目的研究钙离子载体A23187和6-二甲基氨基嘌呤（6-DMAP）及3种酶对兔卵母细胞的激活率,并观察其体外发育情况。方法兔超排后获得卵母细胞,经脱颗粒处理后用A23187和6-DMAP处理不同时间,体外培养5-7d,观察细胞发育与囊胚形成情况。结果钙离子载体A23187激活处理5min后,继续在2.0mmol/L6-DMAP中分别处理3.5、4.0、4.5、5.0h,均能激活兔卵母细胞;以处理4.0h的卵裂率最高（58.82%,20/34）,并有15%（3/20）的囊胚发育率。透明质酸酶、胶原酶和胰酶处理20min后,均能激活兔卵母细胞,其中透明质酸酶处理20min的激活率和卵裂率最高,分别为58.33%（28/48）和63.64%（28/44）。结论兔卵母细胞孤雌激活率与激活方法密切相关。化学激活剂的处理时间及消化酶的不同均对兔卵母细胞的激活产生显著影响。

Objective To investigate the parthenogenetic activation of rabbit ooeytes by chemical agents including calcium ionophore A23187, 6-dimethylaminopurine （6-DMAP） and three different enzymes, and to observe the in vitro parthenogenetic development. Methods Cumulus-oocyte complexes were harvested from ovaries of superovulated rabbits. After removing the cumulus cells, the oocytes were treated with A23187 and 6-DMAP for a different time period. After in vitro culture for 5 - 7 d, parthenogenetic development of stimulated oocytes was assessed by microscopic observation. Results The oocytes treated with A23187 for 5 rain followed by 6- DMAP treatment （2.0 mmoL/L） for 3.5, 4.0, 4.5 and 5 h could activate the cells. However, the highest cleavage rate （58.82% , 20/34） and blastocyst formation rate （ 15% , 3/20） were achieved from 4.0 - h treatment of 6-DMAP group. Treatment of oocytes with hyaluronidase, collagenase or trypsin respectively could also activate the ceils. Twenty minutes of treatment with hyaluronidase yielded a best activation rate（58.33% , 28/48） and cleavage rate （63.64% , 28/44）. Conclusion The parthenogenetic activation of rabbit oocytes is closely related with the chemicals used. Different treatment time period and different digestive enzyme may result in different activation rates.