Comparison of chemical, electrical, and combined activation methods for in vitro matured porcine oocytes
Comparison of chemical, electrical, and combined activation methods for in vitro matured porcine oocytes
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猪体外成熟卵母细胞化学、电和联合激活方法的比较
DOI：
10.1007/s11626-014-9819-1
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发表时间：
2015-02-01
影响因子：
2.1
通讯作者：
Shi, De Shun
中科院分区：
文献类型：
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作者：
Liu, Shuai;Cui, Kuiqing;Shi, De Shun
对影响猪卵母细胞活化的因素进行了系统研究。本研究包括（1）离子霉素加各种化学试剂对激活的影响，（2）不同电激活参数的比较，（3）组合激活的优化，以及（4）优化方案的评估。结果显示：（1）离子霉素（Ion）+6-二甲氨基嘌呤（6-DMAP）的囊胚率（29.7±1.1%）、Ion+细胞松弛素B（CB）+放线菌酮（CHX）（29.8±1.2%）、Ion+CB+6-DMAP (30.4±1.6%)，Ion + CB + CHX + 6-DMAP (30.2±2.7%) 显着高于 Ion + CHX (15.8±1.5%，p<0.05)； (2)当卵母细胞被三个1.00kVcm−1的直流脉冲激活80μs时，电激活的单性生殖囊胚形成最佳(39.5±1.1%)； （3）DC+CB+CHX（55.4±1.2%）和DC+CB+6-DMAP（50.4±2.9%）的囊胚率显着高于DC+6-DMAP、DC+CB+CHX+6-DMAP、电激活和单独化学激活（p<0.05）； (4)优化方案产生的孤雌囊胚中约84%是二倍体，显着高于电激活囊胚(40%)。使用优化的电激活和组合激活方案，胞浆内单精子注射（ICSI）（34.6±1.1%）、细胞质显微注射（CI）（52.3±2.2%）和手工克隆（HMC）（31.2±1.0%）分别产生高囊胚率。本研究得出的结论是，体外成熟猪卵母细胞的最佳激活方案是将激活与参数相结合，即三个 1.00 kV cm−1 的直流脉冲持续 80 μs 加 CB 和 CHX 处理。
Factors influencing porcine oocyte activation were systematically studied. This study included (1) the effect of ionomycin plus various chemical agents on activation, (2) comparison of different electrical activation parameters, (3) optimization of combined activation, and (4) evaluation of the optimized protocols. The results showed that (1) blastocyst rates of ionomycin (Ion) + 6-dimethylaminopurine (6-DMAP) (29.7 ± 1.1%), Ion + cytochalasin B (CB) + cycloheximide (CHX) (29.8 ± 1.2%), Ion + CB + 6-DMAP (30.4 ± 1.6%), and Ion + CB + CHX + 6-DMAP (30.2 ± 2.7%) were significantly higher than Ion + CHX (15.8 ± 1.5%,p< 0.05); (2) the parthenogenetic blastocyst formation of electrical activation was optimal when oocytes were activated by three direct current (DC) pulses of 1.00 kV cm−1for 80 μs (39.5 ± 1.1%); (3) blastocyst rates of DC + CB + CHX (55.4 ± 1.2%) and DC + CB + 6-DMAP (50.4 ± 2.9%) were significantly higher than DC + 6-DMAP, DC + CB + CHX + 6-DMAP, electrical activation, and chemical activation alone (p< 0.05); and (4) approximately 84% of parthenogenetic blastocysts yielded by the optimized protocol were diploid, which was significantly higher than that of electrical activation blastocysts (40%). Using the optimized electrical and combined activation protocol, high blastocyst rates were generated by intracytoplasmic sperm injection (ICSI) (34.6 ± 1.1%), cytoplasmic microinjection (CI) (52.3 ± 2.2%), and handmade cloning (HMC) (31.2 ± 1.0%), respectively. This study concludes that the optimal activation protocol of in vitro matured porcine oocytes was combined activation with parameter as three DC pulses of 1.00 kV cm−1for 80 μs plus CB and CHX treatment.