不同激活方法对小鼠卵母细胞孤雌激活的实验观察

Effects of Different Treatments on the Parthenogenetic Activation of Mouse Oocytes

目的 建立合适的小鼠卵母细胞孤雌激活方法。方法 取不同卵龄小鼠卵母细胞 ,运用不同浓度的氯化锶和不同强度的电脉冲对其进行活化 ,观察小鼠卵母细胞激活率和体外发育状况。结果 ① 14、16和 18h卵龄组卵母细胞经 10mmol LSrCl2 处理后 ,三组的激活率随卵龄的增长而提高 ,分别为 2 1 1%、4 1 8%和 73 7% ,组间差异有统计学意义 (P <0 0 5 ) ;卵母细胞离体后在激活前体外培养 3h也能提高其激活率 ,但当卵龄达到一定时 (18h) ,体外培养激活率不再提高。② 5 0、10 0、15 0mmol L三种浓度的SrCl2 均能有效地激活小鼠卵母细胞 ,激活率分别为 5 8 5 %、6 8 95和 70 9% ,与对照组和 1 6mmol L组相比差异有统计学意义 (P <0 0 5 )。③卵母细胞的激活率随处理时间的延长而提高 ,30、6 0、12 0、2 4 0minSrCl2 处理时间的激活率分别为 4 4 9%、5 6 1%、6 8 9%、77 0 % ,相隔两组之间差异有统计学意义 (P <0 0 5 )。④ 1 5kV cm ,16 0 μs和 1 0kV cm ,32 0 μs脉冲刺激下的卵母细胞激活率显著高于 1 8kv cm ,90 μs脉冲刺激组 (5 8 5 %vs2 7 1%、6 9 1%vs2 7 1% ,P <0 0 5 ) ,前两组之间差异无统计学意义 (P >0 0 5 )。结论 小鼠卵母细胞孤雌激活率与卵龄和激活方案有关 ;氯化锶浓度。

Objective In order to study the mammalian parthenogenesis, increase the efficiency of nuclear transfer, and establish the optimum procedure of oocyte activation in mouse. Methods Oocytes were recovered from mice killed at different hours post denuded of cumulus with hyaluronidase. The cleavage rate and blastocyst development rate of mouse oocytes activated by different strontium chloride (SrCl 2) concentration and pulse strength were investigated. Results ①The cleavage rate of oocytes with the age of 14,16,18 hours were respectively 21.1%, 41.8%, 73.7% after treatment with 10mmol/L SrCl 2 (P<0.05); The cleavage rate could increase after the oocytes were cultured 3 hours in vitro before activation, but it won’t increase when the oocyte age was 18 hours up. ② The cleavage rate of oocytes treated with 5.0, 10.0, 15.0mmol/L SrCl 2 were respectively 58.5%, 68.9%, 70.9%, all were significantly higher than that of the control group and the 1.6mmol/L group (P<0.05). ③ The cleavage rate of oocytes treated with SrCl 2 for 30, 60, 120, 240min were 44.9%, 56.1%, 68.9%, 77.0%. In other words, it increased with the extension of treatment time. Significant differences were existed every other group (P<0.05). ④ The cleavage rates of oocytes treated with 1.8kV/cm for 160μs or 1.0kV/cm for 320μs were higher than that of 1.8kv/cm for 90μs (58.5%vs27.1%,69.1%vs27.1%,P<0.05). There were no remarkable differences between the former two groups(58.5%vs 69.1%, P>0.05). Conclusion The activation rates of mouse oocytes were associated with oocytes age and activation protocols. The SrCl 2 concentration, the duration subjected to SrCl 2 and the pulse strength have great effect on mouse oocytes activation.