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目的:拟通过特定穿刺实验来分析小鼠卵子胞膜在显微穿刺中存活的膜特性因素。方法:采用尖口注射针及37℃恒温台的显微条件下,①单纯穿刺实验:即单纯对卵子穿刺不同时间长度(1 s或60 s)或不同深度(50μm和90μm)共4个组合组(每组n=20),比较膜凹陷的回复时间;②显微授精(ICSI)实验:延时组(n=91),在穿刺时延缓对小鼠卵子抽吸破膜的时间;加深组(n=140),改良显微持卵管,使呈喇叭形开口,可在穿刺持卵时将卵子部分吸入该开口内,以增加注射针穿刺小鼠卵子的深度;对照组(n=115),即相同于传统的人ICSI操作。结果:①穿刺时间更长或更深可显著延长膜凹陷的恢复时间;②与对照组相比,延缓对卵子的抽吸破膜时间使得小鼠卵存活率显著提高(22%vs 48%,P<0.01);而应用改良的新型持卵管后,小鼠卵存活率更加显著地提高(22%vs 82%,P<0.01),且其正常受精率>70%。结论:小鼠卵子胞膜并非易脆;在显微穿刺时小鼠卵子膜凹陷柔顺性随时间和深度增加而增加,从而延缓膜凹陷回复时间,有利卵子胞膜破口修复而存活。
OBJECTIVE: To analyze the membrane characteristics of mouse oocyte membrane during micro-puncture through specific puncture experiments. Methods: Under the microscopic conditions of acupuncture needle and 37 ℃ thermostatic station, ① purely puncture test: 4 pairs of oocyte puncturing for different time (1 s or 60 s) or different depth (50 μm and 90 μm) (N = 20 in each group) to compare the recovery time of membrane depression. ② ICSI: Delayed group (n = 91) (N = 140). The microscopic oviduct was modified to make a trumpet-like opening which could ingest part of the egg into the opening during the process of pricking and holding eggs to increase the depth of the ovum in mice injected with the needle. The control group (n = 115), that is the same as traditional ICSI operations. Results ① The longer or deeper puncture time could significantly prolong the recovery time of membrane denture. ② Compared with the control group, the delay of ovum rupture time significantly increased the survival rate of mice (22% vs 48%, P <0.01). However, the survival rate of mouse ovaries increased significantly (22% vs 82%, P <0.01) after application of a new modified oviduct, and the normal fertilization rate was> 70%. CONCLUSION: The membrane of mouse ovum is not brittle; while the flexibility of mouse ovum membrane increased with time and depth during microscopic puncture, which delayed the recovery time of membrane denture and survived the repair of membrane rupture of ovum.