低温对小鼠卵母细胞纺锤体损伤和发育潜力的影响

Effects of Cooling and Rewarming on the Meiotic Spindle and the Developmental Potential of the Mouse Oocytes

为了评价低温对小鼠卵母细胞纺锤体损伤和发育潜力的影响以及损伤的可逆性,MII期卵母细胞在24℃室温或4℃环境下分别处理1、2、4或24h,并于37℃下恢复培养1或2h,复温或未经复温的卵孤雌激活后分析早期发育潜力,并对处理的卵通过免疫荧光细胞化学技术检测纺锤体的损伤和重建动态。结果显示纺锤体受低温打击的受损程度明显依赖于处理温度和时间,低温处理对纺锤体的损伤具有较强的可逆性,不论24℃或4℃,处理时间不超过4h的卵均能重建正常的纺锤体构像,但孤雌激活胚胎的早期发育潜力仍然受到明显的影响,而24h低温处理导致了纺锤体构像的不可逆破坏。对不同卵龄小鼠卵母细胞的研究发现卵龄大小不影响低温打击后的卵发育潜力。这些结果提示在以卵母细胞为材料的细胞工程技术的操作过程中应尽可能缩短室温操作时间。

In order to evaluate the effects of cooling on the meiotic spindles, development potential of mouse oocytes and the reversibility of the spindle＇ s damage, the Metaphase-II mouse oocytes were exposed at room temperature （24℃） or 4 ℃ for 1, 2, 4 or 24 h and part of the oocytes renew by culturing at 37 ℃ for 1 or 2 h. The early developmental potential of these oocytes was analyzed by parthenogenetic activation and the dynamic of damage and reconstruction of the spindles was detected by confocal microscopy. The results displayed that the damage of the spindles after cooling relied obviously on the temperature and time of the treatment. The damage of spindles after low temperature treatment for less than 4 h, in spite of 24 or 4 ℃, was strongly reversible, and the oocytes could reconstruct normal spindle configuration, but the early developmental potential of parthenogenetic embryos derived form these oocytes was evidently decreased. The different oocyte＇s age had no influence to the injury of the oocytes by cooling. The results suggest that the operative time of cell engineering procedure using oocytes should be shorten as much as possible.