Park Chi-Hun, Lee Sang-Goo, Choi Don-Ho, Lee Chang-Kyu
Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute for Agriculture and Life Science, Seoul National University, Seoul 151-921, Republic of Korea.
Anim Reprod Sci. 2009 Oct;115(1-4):169-81. doi: 10.1016/j.anireprosci.2008.12.004. Epub 2008 Dec 6.
The general method of porcine in vitro fertilization (IVF), involving the co-culture of both gametes in a medium drop, is thought to be the main reason for the high incidence of polyspermy. The aim of this study was to reduce the polyspermic fertilization of porcine embryos during IVF by the modified swim-up method, based on general sperm swim-up technique. Within this design, a 70 microm pore sized cell strainer was used to separate the sperm pellet placed at the bottom of a tube from the mature oocytes placed within the upper region. The separation of gametes using this permeable barrier was to ensure that only motile sperm gained access to the oocytes. It was found that the rate of polyspermy was significantly lowered for the sperm preparations from three boar breeds in modified swim-up method when compared with that of the general microdrop method (p<0.05). However, the penetration rates were found to be similar in both methods for two boar breeds. The average occurrence of blastocysts with more total cell number was higher in the modified swim-up method, while no significant difference in blastocyst rates between the two IVF methods was observed. The frequency of normal diploid embryos was also significantly higher in the modified swim-up method and polyploidy was more frequently observed in microdrop method (p<0.05). Our results demonstrated that the modified swim-up IVF method could reduce polyspermic penetration, and consequently produce better quality and karyotypically normal embryos in porcine IVF.
猪体外受精(IVF)的常规方法是将两性配子在培养基滴中共培养,这被认为是多精受精发生率高的主要原因。本研究的目的是在常规精子上浮技术的基础上,通过改良的上浮法降低猪体外受精过程中猪胚胎的多精受精率。在该设计中,使用孔径为70微米的细胞筛将置于试管底部的精子沉淀与置于上部区域的成熟卵母细胞分离。使用这种可渗透屏障分离配子可确保只有活动精子能够接触到卵母细胞。结果发现,与常规微滴法相比,改良上浮法中来自三个公猪品种的精子制剂的多精受精率显著降低(p<0.05)。然而,发现两个公猪品种在两种方法中的穿透率相似。改良上浮法中总细胞数更多的囊胚平均发生率更高,而两种体外受精方法之间的囊胚率未观察到显著差异。改良上浮法中正常二倍体胚胎的频率也显著更高,多倍体在微滴法中更频繁出现(p<0.05)。我们的结果表明,改良上浮体外受精方法可以减少多精穿透,从而在猪体外受精中产生质量更好且核型正常的胚胎。