中文摘要：

纳米低温保存技术很可能是新一代低温保存技术的重要发展方向,但纳米颗粒应用于卵母细胞玻璃化保存的报道较少.本研究将羟基磷灰石（HA）、二氧化硅、三氧化二铝、二氧化钛等4种纳米颗粒添加到低温保护剂中,使用Cryotop法冷冻猪GV期卵母细胞,使用形态观察和荧光染色的方法,研究其对细胞存活率和发育率的影响.结果显示在实验浓度范围内,HA较其它纳米颗粒对猪卵母细胞的毒性低,当浓度低于0.5％时,细胞发育率为100％；低温保护剂中添加0.1％HA纳米颗粒,发育率比其它组显著提高,可以达到22％,且HA的粒径对结果影响不大；当低温保护剂中添加0.05％粒径为60 nm的HA颗粒时,卵母细胞冷冻复温后发育率会进一步从14.7％提高达到30.4％.在低温保护剂中添加适宜浓度的HA纳米颗粒,可以减少复温过程中的重结晶现象,促进细胞的冷冻存活率和发育率,保存效果与浓度相关,而与纳米颗粒的粒径关系不大.

英文摘要：

Nano-cryopreservation is a promising new way in the next generation of cryopreservation technology; however, using nanoparticles in oocytes vitrification has been rarely reported. This paper investigated the effect of hydroxy apatite （HA） , silica, aluminum oxide, and titanium dioxide nanoparticles in the cryoprotectant on the survival rate and developmental rate of porcine GV- stage oocytes. The cells were cryopreserved in Cryotop and observed using fluorescence staining methods. Results showed that HA nanoparticles have the lowest cytotoxicity among the other nanoparticles, the developmental rate of GV-stage porcine oocytes was 100% when concentrate of HA was lower than 0.5%. When the concentration of HA was 0. 1% in the cryoprotectant, the developmental rate of GV-stage porcine oocytes was 22% in Cryotop, which was significantly higher than that in the other groups. The size of nanoparticles exerted little influence. When 0. 05% HA nanoparticles （60 nm in diameter） were added, the developmental rate increased dramatically from 14.7% in control group to 30. 4%. In conclusion, adding appropriate concentration of HA nanoparticles to cryoprotectant can reduce recrystallization during rewarming and promote survival rate and developmental rate of oocytes after freezing and rewarming. The effect of HA nanoparticles is concentration dependent, while independent to diameters of the nanoparticles.