玻璃化冷冻法对小鼠MⅡ卵纺缍体及染色体的影响

目的:探讨玻璃化冷冻法对小鼠MⅡ卵纺缍体及染色体的毒性。方法:收集小鼠MⅡ卵,分A组(新鲜对照组)、B组、C组和D组(玻璃化冷冻法冻融后分别培养0h、1h和3h固定)及E组、F组和G组(MⅡ卵经玻璃化冷冻液处理后分别培养0h、1h和3h固定),共7组,固定后免疫荧光标记纺锤体和染色体。结果:各组纺锤体正常率无显著差异(P>0.05),染色体正常率:B组显著低于A组(50%vs 87.5%,P<0.01),E组(47.62%)和F组(45%)显著低于A组和G组(75%)(P<0.05)。纺锤体和染色体均正常率:B组显著低于A组(41.67%vs 75%,P<0.05),E组(47.62%)和F组(45%)显著低于A组和G组(75%)(P<0.05)。B、C、D及E、F、G组的细胞骨架正常率依次提高,且玻璃化冷冻组(B组、C组和D组)和冷冻液暴露组(E组、F组和G组)相应时段(0h、1h及3h)比较(即B组与E组、C组与F组及D组与G组比较)均无显著差异(P>0.05)。结论:冷冻保护剂的细胞毒性在玻璃化冷冻损伤中起主要作用,但它对纺缍体的损伤是可逆的。

Effects of vitrification on cytoskeleton of mouse MⅡ oocytes

Objective：To explore the cytoxicity of vitrification on the cytoskeleton of mouse M Ⅱ oocytes. Method： Mice M Ⅱ oocytes were randomly divided into seven groups ： A （control） , B, C, D （ fixed for immunofluorescence of chromosomes and spindles when incubated 0h,1h and 3h respectively,after vitrified-thawed）,E, F and G（fixed when incubated Oh, lh and 3h respectively, after exposed to vitrification protection agents （CPAs） without cooling）. Result： The normality rates of spindles were not significant differences in all groups （ P 〉 0. 05 ）. The normality rate of chromosomes of group B was significantly lower than that of group A （50% vs 87.5% ,P 〈0.01 ） ,and those of group E（47.62% ） and F（45% ） were significantly lower than that of group A and G （ 75 % ） （ P 〈 0.05 ）. The rate of both spindles and chromosomes with normal cdnfiguration of group B was significantly lower than group A（41.67% vs 75% ,P 〈0.05 ） ,and those of group E（47.62% ） and F（45% ） were also significantly lower, compared with group A and G （ 75 % ） （ P 〈 0.05 ）. The normality rates of cytoskeleton were rised from group B to D and from group E to G,and there were no significant differences when group B compared with E, group C compared with F, and group D compared with G （P 〉 0. 05 ）. Conclusion：The toxicity of vitrification protectant agents play an important role in cryoinjury,but the damage to cytoskeleton is reversible.