单层细胞共培养对猪去卵丘卵母细胞体外成熟和孤雌发育的影响

Effects of cell monolayer co-culture on porcine cumulus cells denuded oocytes during in vitro maturation and parthenogenetic development

本试验比较观察第一极体(The first polar body,PbⅠ)、Oosight imaging system观察和hocchst33342染色法对第2次减数分裂中期(Metaphase Ⅱ,MⅡ)卵母细胞判定结果的相关性分析,并探讨卵巢皮质细胞(porcine ovarian cortex cells,pOCCs)、猪输卵管上皮细胞(porcine oviductal epithelial cells,pOECs)和猪卵丘颗粒细胞(porcine cumulus cells,pCCs)等3种单层细胞体外共培养体系对猪去卵丘卵母细胞(cumulus cells denuded oocytes,Dos)体外成熟(in vitro maturation,IVM)和孤雌发育的影响。结果显示:(1)Oosight imaging system判定卵母细胞成熟的结果与hocchst33342染色法判定的结果有很强相关性(R=0.973,P<0.01,N=90);(2)pOCCs单层细胞共培养体系中猪去卵丘卵母细胞成熟率显著高于pOECs((52.5±0.30)%vs(43.8±2.18)%,P<0.05),且这2组成熟率显著高于其余各组(P<0.05);Dos在3种单层细胞共培养体系培养后,成熟卵母细胞GSH含量较非共培养组都能显著升高(P<0.05),pOCCs组GSH含量最高((7.1±0.32)pmol/卵);(3)成熟后的Dos经孤雌激活,pOCCs组和pOECs组卵裂率和囊胚率都显著高于其余2组(P<0.05)。

The objective of the experiment was to compare the correlation of three ways to determine matured MII oocytes,including the first polar body,oosight imaging system observation and hocchst33342 staining,and to evaluate the effects of pOCCs,pOECs and pCCs monolayer co-culture system on porcine cumulus cells denuded oocytes during in vitro maturation and parthenogenesis,respectively.The results show that：（1）determining the outcome of oocyte maturation,the result of oosight imaging system is significant correlation with that of the hocchst33342 staining（R=0.973,P0.01,N=90）;（2） Comparing with pOECs group,the rate of Dos maturation in pOCCs group in vitro maturation is significantly higher（（52.5±0.30）% vs（43.8±2.18）%,P0.05）,and the rates in the pOCCs group and pOECs group are both significantly higher than that in the other groups（P0.05）;After Dos culturing in vitro maturation in three kinds of cell monolayer co-culture,GSH content significantly increased（P0.05）,pOCCs group had the highest content（（7.1±0.32）/（pmol·egg-1）;（3）Mature Dos after parthenogenetic activation,the cleavage rate and blastocyst rate in pOCCs group and pOECs group are significantly higher than in the other two groups（P0.05）.