高浓度葡萄糖对猪卵母细胞体外成熟和早期胚胎发育的影响

试验旨在探究高浓度葡萄糖对猪卵母细胞体外成熟及早期胚胎发育能力的影响。取体外分离处于生发泡期的猪卵丘卵母细胞复合体(COCs),分为3个处理组。分别用含葡萄糖浓度为5.6 mmol/L(C组)、10 mmol/L(G-1组)、15 mmol/L(G-2组)的培养液,进行体外成熟(IVM)处理,42 h后观察,并统计卵丘细胞扩散情况和第一极体排出率;对体外成熟42 h后的卵母细胞孤雌激活,统计2-细胞、4-细胞和第7天囊胚发育。结果发现,G-1组和G-2组卵丘细胞扩散度显著低于C组(P<0.05);G-1组和G-2组的MII期卵母细胞死亡率和存活率与C组相比无显著差异(P>0.05),但G-1组极体率显著降低(P<0.05),G-2组极体率极显著低于C组(P<0.01)。孤雌激活后,与C组相比,G-1组和G-2组的2-细胞分裂率显著降低(P<0.05),4-细胞分裂率以及囊胚发育率均极显著降低(P<0.01),但G-1、G-2组囊胚细胞数量与C组相比无显著性差异(P>0.05)。进一步线粒体染色发现,G-1组和G-2组的线粒体与C组相比分布不均。与C组相比,荧光结果显示G-1组和G-2组不仅活性氧(ROS)水平极显著升高(P<0.01),而且G-1组与G-2组谷胱甘肽(GSH)含量显著降低(P<0.05),虽然G-1组丙二醛(MDA)无显著性差异(P>0.05),但G-2组丙二醛(MDA)含量显著升高(P<0.05)。研究结果表明,葡萄糖浓度高会影响猪卵母细胞线粒体分布,氧化应激水平升高,成熟效率降低,损害早期胚胎的发育潜能。

Effects of high glucose on in vitro maturation and early embryo development of porcine oocytes

The present study was designed to investigate the effects of high glucose on oocytes maturation and early embryo development in vitro.Cumulus oocyte complexes(COCs)were isolated from pig ovaries.Then,5.6 mmol/L glucose(the Control group),10 mmol/L glucose(the G-1 group)and 15 mmol/L glucose(the G-2 group)were applied to in vitro maturation(IVM)of the oocytes,respectively.After 42 h of maturation,the expansion of cumulus cells and the excretion rate of the first polar body were observed in the culture,and the developmental abilities of 2-cells,4-cells and Day 7 blastocysts were analyzed.The results showed that the expansion of cumulus cells in the G-1 and G-2 groups were significantly lower than in the Control group(P<0.05).However,no significant difference was observed in the mortality and survival rates of the MII oocytes among the three groups(P>0.05).After maturation,the rate of oocytes with an extruded first polar body was relatively lower both in the G-1 and G-2 groups(P<0.05,P<0.01).After parthenogenetic activation,the developmental abilities of the embryos from the G-1 and G-2 groups were significantly reduced not only at the cleavage rate at 2-cell and 4-cell stages,but also at the rate of the blastocyst(P<0.01).But no significant difference was observed in the number of blastocyst cells(P>0.05).Further mitochondrial staining revealed that mitochondria in the G-1 and G-2 groups were unevenly distributed,compared with the Control group.In addition,the reactive oxygen species(ROS)level evaluated by staining was significantly increased in the G-1 and G-2 groups(P<0.01),the expression of glutathione(GSH)was also significantly reduced(P<0.05).No significant difference was observed in the malondialdehyde(MDA)of the G-1 group,compared with the Control(P>0.05),but a significantly increased expression of malondialdehyde(MDA)was observed in the G-2 group(P<0.05).The present study indicated that high glucose concentration in IVM affected not only the quality of oocytes but also their development potential.