肌肽对小鼠卵母细胞体外成熟和胚胎发育的影响  

Effects of carnosine on mouse oocyte in vitro maturation and embryo development

作　　者：包云 余柯达[1] 柴娟[1] 毛佳婷 师帅[1] Bao Yun;Yu Keda;Chai Juan;Mao Jiating;Shi Shuai(Reproductive Medical Center,Jinhua People's Hospital,Jinhua 321000,China)

机构地区：[1]金华市人民医院生殖医学中心,金华321000

出　　处：《中华生殖与避孕杂志》2023年第11期1146-1152,共7页Chinese Journal of Reproduction and Contraception

基　　金：浙江省医药卫生科技计划项目(2022KY1333);金华市科学技术研究计划项目(2019-4-030)。

摘　　要：目的探讨肌肽对小鼠卵母细胞体外成熟和胚胎发育的影响。方法获取小鼠卵丘-卵母细胞复合体(cumulus-oocyte complexes,COC),分别用含10μmol/L、30μmol/L、50μmol/L、100μmol/L、200μmol/L肌肽的体外成熟培养液培养18 h,不含肌肽的培养液设为对照组。根据卵母细胞成熟率、卵裂率和囊胚形成率确定作用的最优浓度。通过检测活性氧(reactive oxygen species,ROS)水平、总谷胱甘肽(total glutathione,T-GSH)含量、皮质颗粒分布、线粒体分布和线粒体拷贝数进一步验证肌肽的作用。结果肌肽浓度为100μmol/L时,卵母细胞成熟率[69.23%(135/195)]、卵裂率[66.06%(72/109)]和囊胚形成率[48.61%(35/72)]最高,与对照组[44.44%(84/189)、38.67%(29/75)、20.69%(6/29)]相比,差异具有统计学意义(P<0.001、P<0.001、P=0.010),故选定100μmol/L作为肌肽作用的最优浓度。对照组卵母细胞中的ROS和T-GSH水平为100μmol/L肌肽组的2.04倍和0.64倍,差异具有统计学意义(均P<0.001)。100μmol/L肌肽组的皮质颗粒Ⅲ级的比例显著高于对照组(P<0.001)。对照组卵母细胞的线粒体均匀分布在胞质中,100μmol/L肌肽组的线粒体聚集在胞质中央区域,且荧光强度高于对照组(P=0.040)。100μmol/L肌肽组的线粒体拷贝数是对照组的1.72倍,差异具有统计学意义(P<0.001)。结论肌肽能够有效改善未成熟卵母细胞的氧化应激,促进小鼠卵母细胞的体外成熟和胚胎发育。ObjectiveTo explore the effects of carnosine on in vitro maturation(IVM)of mouse oocytes and the developmental competency of embryos.MethodsThe mouse cumulus-oocyte complexes(COC)were isolated and cultured in the IVM medium containing 10μmol/L,30μmol/L,50μmol/L,100μmol/L,or 200μmol/L carnosine for 18 h.IVM medium without carnosine was acted as control group.The optimal concentration was chosen according to the rates of oocyte maturation,embryo cleavage and blastocyst formation.The effect of carnosine was further verified by detecting the levels of reactive oxygen species(ROS),and total glutathione(T-GSH),the distribution of cortical granules and mitochondria,and also the copy numbers of mitochondria.ResultsIn the 100μmol/L carnosine group,the rates of oocyte maturation[69.23%(135/195)],embryo cleavage[66.06%(72/109)]and blastocyst formation[48.61%(35/72)]were the highest and showed statistically significant differences(P<0.001,P<0.001,P=0.010)when compared with control group[44.44%(84/189),38.67%(29/75),20.69%(6/29)].Therefore,100μmol/L was selected as the optimal concentration of carnosine.The levels of ROS and T-GSH in control group were 2.04 and 0.64 times as those in the 100μmol/L carnosine group(both P<0.001).The percentage of gradeⅢcortical granules in the 100μmol/L carnosine group was significantly higher than that in control group(P<0.001).The mitochondria of oocytes in control group were uniformly distributed,while the mitochondria in the 100μmol/L carnosine group gathered in the central cytoplasmic region,and the fluorescence intensity was higher than that in control group(P=0.040).The mitochondrial copy number in the 100μmol/L carnosine group was as much 1.72 times as that in control group(P<0.001).ConclusionCarnosine effectively improves oxidative stress in the immature oocytes and promotes the mouse oocyte IVM and embryo development.

关 键 词：肌肽 胚胎发育 线粒体 未成熟卵母细胞 体外成熟 

分 类 号：R714.8[医药卫生—妇产科学]