肥胖引起的线粒体功能及抗氧化异常促进小鼠卵母细胞排卵后老化

Promoting oocyte aging with abnormal mitochondrial function and antioxidant post ovulation in obesity mouse

目的:探讨肥胖对小鼠卵母细胞体内老化、体外受精和胚胎发育的影响及其机制。方法:分别获取人绒毛膜促性腺激素(HCG)注射后14 h、18 h和22 h的卵母细胞-卵丘颗粒细胞复合体,进行体外受精胚胎培养,采用Hoechst 33342和碘化丙啶双染检测卵丘颗粒细胞的凋亡率,JC-1和DCFH-DA分别检测卵母细胞中线粒体膜电位(MMP)和活性氧类(ROS)水平,并检测卵母细胞中ATP和谷胱甘肽(GSH)水平。结果:刚排卵时肥胖小鼠卵母细胞GSH低于对照组(P<0.05),囊胚形成率、ATP、ROS和卵丘颗粒细胞凋亡率两组无显著差别(P>0.05);HCG注射后18 h,肥胖小鼠卵母细胞中ATP含量及囊胚细胞数目开始明显减少并低于对照组(P<0.05),而卵母细胞中ROS水平和卵丘颗粒细胞凋亡率则开始升高并高于对照组(P<0.05);HCG注射后22 h,肥胖小鼠卵母细胞MMP和体外受精囊胚形成率显著下降(P<0.05),并且低于对照组(P<0.05)。结论:肥胖促进排卵后卵母细胞的老化,氧自由基对卵母细胞的损伤可能是引起母源老化不孕的重要原因。

AIM： To investigate the effects of obesity on mouse oocyte in vivo aging, its in vitro fertilization and embryo development. METHODS ： The oocyte - cumulus granulosa cell complexes were obtained at the time points of 14 h, 18 h and 22 h after HCG injection. The in vitro fertilization and embryo culture were conducted. Hoechst 33342 and propidium iodide double staining were performed to determine the apoptotic rate of cumulus granulosa ceils. JC - 1 and DCFH- DA were used to detect the levels of mitochondrial membrane potential （MMP） and reactive oxygen species （ ROS）, respectively. The levels of ATP and glutathione （GSH） in oocytes were also detected. RESULTS： The GSH level in obese mouse oocytes just after ovulation was higher than that in control group （P 〈 0. 05 ）. No difference of blastocyst formation rate, ATP, ROS and cumulus granulosa cell apoptotic rate between obesity group and control group was observed （P 〉0. 05）. However, 18 h after HCG injection, ATP content and blastocyst cell numbers in obese mouse oocytes were less than those in control group （P 〈 0. 05 ）, while ROS level in oocytes and the apoptotic rate of cumulus granulosa cells were higher than those in control group （ P 〈 0. 05 ）. Compared with control group, MMP and blastocyst formation rate were significantly decreased in obesity group 22 h after HCG injection （P 〈 0. 05）. CONCLUSION： Obesity promotes oocyte aging after ovulation, and oxygen free radicals may play an important role in obesity - induced decrease of maternal infertility.