| 马景.反复超促排卵对老龄小鼠生殖能力的影响[J].浙江中西医结合杂志,2022,32(12): |
| 反复超促排卵对老龄小鼠生殖能力的影响 |
| Effects Of Consecutive Superovulation On ROS-JNK-Mitochondrial Pathway and Ovarian Reserve In Mice |
| 投稿时间：2022-07-01 修订日期：2022-11-17 |
| DOI： |
| 中文关键词: 反复超促排 卵巢储备功能下降 氧化应激 线粒体功能 |
| 英文关键词:Repeated Superovulation Diminished ovarian reserve Oxidative stress Mitochondrial function |
| 基金项目:]:浙江省自然基金(LQ19H270008),全国名老中医药专家传承工作室建设项目(国中医药人教函｛2022｝75号),浙江省中医药“新苗”计划项目(No.浙中医药 [2021]1号) |
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| 中文摘要: |
| 目的 探索反复促排卵对老龄小鼠生殖功能的影响。方法 30只7月龄雌性C57BL/6小鼠，随机分成对照组与模型组，模型组予孕马血清促性腺激素（（Pregnant Mare Serum Gonadotropin, PMSG）腹腔注射，隔48小时后注射人绒毛膜促性腺激素（human chorionic gonadotropin，HCG），隔周一次，连续4周，建立反复超促排卵老龄小鼠模型；对照组仅在第四周注射PMSG、HCG促排卵。第4次促排卵16小时后检测对照组、模型组小鼠血清雌二醇（estrogen ,E2）、孕酮（P）、抑制素B（Inhibin-B ,INHB）水平；HE染色卵巢组织形态学变化，透射电镜观察颗粒细胞超微结构；RT-qPCR检测卵巢JNK、BAX、P53、FOXO3a等基因的表达；体式显微镜下观察卵母细胞形态并检测其活性氧ROS水平。结果 与对照组比较，模型组小鼠血清E2、P、INHB均显著降低P<0.01），卵巢原始卵泡显著减少（P<0.01）、发育卵泡亦减少（P<0.05），黄体数量显著增加（P<0.05）；模型组小鼠卵巢颗粒细胞中嵴模糊及空泡征的线粒体明显增多（P<0.01）；JNK、BAX、P53水平较对照组显著增加（P<0.05），FOXO3a水平较对照组降低（P<0.01）；同时，模型组小鼠的卵母细胞碎裂率及ROS水平均显著增加（P<0.01）。结论 经历4周连续超促排的老龄小鼠卵巢储备功能下降，可获得的卵子质量和数量减少，可能与连续超促排卵导致卵泡内ROS积聚，卵巢氧化应激水平激增，JNK、BAX、P53等凋亡相关基因表达增加，颗粒细胞线粒体功能紊乱有关。由过度氧化应激反应导致这一系列变化后最终导致老龄小鼠卵母细胞质量降低甚至碎裂。 |
| 英文摘要: |
| Objective: to establish a new mouse model of diminished ovarian reserve induced by consecutive superovulation, study the effect of consecutive superovulation on mouse ovarian reserve and demonstrate its deep molecular mechanism. Methods: Thirty C57BL / 6 female mice were selected and randomly divided into control group and model group. The model group was injected intraperitoneally with 15 IU pregnant horse serum gonadotropin. After 48 hours, human chorionic gonadotropin (hCG) was injected, and this procedure was implemented once every other week for 4 weeks to establish the superovulation mouse model. The levels of serum estradiol, progesterone and inhibin B in mice were detected after the last superovulation; The morphological changes of ovary were stained with HE, and the ultrastructure of granulosa cells was observed by transmission electron microscope; RT qPCR was used to detect the expression of JNK, Bax, p53 and FOXO3a genes expressions in ovary; After oocytes were obtained, their morphology was observed under the stereomicroscope and the level of ROS was detected. Results: compared with the control group, the serum E2, P and inhb of the model group were significantly decreased (P < 0.01), the initial ovarian follicles were significantly reduced (P < 0.01), the developmental follicles were also reduced (P < 0.05), and the number of corpus luteum was significantly increased (P < 0.05); In the model group, the mitochondria with fuzzy ridge and vacuolated mitochondria increased significantly (P < 0.01); Compared with the control group, the levels of JNK, Bax and p53 increased significantly (P < 0.05), and the level of FOXO3a decreased (P < 0.01); Meanwhile, the oocyte fragmentation rate and ROS level in the model group increased significantly (P < 0.01). Conclusions：The ovarian reserve function of mice undergoing 4 weeks of continuous superovulation decreased. This may be due to the accumulation of reactive oxygen species in follicles and the surge of ovarian oxidative stress, resulting in the increased expression of apoptosis related genes such as JNK, Bax and p53, and the dysfunction of mitochondria in granulosa cells. This process leads to the reduction or even fragmentation of oocyte quality. |
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