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目的:研究骨髓间充质干细胞(mesenchymal stem cell,MSC)条件培养液对小鼠未成熟卵母细胞体外成熟、受精及胚胎发育的影响。方法:分离、培养小鼠MSC,获取MSC条件培养液(conditioned medium of MSC,CM)。收集小鼠生殖泡(germinal vesicle,GV)期卵母细胞,根据有无完整颗粒细胞层包裹颗粒细胞-卵母细胞复合物(cumulus-oocyte complex,COC)分为2类,分别在4种培养基(CM、DMEM、α-MEM、HTF)中培养不同时间,观察卵母细胞核成熟情况,荧光标记后激光共聚焦检测皮质颗粒(cortical granules,CG)分布和含量变化;体外受精后观察受精率和囊胚形成率。结果:2类COC卵母细胞的成熟率CM组明显高于DMEM组、α-MEM组、HTF组(P<0.01);体外受精率和囊胚形成率CM组明显高于DMEM组、HTF组(P<0.01),与α-MEM组和体内成熟组间差异无统计学意义(P>0.05);CM组卵母细胞内CG分布和含量变化与核成熟同步。结论:CM能使小鼠卵母细胞核、质同步成熟,提高了成熟率和卵母细胞质量。
Objective: To study the effect of mesenchymal stem cell (MSC) conditioned medium on mouse immature oocyte maturation, fertilization and embryo development. METHODS: Mouse MSC were isolated and cultured to obtain conditioned medium of MSC (CM). The oocytes of mice with germinal vesicle (GV) were collected and divided into two groups according to the presence or absence of intact granulosa cells (COC) (CM, DMEM, α-MEM, HTF) were cultured in different time to observe the nuclear maturation of oocytes. The distribution and content of cortical granules (CG) were detected by confocal laser scanning microscopy after in vitro fertilization. And blastocyst formation rate. Results: The maturation rates of two types of COC oocytes in CM group were significantly higher than those in DMEM group, α-MEM group and HTF group (P <0.01). In vitro fertilization rate and blastocyst formation rate in CM group were significantly higher than those in DMEM group and HTF group (P <0.01). There was no significant difference between α-MEM group and in vivo maturation group (P> 0.05). The distribution and content changes of CG in oocytes in CM group were synchronized with nuclear maturation. Conclusion: CM can make mouse oocytes mature synchronously in nuclei and cytoplasm, improve maturation rate and oocyte quality.