维生素A对牦牛卵母细胞体外成熟及后续胚胎发育能力的影响

doi:10.16431/j.cnki.1671-7236.2022.12.020

摘要/Abstract

摘要： 【目的】探究维生素A对牦牛卵母细胞体外成熟及后续胚胎发育能力的影响。【方法】以牦牛卵母细胞为研究对象，在其体外成熟培养液中分别添加0（对照组）、2、5、10和20 μmol/L维生素A，体外培养24 h统计第一极体排出率；对成熟后各组卵母细胞进行孤雌激活，在孤雌激活胚胎培养的第2和8天分别统计卵裂率和囊胚率；用实时荧光定量PCR检测各组MⅡ期卵母细胞中维生素A调控卵母细胞成熟典型信号通路中的节点基因RARα、RARβ、RARγ、RXRα、RXRβ、RXRγ、STRA8及非典型信号通路中的节点基因MEK和ERK1的相对表达量，筛选最佳维生素A处理浓度。在体外成熟培养液中添加最佳浓度维生素A，成熟6和24 h分别收集MⅠ和MⅡ期卵母细胞，将部分MⅡ期卵母细胞进行孤雌激活，收集激活8 d的囊胚，用实时荧光定量PCR检测GV、MⅠ和MⅡ期卵母细胞及孤雌激活囊胚中RARα、RXRα、STRA8基因的相对表达量。【结果】与对照组相比，2、5和10 μmol/L维生素A组第一极体排出率和卵裂率均显著提高（P<0.05），且2 μmol/L维生素A组均达到最高；2 μmol/L维生素A组囊胚率显著提高（P<0.05），20 μmol/L维生素A组第一极体排出率、卵裂率和囊胚率均显著降低（P<0.05）。实时荧光定量PCR结果表明，与对照组相比，2、5、10和20 μmol/L维生素A组RARα、RXRα和STRA8基因的相对表达量均显著增加（P<0.05），其中2 μmol/L维生素A组均达到最高，因此2 μmol/L维生素A对牦牛卵母细胞体外成熟的效果最好。与GV期卵母细胞相比，STRA8、RXRα、RARα基因的相对表达量在MⅡ期卵母细胞均极显著增加（P<0.01），在MⅠ及囊胚期差异均不显著（P>0.05）。【结论】在体外成熟过程中，添加2 μmol/L维生素A可以促进牦牛卵母细胞的成熟，能够显著提高孤雌激活胚胎的卵裂率，且维生素A主要通过典型信号通路调控牦牛卵母细胞的成熟。

关键词: 维生素A; 卵母细胞; 孤雌激活; 典型信号通路

Abstract: 【Objective】 The aim of this study was to explore the effects of vitamin A on in vitro maturation and subsequent embryo development of yak oocytes.【Method】 Yak oocytes were taken as the research object, and vitamin A of 0 (control group), 2, 5, 10 and 20 μmol/L was added to the in vitro maturation culture medium, and the first polar body excretion rate was counted after in vitro culture for 24 h.Parthenogenetic activation was carried out on matured oocytes in each group.After activation, the cleavage rate and blastocyst rate were counted at the 2nd and 8th days, respectively.The relative expression of node genes, including RARα, RARβ, RARγ, RXRα, RXRβ, RXRγ and STRA8 genes in typical signaling pathways and MEK and ERK1 genes in atypical signaling pathways were detected by Real-time quantitative PCR in MⅡ oocyte stage, and the optimal treatment concentration of vitamin A was selected.The optimal concentration of vitamin A was added to the in vitro maturation medium, and oocytes in MⅠ and MⅡ stage were collected at 6 and 24 hours of maturation.Some MⅡ oocytes were parthenogeneously activated, and blastocysts were collected at 8 days after activation.The relative expression of RARα, RXRα and STRA8 genes in GV, MⅠ and MⅡ oocytes and parthenogeneously activated blastocysts were detected by real-time fluorescence quantitative PCR.【Result】 Compared with control group, the first polar body excretion rate and cleavage rate of 2, 5 and 10 μmol/L vitamin A groups were significantly increased (P<0.05), and reached the highest in 2 μmol/L vitamin A group.The blastocyst rate of 2 μmol/L vitamin A group was increased significantly (P<0.05), while the first polar body excretion rate, cleavage rate and blastocyst rate of 20 μmol/L vitamin A group were decreased significantly (P<0.05).The results of Real-time quantitative PCR showed that compared with control group, the relative expressions of RARα, RXRα and STRA8 genes in 2, 5, 10 and 20 μmol/L vitamin A groups were significantly increased (P<0.05), among which the 2 μmol/L vitamin A group was the highest, so 2 μmol/L vitamin A had the best effect on the in vitro maturation of yak oocytes.Compared with GV stage oocytes, the relative expression of STRA8, RXRα and RARα genes in MⅡ stage oocytes were extremely significantly increased (P<0.01), but there was no significant difference between MⅠ stage oocytes and blastocyst (P>0.05).【Conclusion】 The addition of 2 μmol/L vitamin A could promote the maturation of yak oocytes in vitro and significantly increase the cleavage rate of the parthenogeneously activated blastocysts, and vitamin A regulated the maturation of yak oocytes mainly through typical signaling pathways.

Key words: vitamin A; oocytes; parthenogenetic activation; typical signal pathway

中图分类号:

S814.6

吉文汇, 王玉玲, 何翃闳, 付伟, 兰道亮. 维生素A对牦牛卵母细胞体外成熟及后续胚胎发育能力的影响[J]. 中国畜牧兽医, 2022, 49(12): 4707-4714.

JI Wenhui, WANG Yuling, HE Honghong, FU Wei, LAN Daoliang. Effects of Vitamin A on the Maturation and Subsequent Development of Yak Oocytes in vitro[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(12): 4707-4714.

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