摘要
旨在探索一种自然植物提取的抗氧化分子——β-谷甾醇(β-sitosterol,SITO)对猪卵母细胞体外成熟效率和早期胚胎体外发育质量的影响。本研究以在猪卵母细胞体外成熟培养基中添加不同浓度SITO为试验组,不添加SITO(含0.16%DMSO)的培养基为对照组,对屠宰场所得猪卵巢中收集到的卵母细胞进行体外培养,试验组浓度分别为10、20、40μmol·L~(-1),不同浓度各重复3次,每组培养卵母细胞150~200枚。培养46 h后,对卵母细胞成熟率进行统计,并收集成熟卵母细胞进行活性氧、细胞凋亡、线粒体膜电位染色以及相关基因的实时荧光定量PCR;对成熟卵母细胞进行孤雌激活,2 d后统计卵裂率,7 d后统计囊胚率,并统计囊胚细胞总数。结果发现,20μmol·L~(-1)SITO处理显著提高了猪卵母细胞的体外成熟率,且促进了孤雌激活后囊胚的形成并增加了囊胚细胞数,降低了卵母细胞中ROS含量,提高了抗氧化应激基因SOD和CAT的表达;同时,SITO降低卵母细胞的凋亡水平,增加抗凋亡基因BCL-2的表达,降低促凋亡基因Caspase 3和BAX的表达。此外,SITO可增强线粒体膜电位(ΔΨm),增加TFAM基因的表达。综上所述,本研究结果表明,在猪卵母细胞体外成熟培养基中添加20μmol·L^(-1)SITO能够显著促进猪卵母细胞的体外成熟率、提高胚胎的体外发育能力和胚胎质量。
The aim of this study was to investigate the effects ofβ-sitosterol(SITO),a natural plant-derived antioxidant molecule,on the in vitro maturation efficiency of porcine oocytes and the quality of early embryo development in vitro.In this study,SITO with different concentrations were added into porcine oocytes in vitro maturation medium,which was the experimental group,and the medium without SITO(containing 0.16%DMSO)was the control group,the concentrations in experimental groups were 10,20 and 40μmol·L-1 respectively,and they were used to culture the oocytes in vitro collected from porcine ovaries in slaughterhouse.Different concentrations were repeated 3 times,and 150-200 oocytes were cultured in each group.After 46 h of culture,measured the maturation rate of oocytes,and mature oocytes were collected for reactive oxygen species,apoptosis,mitochondrial membrane potential staining and qRT-PCR of related genes;the adult oocytes were activated by parthenogenesis,the cleavage rate was calculated after 2 days,the blastocyst rate was calculated after 7 days,and the total number of blastocysts was calculated.The results showed that 20μmol·L^(-1) SITO treatment significantly improved the maturation rate of porcine oocytes in vitro,promoted the formation of blastocysts after parthenogenetic activation,decreased the ROS in oocytes,and increased the expression of anti-oxidative stress genes SOD and CAT.At the same time,SITO decreased the apoptosis level of oocytes,increased the expression of anti-apoptotic gene BCL-2,and decreased the expression of pro-apoptotic genes Caspase 3 and BAX.In addition,SITO could enhance mitochondrial membrane potential(ΔΨm)and increase the expression of TFAM gene.In summary,the results of this study showed that adding 20μmol·L-1 SITO to porcine oocyte in vitro maturation medium can significantly promote the rate of porcine oocytes,and improve the in vitro development ability and embryo quality.
作者
蓝昕蕊
赵宝宝
张碧菡
林晓语
马会明
王勇胜
LAN Xinrui;ZHAO Baobao;ZHANG Bihan;LIN Xiaoyu;MA Huiming;WANG Yongsheng(College of Veterinary Medicine,Northwest Agriculture and Forestry University,Yangling 712100,China;Key Laboratory of Fertility Maintenance of Ministry of Education,Ningxia Medical University,Yinchuan 750004,China)
出处
《畜牧兽医学报》
CAS
CSCD
北大核心
2024年第4期1629-1637,共9页
ACTA VETERINARIA ET ZOOTECHNICA SINICA
基金
国家重点研发计划(2021YFD1200303)
陕西省重点研发计划(2022NY-048)。
