活性氧对猪卵母细胞SUMO-1表达及其质量的影响  

Effects of ROS on Expression of SUMO-1 and Quality of Porcine Oocytes

作　　者：徐妲[1,2] 罗晓彤 陈璇[1] 吕艳秋 金一[1] XU Da;LUO Xiaotong;CHEN Xuan;LYU Yanqiu;JIN Yi(Agricultural College,Yanbian University,Yanji 133002,China;Jilin Academy of Agricultural Sciences,Jilin 132001,China)

机构地区：[1]延边大学农学院,延吉133002 [2]吉林市农业科学院,吉林132001

出　　处：《中国畜牧兽医》2020年第8期2544-2552,共9页China Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金(31960659);吉林省科技厅重点科技攻关项目(20190301037NY)。

摘　　要：本研究旨在调查活性氧过氧化氢(hydrogen peroxide,H2O2)对猪卵母细胞体外成熟(in vitro maturation,IVM)过程中蛋白质类泛素SUMO-1表达及精-卵结合能力的影响。试验分为0(control)、10、50、75和100μg/mL H2O2处理组。利用Western blotting、流式细胞术、实时荧光定量PCR、Hoechst染色等方法检测猪卵母细胞体外成熟、蛋白质类泛素SUMO-1含量、细胞活力、凋亡基因mRNA表达、透明带(zona pellucid,ZP)溶解度和精子-卵母细胞结合的表达。结果表明,75μg/mL H2O2组与对照组、10和50μg/mL H2O2组比较卵母细胞体外成熟率及细胞活力显著降低;75μg/mL H2O2组与其他H2O2组相比ZP溶解时间显著延长,并减少了精子黏附在成熟卵母细胞透明带上的数量(P<0.05)。在77和18 ku处出现SUMO-1蛋白标记,75μg/mL H2O2组与对照组、10和50μg/mL H2O2组比较SUMO-1蛋白含量显著降低(P<0.05)。75μg/mL H2O2与对照组、10和50μg/mL H2O2组比较显著下调了Bcl-2基因,而Caspase-3基因表达与对照组和10μg/mL H2O2组比较显著升高(P<0.05),50μg/mL H2O2与对照组和10μg/mL H2O2组相比显著上调了Bax基因水平(P<0.05)。综上所述,H2O2能调控猪卵母细胞体外成熟过程中类泛素化水平以及精-卵结合能力。The purpose of this study was to investigate the effects of reactive hydrogen peroxide(H2O2)on the expression of the protein ubiquitin SUMO-1 and the ability of sperm-oocyte binding of porcine oocyte during in vitro maturation(IVM).The test was divided into 5 groups:0(control),10,50,75 and 100μg/mL of H2O2 treatment group.Western blotting,flow cytometry,Real-time quantiative PCR,Hoechst staining and other methods were used to detect porcine oocyte in vitro maturation,content of protein SUMOylation SUMO-1,cell viability,expression of apoptotic gene mRNA,solubility of zona pellucida(ZP)and sperm-oocyte binding.The results showed that 75μg/mL H2O2 group compared with the control,10 and 50μg/mL H2O2 groups,the in vitro maturation rate and cell viability of oocytes decreased significantly,while 75μg/mL H2O2 group compared with other H2O2 groups,the ZP lysis time prolonged significantly,and the number of sperm adhering to the ZP of mature oocytes was reduced(P<0.05).SUMO-1 protein markers appeared at 77 and 18 ku,and the content of SUMO-1 protein was significantly reduced in 75μg/mL H2O2 group when compare with the control,10 and 50μg/mL H2O2 groups(P<0.05).75μg/mL H2O2 group compared with the control,10 and 50μg/mL H2O2 groups,Bcl-2 gene was significantly down-regulated(P<0.05),while 50μg/mL H2O2 significantly up-regulated Bax gene level when compared with the control and 10μg/mL H2O2 groups(P<0.05).In summary,H2O2 could regulate the SUMOylation level and sperm-oocyte binding ability of porcine oocytes during in vitro maturation.

关 键 词：活性氧 过氧化氢 卵母细胞 类泛素化 细胞活力 猪 

分 类 号：S828.9[农业科学—畜牧学]