阴道毛滴虫分泌蛋白对小鼠精子质量的影响  

Affect of Trichomonas vaginalis excretory secretory protein on sperm quality of mice

作　　者：罗澄扬 郝利霞 郭丽华 王兵立 纪思帆 连安娜 朱晓慧 梅雪芳 田晓薇 王帅[1,2] 张振超[1,2] LUO Cheng-yang;HAO Li-xia;GUO Li-hua;WANG Bing-li;JI Si-fan;LIAN An-na;ZHU Xiao-hui;MEI Xue-fang;TIAN Xiao-wei;WANG Shuai;ZHANG Zhen-chao(School of Basic Medicine,Xinxiang Medical College,Xinxiang 453003,China;Key Laboratory of Pathogen Biology,Xinxiang Medical College,Xinxiang 453003,China;Xinxiang Maternal and Child Health Hospital,Xinxiang 453003,China)

机构地区：[1]新乡医学院基础医学院,新乡453003 [2]新乡医学院新乡市病原生物学重点实验室,新乡453003 [3]新乡市妇幼保健院,新乡453003

出　　处：《中国寄生虫学与寄生虫病杂志》2022年第2期236-241,共6页Chinese Journal of Parasitology and Parasitic Diseases

基　　金：国家自然科学基金(81802028);河南省科技攻关计划项目(212102310716);河南省本科高校省级大学生创新创业训练项目(S202110472024);新乡市科技攻关计划项目(GG2019034,GG2021011)。

摘　　要：收集阴道毛滴虫分泌蛋白(TvESP),用蛋白浓度测定试剂盒测定TvESP浓度,并进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)鉴定。从昆明小鼠的附睾收集精子并调整精子密度为1×10^(7)/ml。将小鼠精子分为0、5、10、20、40、80 ng/μl浓度组(每组设3个平行孔),分别与终浓度为0、5、10、20、40、80 ng/μl的TvESP共培养1 h,倒置显微镜下观察精子活力,细胞流式检测精子凋亡情况;小鼠精子与TvESP作用0.5、1、2 h后,台盼蓝染液染色,计算精子死亡率;小鼠精子分别与终浓度为0和40 ng/μl的TvESP共培养1 h,荧光染料染色,荧光显微镜下观察精子顶体结构。组间比较采用单因素方差分析。结果显示,TvESP作用1 h后,5、10、20、40、80 ng/μl浓度组活力精子占比分别为(31.90±4.10)%、(23.00±4.90)%、(18.80±3.50)%、(13.60±1.20)%和(8.60±1.20)%,均低于0 ng/μl浓度组的(44.50±3.20)%(F=30.76,P<0.01);5、10、20、40、80 ng/μl浓度组精子的晚期凋亡率分别为(17.99±1.73)%、(22.43±1.53)%、(26.76±2.20)%、(32.05±1.68)%和(41.37±2.28)%,均高于0 ng/μl浓度组的(15.42±1.10)%(F=256.79,P<0.01)。TvESP作用0.5 h后,10、20、40和80 ng/μl浓度组精子的死亡率分别为(11.90±1.70)%、(14.10±1.40)%、(16.30±1.20)%、(17.90±1.30)%,均高于0 ng/μl浓度组的(7.80±1.10)%(F=15.72,P<0.05);TvESP作用1 h后,5、10、20、40、80 ng/μl浓度组精子的死亡率分别为(12.90±0.90)%、(14.60±0.90)%、(16.20±1.30)%、(18.10±1.10)%、(20.00±1.40)%,均高于0 ng/μl浓度组的(9.90±1.90)%(F=15.76,P<0.01);TvESP作用2 h后,10、20、40、80 ng/μl浓度组精子的死亡率分别为(17.00±1.20)%、(18.30±1.20)%、(20.50±1.20)%、(22.80±1.50)%,均高于0 ng/μl浓度组的(12.90±0.50)%(F=22.22,P<0.01)。TvESP作用1 h后,40 ng/μl浓度组精子的顶体轮廓不清、结构完整性被破坏。TvESP可降低精子质量。Trichomonas vaginalis excretory secretory protein(TvESP)was collected for determining the concentration using Bradford method kit and further analyzed of TvESP by thesodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE).Mouse sperm were collected from the epididymis of Kunming mice,and the sperm density was adjusted to 1×10^(7)/ml.Sperm were divided into 6 groups and co-cultured with TvESP at final concentrations of 0,5,10,20,40 and 80 ng/μl,respectively.After co-culturing with TvESP for 1 h,sperm motility was observed using an inverted microscope,and sperm apoptosis was detected by flow cytometry.After co-culturing for 0.5,1 and 2 h,the sperm viability was analyzed by trypan blue staining.After sperm were incubated with TvESP at final concentrations of 0 and 40 ng/μl for 1 h,sperm were stained with a fluorescent dye and the acrosome was observed under a fluorescence microscope.One-way ANOVA was used for comparing between groups.The results showed that the percentage of active sperm in the 5,10,20,40 and 80 ng/μl groups were(31.90±4.10)%,(23.00±4.90)%,(18.80±3.50)%,(13.60±1.20)%and(8.60±1.20)%,respectively,which were significantly lower than that of the 0 ng/μl concentration group(44.50±3.20)%(F=30.76,P<0.01).The apoptosis rates of sperm in the 5,10,20,40 and 80 ng/μl groups were(17.99±1.73)%,(22.43±1.53)%,(26.76±2.20)%,(32.05±1.68)%and(41.37±2.28)%,respectively,which were significantly higher than that of the 0 ng/μl group(15.42±1.10)%(F=256.79,P<0.01).After treatment with TvESP for 0.5 h,the mortality rates of sperm treated with 10 ng/μl(11.90±1.70)%,20 ng/μl(14.10±1.40)%,40 ng/μl(16.30±1.20)%,80 ng/μl(17.90±1.30)%of TvESP were significantly higher than that of the 0 ng/μl group(7.80±1.10)%(F=15.72,P<0.05).The sperm mortality rate in the 5 ng/μl group(10.10±1.40)%was not statistically different from that of the 0 ng/μl group.After treatment with TvESP for 1 h,the sperm mortality rates in the 5,10,20,40 and 80 ng/μl groups were(12.90±0.90)%,(14.60±0.90)%,(16.20±1

关 键 词：阴道毛滴虫 分泌蛋白 小鼠 精子质量 

分 类 号：R383.211[医药卫生—医学寄生虫学]