阴道毛滴虫分泌蛋白对小鼠精子质量的影响

doi:10.12140/j.issn.1000-7423.2022.02.016

中国寄生虫学与寄生虫病杂志 ›› 2022, Vol. 40 ›› Issue (2): 236-241.doi: 10.12140/j.issn.1000-7423.2022.02.016

阴道毛滴虫分泌蛋白对小鼠精子质量的影响

罗澄扬¹^,²(), 郝利霞³, 郭丽华³, 王兵立¹, 纪思帆¹, 连安娜¹, 朱晓慧¹, 梅雪芳¹^,², 田晓薇¹^,², 王帅¹^,², 张振超¹^,²^,^*()

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

收稿日期:2021-07-30 修回日期:2021-11-19 出版日期:2022-04-30 发布日期:2022-04-13
通讯作者: 张振超
作者简介:罗澄扬（2000-）,男,本科生,从事男性滴虫感染与致病的研究。E-mail： 1835828386@qq.com
基金资助:
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);新乡市科技攻关计划项目(GG2021011)

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

LUO Cheng-yang¹^,²(), HAO Li-xia³, GUO Li-hua³, WANG Bing-li¹, JI Si-fan¹, LIAN An-na¹, ZHU Xiao-hui¹, MEI Xue-fang¹^,², TIAN Xiao-wei¹^,², WANG Shuai¹^,², ZHANG Zhen-chao¹^,²^,^*()

1. School of Basic Medicine, Xinxiang Medical College, Xinxiang 453003, China
2. Key Laboratory of Pathogen Biology, Xinxiang Medical College, Xinxiang 453003, China
3. Xinxiang Maternal and Child Health Hospital, Xinxiang 453003, China

Received:2021-07-30 Revised:2021-11-19 Online:2022-04-30 Published:2022-04-13
Contact: ZHANG Zhen-chao
Supported by:
National Natural Science Foundation of China(81802028);Science and Technology Project of Henan Province(212102310716);Innovation and entrepreneurship training program for College Students of Henan Province(S202110472024);Science and Technology Project of Xinxiang City(GG2019034);Science and Technology Project of Xinxiang City(GG2021011)

摘要/Abstract

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 × 10⁷/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。

关键词: 阴道毛滴虫, 分泌蛋白, 小鼠, 精子质量

Abstract:

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 × 10⁷/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 ± 1.30)%, (18.10 ± 1.10)%, (20.00 ± 1.40)%, which were significantly higher than that of the 0 ng/μl group (9.90 ± 1.90)% (F = 15.76, P < 0.01). After 2 h treatment, the mortality rates in the 10, 20, 40 and 80 ng/μl groups were (17.00 ± 1.20)%, (18.30 ± 1.20)%, (20.50 ± 1.20)% and (22.80 ± 1.50)%, respectively, which were significantly higher than that of the 0 ng/μl group (12.90 ± 0.50)% (F = 22.22, P < 0.01), and the mortality rate in the 5 ng/μl group (15.00 ± 0.40)% was not statistically different from that of the 0 ng/μl group. After co-culturing the sperm with TvESP at a final concentration of 40 ng/μl for 1 h, the integrity of acrosome was destroyed. All results suggested that TvESP can reduce sperm quality.

Key words: Trichomonas vaginalis, Secretory protein, Mouse, Sperm quality

中图分类号:

R383.211

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, 张振超. 阴道毛滴虫分泌蛋白对小鼠精子质量的影响[J]. 中国寄生虫学与寄生虫病杂志, 2022, 40(2): 236-241.

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. Affect of Trichomonas vaginalis excretory secretory protein on sperm quality of mice[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2022, 40(2): 236-241.

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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

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