摘要
目的:探讨枸杞多糖(LBP)对氧化应激(OS)模型小鼠睾丸生精功能损伤的保护作用及其机制。方法:利用线粒体内膜肽样-2突变(Immp2l^(-/-))小鼠作为独特的OS动物模型,以野生型Immp2l^(+/+)作为正常对照、Immp2l^(-/-)突变小鼠作为阴性对照,通过饮水喂养给予1.5月龄Immp2l^(-/-)小鼠20 mg/kg浓度LBP作为治疗干预组,3组小鼠于13月龄处死,采用HE染色检测睾丸组织病理变化情况,获取小鼠附睾精子计算其精子常规参数,荧光TUNEL法检测睾丸组织生精细胞凋亡情况,通过免疫组化和Western印迹检测谷胱甘肽过氧化物酶4(GPX4)和凋亡诱导因子(AIF)表达水平。结果:Immp2l^(-/-)小鼠睾丸组织皮质变薄,内见大量空泡样变性且生殖细胞减少,管腔内长形精子数量降低,LBP干预后睾丸组织空泡样病理变化减少,各级生精细胞排列较为整齐。与野生型Immp2l^(+/+)小鼠比较,Immp2l^(-/-)小鼠精子计数[(20.78±1.45)×10^(6)vs(72.89±8.28)×10^(6),P<0.01]、活率[(18.37±2.67)%vs(58.62±6.15)%,P<0.01]和正常形态精子百分率[(20.33±3.17)%vs(65.81±7.69),P<0.01]均显著降低,LBP干预后显著提高了Immp2l^(-/-)小鼠精子计数[(45.25±3.39)×10^(6),P<0.05]、活率[(36.34±4.56)%,P<0.05]和正常形态精子百分率[(38.72±3.63)%,P<0.05]。Immp2l^(-/-)小鼠睾丸组织生精细胞凋亡显著增加,与Immp2l^(+/+)相比具有显著性差异[(7.14±0.78)%vs(1.45±0.43)%,P<0.01],LBP干预Immp2l^(-/-)小鼠后凋亡率显著降低[(2.28±0.07)%,P<0.01]。免疫组化结果显示,LBP干预Immp2l^(-/-)小鼠后GPX4表达显著增加[(1.43±0.17)vs(2.75±0.48),P<0.05],AIF表达显著降低[(2.43±0.15)vs(1.35±0.51),P<0.05]。Western印迹结果显示,LBP可显著提高GPX4并降低AIF在Immp2l^(-/-)小鼠中的表达(P<0.01)。结论:20 mg/kg LBP可能通过GPX4-AIF通路改善Immp2l^(-/-)OS小鼠睾丸生精损伤。
Objective:To investigate the protective effect of Lycium barbarum polysaccharide(LBP)against testicular spermatogenic injury in mice with oxidative stress(OS)and its mechanism.Methods:A unique OS model was made in 1.5-month-old mice with mitochondrial inner membrane-like peptide-2 mutation(Immp2l^(-/-)),which were fed with water(the negative control group)or LBP in water at the concentration of 20 mg/kg(the LBP intervention group),and wild-type Immp2l^(+/+) mice used as normal controls and fed with water only.Then all the mice were sacrificed at 13 months old and the testis tissue harvested for observation of pathological changes by HE staining,measurement of routine semen parameters,and detection of the apoptosis of spermatogenic cells by TUNEL and the expression levels of glutathione peroxidase 4(GPX4)and apoptosis-inducing factor(AIF)by immunohistochemistry and Western blot.Results:Thinned testicular cortex was observed in the negative controls,with evident vacuolar degeneration and reduced numbers of germ cells and elongated spermatids in the lumen of the seminiferous tubules,but all these pathological changes were improved and the germ cells at different levels orderly arranged in the LBP intervention group.Compared with the normal controls,the mice in the negative control group showed dramatically reduced sperm count([72.89±8.28]vs[20.78±1.45]×10^(6),P<0.01)and the percentages of progressively motile sperm(PMS)([58.62±6.15]%vs[18.37±2.67]%,P<0.01)and morphologically normal sperm(MNS)([65.81±7.69]%vs[20.33±3.17]%,P<0.01)and increased apoptosis of spermatogenic cells([1.45±0.43]%vs[7.14±0.78]%,P<0.01).LBP intervention,however,significantly increased the sperm count([45.25±3.39]×10^(6),P<0.05),PMS([36.34±4.56]%,P<0.05)and MNS([38.72±3.63]%,P<0.05)and decreased the apoptosis of spermatogenic cells([2.28±0.07]%,P<0.01).The mice in the LBP intervention group,in comparison with the negative controls,exhibited remarkably up-regulated expression of GPX4(2.75±0.48 vs 1.43±0.17,P<0.05)and down-regulated expression of AIF(2.43±0.15 vs 1.35±0.51,P<0.05).Conclusion:Lycium barbarum polysaccharide at 20 mg/kg can reduce testicular spermatogenic injury in Immp2l^(-/-) mice with oxidative stress through GPX4 and AIF pathways.
作者
刘春莲
张倩
张少华
母春兰
陈耀平
焦海燕
徐仙
霍正浩
LIU Chun-lian;ZHANG Qian;ZHANG Shao-hua;MU Chun-lan;CHEN Yao-ping;JIAO Hai-yan;XU Xian;HUO Zheng-hao;无(Center of Reproductive Medicine,General Hospital of Ningxia Medical University,Yinchuan,Ningxia 750004,China;Key Laboratory of the Ministry of Education for Fertility Preservation and Maintenance,Yinchuan,Ningxia 750004,China;Department of Pathology,Yinchuan First Peopled Hospital,Yinchuan,Ningxia 750001,China;Department of Genetics and Cell Biology,Ningxia Medical University,Yinchuan,Ningxia 750004,China)
出处
《中华男科学杂志》
CAS
CSCD
北大核心
2021年第5期387-393,共7页
National Journal of Andrology
基金
国家自然科学基金(81660690,81960753)
宁夏回族自治区自然科学基金(2019AAC03188)。