聚苯乙烯纳米塑料致雄性小鼠生殖毒性效应及初步机制

Reproductive toxicity and preliminary mechanism of polystyrene nanoplastics in male mice

目的分析聚苯乙烯纳米塑料(polystyrene nanoparticles,PS-NPs)亚慢性暴露对成年雄性小鼠生殖功能的影响,并探讨其影响睾酮(testosterone,T)合成的可能分子机制。方法60只8周龄C57BL/6雄性小鼠完全随机分成对照组(去离子水组)、PS-NPs低暴露组(10 mg/kg,10 PS组)和高暴露组(40 mg/kg,40 PS组),每组20只。每日灌胃1次,连续60 d。末次染毒24 h后,称量并计算各组小鼠睾丸、附睾、脑、肝、脾、肺、肾的质量和脏器系数;运用HE染色分析睾丸组织病理形态差异;透射电子显微镜观察睾丸细胞的超微结构改变;采用计算机辅助精液分析系统分析精液参数;采用酶联免疫吸附实验(ELISA)检测血清T、黄体生成素(luteinizing hormone,LH)和卵泡刺激素(follicle-stimulating hormone,FSH)含量;结合转录组测序数据分析结果,采用荧光定量PCR(qPCR)法检测各组小鼠睾丸组织中T合成关键基因的表达水平。结果与对照组相比,40 PS组小鼠附睾质量显著增加(P<0.05);10 PS组小鼠睾丸脏器系数和40 PS组小鼠附睾脏器系数均显著增加(P<0.05);10 PS和40 PS组小鼠肝和脾的质量以及脏器系数均显著下降(P<0.05)。病理学结果显示,与对照组相比,10 PS和40 PS组小鼠睾丸生精小管管腔增大,生精细胞松散,空泡化增多;透射电镜下可见10 PS和40 PS组睾丸间质细胞胞浆中线粒体肿胀和内质网扩张。10 PS组小鼠精子活力较对照组显著下降(P<0.05);10 PS和40 PS组小鼠血清T水平均降低,其中40 PS组下降显著(P<0.05);10 PS和40 PS组小鼠血清LH和FSH水平较对照组均显著上升(P<0.05)。10 PS和40 PS组小鼠睾丸中CYP11A1的mRNA表达较对照组均显著下调(P<0.05)。结论PS-NPs会导致小鼠睾丸病理和功能损伤,并可能通过影响睾丸间质细胞结构下调T生成关键基因CYP11A1的表达,降低T的合成。

Objective To analyze the effect of subchronic exposure of polystyrene nanoplastics(PSNPs)on reproductive function in adult male mice,and to explore the preliminary molecular mechanism of its impact on testosterone synthesis.Methods Sixty 8-week-old C57BL/6 male mice were fully randomly divided into control group(deionized water)and PS-NPs low(10 mg/kg,10 PS)and high exposure group(40 mg/kg,40 PS),with 20 mice in each group.The mice were treated once per day for 60 d by gavage.The weights and organ coefficient of the testes,epididymis,brain,liver,spleen,lung and kidney of each group of mice were weighed and calculated at 24 h after the last exposure.HE staining was used to analyze the histopathological differences of testicular tissue,and transmission electron microscopy was used to observe the ultrastructural changes of testicular cells.The computer assisted sperm assay system was used to analyze semen parameters,the contents of serum testosterone(T),luteinizing hormone(LH)and follicle-stimulating hormone(FSH)were detected by enzyme-linked immunosorbent assay(ELISA),and the expression levels of key genes for T synthesis,which had been screened by transcriptome sequencing,were determined in testicular tissues by real-time PCR.Results Compared with the control group,the weight of the epididymis in the 40 PS group was increased significantly(P<0.05),the testicular organ coefficients of mice in the 10 PS group and the epididymis organs coefficients of mice in the 40 PS group were increased obviously(P<0.05),and the weight and organ coefficients of the liver and spleen of the mice in the 10 PS and 40 PS groups were decreased remarkably(P<0.05).The pathological results showed that compared with the testis of the control group,the lumen of the semininogenic tubule in the 10 PS and 40 PS groups were enlarged,the semimatogenic cells were loose,and the vacuoleization was increased.Transmission electron microscopy displayed that mitochondrial swelling and endoplasmic reticulum expansion were observed in the cytoplasm of Leydig cells in 10 PS and 40 PS groups.The sperm motility of mice in the 10 PS group was decreased significantly compared with the control group(P<0.05).The serum T level of mice in the 10 PS and 40 PS groups was decreased,with that of the 40 PS group more significant(P<0.05).The serum LH and FSH levels of mice in the 10 PS and 40 PS groups were increased notably.Compared with the control group,the mRNA levels of CYP11A1 in the testes of mice were significantly down-regulated in both 10 PS and 40 PS groups(P<0.05).Conclusion PS-NPs can cause damage to the testes pathologically and functionally in mice,and may reduce T synthesis by inducing structural abnormalities of Leydig cells and down-regulating CYP11A1 expression.