育龄期男性暴露双酚A及替代物与生殖健康损害的关联及精子miRNA的介导作用

Bisphenol A and its alternatives are the ubiquitous endocrine disrupting chemicals present in the environment. Toxicological studies have consistently demonstrated that bisphenol A and its alternatives exposures cause adverse effects on male reproductive health. However, the evidence from epidemiological studies is limited and inconsistent. This project is conducted to examine the effects of exposure to bisphenol A and its alternatives on reproductive health among reproductive-aged men. The concentrations of bisphenol A and its alternatives in two urine samples and seminal plasma collected from each subject are measured to comprehensively and accurately assess the individuals’ exposure levels. The conventional semen quality parameters, spermatozoa apoptosis, sperm DNA damage and sperm protamine, as well as reproductive hormones are assessed as biomarkers of reproductive effects. The associations between bisphenol A and its alternatives exposures and biomarkers of reproductive effects are examined. Furthermore, the differences of sperm miRNA expression are analyzed in subjects using two-stage study, and the mediating role of sperm miRNA in the associations between exposure and outcomes are assessed. The results from this project will uncover the effects of exposure to bisphenol A and its alternatives on male reproductive health in agreement of exposures, epigenetic modification and reproductive effects, and also provide the early biomarkers and population-based scientific evidence for the reproductive risk assessment of bisphenol A and its alternatives exposure.

双酚A（BPA）及替代物是环境中一类广泛存在的内分泌干扰物，毒理学研究已表明其具有明确的雄性生殖毒性，但流行病学证据还较为缺乏，并且结论尚不一致。本项目拟在我国育龄期男性中开展BPA及替代物暴露对生殖健康影响的流行病学研究，通过测定研究对象双份尿样和精浆中BPA、BPF和BPS含量准确而全面评估个体暴露水平，以精液质量常规参数、精子凋亡、精子DNA损伤、精子鱼精蛋白表达和生殖激素为生殖效应指标，分析BPA及替代物暴露与生殖效应指标的关联。在此基础上，进一步采用两阶段研究策略分析研究对象精子miRNA差异表达谱，明确精子miRNA在BPA及替代物暴露与男性生殖健康损害关联中的介导作用。研究结果将在“暴露-表观遗传修饰-生殖效应”一致原则上深入认识BPA及替代物暴露对男性生殖健康的影响，也为BPA及替代物的生殖健康风险评估提供早期生物标志物和基于人群的科学依据。

双酚A（BPA）及替代物（BPF和BPS）作为环境中一类广泛存在的内分泌干扰物，毒理学研究已表明其具有降低精液质量、干扰内分泌系统等雄性生殖毒性，但流行病学证据还较为缺乏，并且其作用机制也有待阐明。本项目以来湖北省武汉市同济医院生殖中心寻求精液分析的984名育龄期男性为研究对象，通过测定研究对象双份尿样中BPA及替代物浓度准确评估个体环境暴露水平，以精液质量常规参数、精子凋亡、精子DNA损伤、精子鱼精蛋白和血清生殖激素为生殖效应指标，分析了BPA及替代物环境暴露与男性生殖损害之间的关系。在此基础上，采用Small RNA高通量测序技术对10名精液质量正常和异常的精子miRNA表达谱进行检测，根据精子miRNA表达量在两组间的差异，并结合生物信息学方法筛选出6种与精子发生相关的候选miRNA，采用荧光定量PCR仪测定了215名育龄期男性6种精子候选miRNA表达量，进一步采用中介分析探讨了候选精子miRNA在双酚A及替代物与精液质量降低关联中的介导作用。研究结果显示，BPA、BPF和BPS在研究人群的尿样中检出率均大于83%，表明我国育龄期男性人群已广泛暴露BPA及替代物；尿液中BPA和/或替代物（BPF和BPS）平均浓度升高与精液质量参数降低、精子凋亡、精子DNA损伤、精子鱼精蛋白表达改变以及生殖激素（雌二醇和性激素结合蛋白）改变有关，表明我国育龄期男性人群环境暴露BPA及替代物可能与生殖健康损害存在关联；初筛阶段共有60种精子miRNA表达在两组间存在差异（fold change>2），其中表达上调13种，表达下调47种；验证阶段选择了6种精子发生相关的候选miRNA（miRNA3613-3p、miRNA4455、miRNA4469、miRNA4480、miRNA4487、miRNA444804-3p），其中精子miRNA4487和miRNA4804-3p在BPA暴露与精子密度和精子总数降低关联中起着中介作用，中介比在16.43%-24.16%，表明BPA暴露可能通过诱导miRNA4487和miRNA4804-3p表达升高进而降低精液质量。研究成果为BPA及替代物的生殖健康损害的风险评估提供了新的基于中国人群的研究资料，同时也为深入认识BPA及替代物的生殖毒作用机制提供了新线索。