环境内分泌干扰物通过抑制miR-133a负调控IP3R致尿道下裂的发生

As the starting of two-child policy, demands on promoting good prenatal and postnatal care is getting more pressing. The incidence of hypospadias has gradually increased, but there is still a lack of effective screening methods during pregnancy. The occurrence of hypospadias is considered to be closely related to the environmental endocrine disruptor (EEDs). Intrauterine EEDs exposure can lead to abnormal embryonic development through epigenetic modification. MiRNA is one of the common epigenetic modifications. In our previous research, we investigated the differential expression of miRNA and mRNA in genital tubercles (GT) of DBP induced hypospadiac male rat and normal male rat. The results suggest that the expression of miR-133a in hypospadias group is significantly decreased and the expression of target gene IP3R is significantly increased in hypospadias group. We speculate that DBP can influence the development of GT through its oxidative stress effect, which significantly increases the expression of IP3R, the concentration of calcium and the level of autophagy, and inhibits EMT in urethral epithelial cells, and block the fusion process of urethral groove, causing the occurrence of hypospadias. In this study, miR-133a was used as a breakthrough point to study the mechanism of hypospadias induced by IP3R through autophagy, and to explore the feasibility of miR-133a detection in peripheral blood and amniotic fluid of pregnant women for malformation screening.

二胎政策的启动对优生优育提出了更高要求。近年来尿道下裂发病率逐步上升，但临床仍然缺乏有效的孕期筛查方法。尿道下裂的发生被认为与环境内分泌干扰物（EEDs）密切相关。宫内EEDs暴露可通过表观遗传修饰致胚胎发育异常。miRNA是常见的表观遗传修饰之一。前期实验中，利用miRNA和mRNA芯片检测、综合分析正常子鼠和DBP孕期染毒致尿道下裂子鼠生殖结节（GT）中的差异表达。结果提示尿道下裂组中miR-133a表达明显下降，其靶基因IP3R的表达显著升高。IP3R对维持细胞内钙离子动态平衡起到至关重要的作用。实验发现尿道上皮细胞内钙离子浓度上升，诱发细胞自噬，使上皮间质转化（EMT）水平降低，导致尿道下裂的发生。本课题以miR-133a为切入点，研究其靶基因IP3R通过诱发自噬，致尿道下裂发生的机制，同时探索孕妇外周血和羊水中miR-133a检测用于畸形筛查的可行性。

近年来尿道下裂发病率逐步上升，但临床仍然缺乏有效的孕期筛查方法。尿道下裂的发生被认为与环境内分泌干扰物（EEDs）密切相关。项目组通过研究发现，尿道下裂子鼠GT中IP3R的表达明显上升，伴随着EMT减弱。在体外实验中发现，DBP处理后能导致尿道上皮细胞中IP3R的表达明显上调，EMT上皮相关标记表达增高，而间质相关标记的表达明显减弱，并伴有细胞内钙离子浓度的升高。DBP处理后能提高尿道上皮细胞内ROS水平，提示DBP对尿道上皮细胞所产生的影响与其氧化应激效应相关。研究结果提示妊娠期DBP可通过其氧化应激作用影响GT的正常发育，使得尿道上皮细胞内钙离子浓度明显升高，导致细胞EMT被抑制，使得尿道板的发育及尿道沟的融合过程受到影响，引起了尿道下裂的发生。.同时在研究中发现孕期DBP暴露除了对子代的泌尿生殖系统发育产生影响以外，还会造成母鼠空腹血糖水平的升高。为了进一步研究其中的具体机制，项目组展开相关研究。结果显示，无论是空腹血糖检测还是OGTT，STZ+DBP组母鼠血糖水平更高，而胰岛素分泌水平更低。孕期DBP暴露使得FoxM1被显著抑制，而pSTAT1则明显高表达。FoxM1可以由pSTAT1调节。DBP可通过其毒理学作用显著增加pSTAT1的表达，抑制其下游FoxM1的表达，从而导致胰岛β细胞活性下降、凋亡增加，最终导致妊娠期糖尿病的发生。.以往对尿道下裂的研究主要集中在生殖结节上。项目组同时发现DBP可以影响血管内皮的外分泌功能，从而干扰生殖结节的发育和诱导尿道下裂。结果表明，孕期DBP暴露可导致生殖结节血管内皮中RhoA/ROCK信号通路异常激活和ROS过量积聚。通过RhoA/ROCK/ROS途径促进血管内皮NAP-2分泌，并通过TGF-β途径进一步促进尿路上皮细胞EMT，最终促进后代大鼠尿道下裂的发生。本研究为研究尿道下裂的发生提供了新的方向，并有可能为今后尿道下裂的发生和畸形筛查提供一个预测指标。.项目研究成果在Ecotoxicol Environ Saf期刊发表论文2篇。

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