B[a]P致睾酮合成障碍中线粒体-内质网通路损害的作用及分子机制研究

Benzo[a]pyrene (B[a]P) is a widely existed environmental endocrine disruptor, which is harmful to male reproductive system. Previous studies have shown that B[a]P can damage Leydig cells and affect the process of testosterone synthesis, cause male reproductive damage, however, the target organelles and underlying molecular mechanisms involved are still unclear. Our previous study found that B[a]P can destroy the mitochondria and endoplasmic reticulum integrity of the mouse Leydig TM3 cell line, and the process of testosterone biosynthesis requires the participation of mitochondria and endoplasmic reticulum. From this, we speculated that mitochondria and endoplasmic reticulum may be important targets of testosterone synthesis disorder induced by B[a]P. In order to investigate the roles and molecular mechanisms of mitochondrial damage and endoplasmic reticulum stress in testosterone biosynthesis in Leyding cells exposed to B[a]P, the present study plans to establish in vitro model of BPDE-exposed the purified mouse Leydig cells and in vivo model of B[a]P-exposed mice, Testosterone concentration, mitochondrial ultrastructure, mitochondrial membrane potential, ATP level, mitochondrial DNA copy number, and expression of CHOP, PGC-1α signaling pathway related proteins, testosterone biosynthesis related proteins were measured through ELISA, transmission electronmicroscope, flow cytometry , ATP detection kit, Q-PCR and western blot, respectively. These findings will provide new ideas for further understanding the toxic mechanism of B[a]P-induced male reproductive damage.

苯并[a]芘（B[a]P）是一种广泛存在的环境内分泌干扰物，对雄性生殖系统具有危害作用。研究表明B[a]P可损伤睾丸间质细胞并影响睾酮合成过程造成雄性生殖损害，但其中所涉及的靶细胞器和具体的分子机制尚不清楚。我们前期研究发现B[a]P可诱导小鼠睾酮间质细胞株TM3线粒体和内质网损伤，且睾酮合成过程需要线粒体和内质网的参与，推测线粒体和内质网可能是B[a]P致睾酮合成障碍的重要损伤靶点。本项目拟建立B[a]P的活性代谢产物BPDE染毒原代小鼠睾酮间质细胞模型和B[a]P染毒动物模型，采用电镜、western blot、Q-PCR、流式细胞术、ELISA等方法检测睾酮水平、线粒体损伤、内质网应激和CHOP介导的PGC-1α通路关键蛋白的变化，旨在阐明线粒体损伤和内质网应激在B[a]P致睾丸间质细胞睾酮合成障碍中的作用及分子机制，为深入理解B[a]P致雄性生殖损伤的毒理机制提供新的思路。

苯并[a]芘（B[a]P）是一种广泛存在的环境内分泌干扰物，对雄性生殖系统具有危害作用。研究表明B[a]P可损伤睾丸间质细胞并影响睾酮合成过程造成雄性生殖损害，但其中所涉及的靶细胞器和具体的分子机制尚不清楚。前期研究发现B[a]P导致的睾酮合成障碍可能与影响线粒体有关，推测线粒体可能是B[a]P致睾酮合成障碍的重要损伤靶点。本项目通过构建B[a]P染毒动物模型和B[a]P的活性代谢产物BPDE染毒小鼠睾丸间质细胞模型，评价了B[a]P/BPDE对睾酮合成和线粒体的损伤效应；探讨了线粒体损伤在BPDE致睾酮合成障碍中的作用及机制；分析了线粒体复合体Ⅰ组装相关基因Ndufa10在BPDE诱导的线粒体损伤和睾酮合成障碍中的作用。研究结果表明：（1）证实了B[a]P暴露可诱导小鼠睾酮合成障碍及睾丸间质细胞线粒体损伤；（2）证实了睾丸间质细胞线粒体损伤是介导B[a]P抑制睾酮合成的重要靶点；（3）BPDE可能通过抑制Ndufa10基因的表达导致能量代谢紊乱，进而引起线粒体功能损害及睾丸间质细胞睾酮合成降低的重要机制；（4）初步发现线粒体损伤在1-硝基芘致小鼠睾酮合成障碍中发挥作用。我们的研究首次证明了睾丸间质细胞线粒体损伤是多环芳烃类污染物致类固醇合成紊乱中极其关键的靶点，为深入了解多环芳烃及其衍生物致雄性生殖毒性的作用机制提供了基础资料，对于阐明多环芳烃类污染物的健康危害、开展合理的危险性评估以及防护具有一定的现实意义。项目的各项研究工作按照计划进行，在各个阶段都取得了较好的进展和成果，达到了预期目标。目前，在本项目的资助下，项目组成员已发表SCI论著2篇，并有其它研究成果正在陆续整理发表中。