Department of Urology, Pediatric Research Institute, Chongqing Key Laboratory of Structural Birth Defect and Reconstruction, Ministry of Education Key Laboratory of Child Development and Disorders, National Clinical Research Center for Child Health and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China; Department of Urology, the Second Affiliated Hospital of Chongqing Medical University, Chongqing, China.
Department of Urology, Pediatric Research Institute, Chongqing Key Laboratory of Structural Birth Defect and Reconstruction, Ministry of Education Key Laboratory of Child Development and Disorders, National Clinical Research Center for Child Health and Disorders, Children's Hospital of Chongqing Medical University, Chongqing, China.
Toxicology. 2024 Dec;509:153956. doi: 10.1016/j.tox.2024.153956. Epub 2024 Sep 20.
As one of the most prevalent environmental endocrine disruptors, di-(2-ethylhexyl) phthalate (DEHP) is known for its significant developmental toxicity to the male reproductive system in humans and mice. Prepubertal exposure to DEHP has been shown to cause testicular damage, but the underlying mechanisms require further investigation. To investigate this effect, prepubertal mice were exposed to 100, 250 or 500 mg/kg body weight (bw) of DEHP for 14 days, which resulted in impaired histological structure and increased apoptosis of the testes. RNA sequencing (RNA-seq) of testicular tissue suggested that DEHP led to injury in Leydig and Sertoli cells. To further elucidate these mechanisms, we conducted experiments using immature mouse Leydig (TM3) and Sertoli (TM4) cells, and exposed them to 200 μM mono-(2-ethylhexyl) phthalate (MEHP), the primary metabolite of DEHP, for 24 h. We found that MEHP exposure induced oxidative stress injury and promoted cell apoptosis, and that cotreatment with N-acetylcysteine partially reversed these injuries. Given the close association between oxidative stress and mitochondrial calcium levels, we demonstrated that MEHP exposure disrupted mitochondria and increased mitochondrial calcium levels. In addition, MEHP exposure facilitated the formation of mitochondria-associated endoplasmic reticulum membranes (MAMs), upregulated protein expression and enhanced the interactions of the IP3R3-Grp75-VDAC1 complex. Furthermore, inhibition of calcium transfer in the IP3R3-Grp75-VDAC1-MCU axis relieved MEHP-induced mitochondrial injury, oxidative stress and apoptosis in TM3 and TM4 cells. This study highlights the importance of MAM-mediated mitochondrial calcium overload and the subsequent apoptosis of Leydig and Sertoli cells as pivotal factors contributing to testicular injury induced by prepubertal exposure to DEHP.
作为最常见的环境内分泌干扰物之一，邻苯二甲酸二（2-乙基己基）酯（DEHP）已知对人类和小鼠的雄性生殖系统具有显著的发育毒性。青春期前暴露于 DEHP 已被证明会导致睾丸损伤，但潜在机制仍需进一步研究。为了研究这种影响，将青春期前的小鼠暴露于 100、250 或 500mg/kg 体重（bw）的 DEHP 中 14 天，导致睾丸组织的结构受损和细胞凋亡增加。睾丸组织的 RNA 测序（RNA-seq）表明，DEHP 导致 Leydig 和 Sertoli 细胞损伤。为了进一步阐明这些机制，我们使用未成熟的小鼠 Leydig（TM3）和 Sertoli（TM4）细胞进行了实验，并将它们暴露于 200μM 的邻苯二甲酸单（2-乙基己基）酯（MEHP），即 DEHP 的主要代谢物中 24 小时。我们发现，MEHP 暴露诱导氧化应激损伤并促进细胞凋亡，而用 N-乙酰半胱氨酸共处理可部分逆转这些损伤。鉴于氧化应激与线粒体钙水平之间的密切关系，我们证明 MEHP 暴露破坏了线粒体并增加了线粒体钙水平。此外，MEHP 暴露促进了线粒体相关内质网膜（MAMs）的形成，上调了蛋白质表达并增强了 IP3R3-Grp75-VDAC1 复合物的相互作用。此外，抑制 IP3R3-Grp75-VDAC1-MCU 轴中的钙转移缓解了 MEHP 诱导的 TM3 和 TM4 细胞中线粒体损伤、氧化应激和凋亡。这项研究强调了 MAM 介导的线粒体钙超载以及随后的 Leydig 和 Sertoli 细胞凋亡作为青春期前暴露于 DEHP 导致睾丸损伤的关键因素的重要性。
