氧化应激在TOCP诱导小鼠睾丸间质细胞自噬中的作用及机制研究

Leydig cells play a crucial role in spermatogenesis. Tri-ortho -cresyl phosphate (TOCP) has been widely used as plasticizers and flame-retardants in industry. We found that TOCP can disrupt the seminiferous epithelium in the mouse testis and inhibit spermatogenesis (Toxicol Lett, 2012), and induce autophagy of rat spermatogonial stem cells (Reproduction, 2015). Our preliminary results found that TOCP could induce autophagy and oxidative stress in the mice Leydig cells and increase lysophosphatidylcholine (LPC) content of the cells. Inasmuch as LPC can induce oxidative stress and cell damage. These results indicate that LPC might be involved in TOCP-induced oxidative stress, and subsequently induce autophagy of mice Leydig cells. Therefore, we will test these hypotheses: (1) we will investigate whether TOCP induces autophagy of mice Leydig cells, (2) we will assess the effect of TOCP on LPC content and oxidative stress in these cells and (3), we will determine whether TOCP induces oxidative stress via LPC, and finally induces autophagy of mice Leydig cells. The present study will provide a novel insight into the mechanism for TOCP- induced testis injury in male mice.

睾丸间质细胞（Leydig）在精子发生中起着非常重要的作用。三邻甲苯磷酸酯（TOCP）常作为增塑剂及阻燃剂等，在工业中使用非常广泛。我们的前期研究表明，TOCP可破坏小鼠的生精上皮并抑制精子发生（Toxicol Lett, 2012）；且可诱导大鼠精原干细胞自噬（Reproduction, 2015）。我们的初步结果发现，TOCP可诱导小鼠睾丸Leydig细胞自噬及氧化应激，并增加细胞内溶血磷脂酰胆碱（LPC）的含量。鉴于LPC可诱导细胞氧化应激及细胞损伤，我们推测LPC可能参与TOCP导致的细胞氧化应激，从而诱导细胞自噬。本课题将首先确定TOCP是否诱导Leydig细胞自噬，并明确TOCP对细胞内LPC含量及氧化应激的影响；然后在抑制细胞内LPC含量及氧化应激的情况下，最终确定TOCP可通过LPC诱导细胞氧化应激，从而诱导细胞自噬。为深入探讨TOCP的生殖毒性机理提供理论基础和实验依据。

三邻甲苯磷酸酯（TOCP）常作为增塑剂及阻燃剂等，在工业中使用非常广泛，可导致精子发生障碍。我们的研究发现，TOCP可诱导小鼠睾丸Leydig细胞自噬，并诱导细胞产生氧化应激，同时发现，氧化应激可抑制细胞活性并诱导小鼠睾丸Leydig细胞产生自噬，并明确TOCP可通过氧化应激诱导小鼠睾丸Leydig细胞产生自噬。TOCP可增加小鼠睾丸Leydig细胞内LPC含量，而且LPC可抑制小鼠睾丸Leydig细胞活性和诱导细胞凋亡。LPC还可诱导小鼠睾丸Leydig细胞产生氧化应激，而抑制氧化应激则可挽救LPC诱导的细胞凋亡，提示LPC可通过氧化应激诱导细胞凋亡。我们还发现LPC可抑制小鼠睾丸Leydig细胞自噬，而用雷帕霉素诱导细胞自噬后，可显著抑制LPC诱导的小鼠睾丸间质TM3细胞凋亡。通过本课题的研究，提示氧化应激很参与了TOCP诱导小鼠睾丸间质Leydig细胞自噬，TOCP虽可增加细胞内LPC的含量，但LPC却抑制小鼠睾丸间质Leydig细胞自噬。本课题的研究为深入探讨TOCP对男性生殖毒性机理提供理论基础和实验依据。