Institute of Pharmacology and Toxicology, Zhejiang University, 866 Yuhangtang Road, Hangzhou 310058, China.
Undergraduate Students in Research Training Project at Zhejiang University, Hangzhou 310058, China.
Toxicol In Vitro. 2020 Dec;69:104988. doi: 10.1016/j.tiv.2020.104988. Epub 2020 Aug 28.
Perfluorooctane sulfonate (PFOS), a classic environmental pollutant, is reported to cause cardiotoxicity in animals and humans. It has been demonstrated that PFOS exposure down-regulates expression of cardiac-development related genes and proteins. However, the related mechanism of PFOS has not been fully elucidated. In the present study, the embryonic stem (ES) cells-derived cardiomyocytes (ESC-CMs) was employed to investigate PFOS-mediated mechanism in developmental toxicity of cardiomyocytes. Our previous study shows that PFOS induces cardiomyocyte toxicity via causing mitochondrial damage. Nevertheless, the underlying mechanism by which PFOS affects the autophagy-related mitochondrial toxicity in ESC-CMs remains unclear. Here, we found that PFOS induced the swelling of mitochondria and the autophagosome accumulation in ESC-CMs at 40 μM concentration. PFOS increased the levels of LC3-II, p62, and ubiquitinated proteins. PFOS also induced an increase of LC3 and p62 localization into mitochondria, indicating that mitophagy degradation was impaired. The results of autophagic flux using chloroquine and RFP-GFP-LC3 analysis showed that the accumulation of autophagosome was not caused by the formation but by the impaired degradation. PFOS was capable of blocking the fusion between autophagosome and lysosome. PFOS caused dysfunction of lysosomes because it down-regulated Lamp2a and cathepsin D, but it did not induced lysosome membrane permeabilization. Meanwhile, PFOS-mediated lysosomal function and the inhibitory effect of autophagic flux could be reversed by PP242 at 40 nM concentration, an mTOR inhibitor. Furthermore, PP242 restored PFOS-induced ATP depletion and mitochondrial membrane potential. In conclusion, PFOS induced mitochondrial dysfunction via blocking autophagy-lysosome degradation, leading to cardiomyocyte toxicity from ES cells.
全氟辛烷磺酸（PFOS）是一种经典的环境污染物，据报道可引起动物和人类的心脏毒性。已经证明，PFOS 暴露会下调与心脏发育相关的基因和蛋白质的表达。然而，PFOS 的相关机制尚未完全阐明。在本研究中，胚胎干细胞（ES）细胞衍生的心肌细胞（ESC-CMs）被用于研究 PFOS 介导的心肌细胞发育毒性的机制。我们之前的研究表明，PFOS 通过引起线粒体损伤诱导心肌细胞毒性。然而，PFOS 如何影响 ESC-CMs 中的自噬相关线粒体毒性的潜在机制尚不清楚。在这里，我们发现 40 μM 浓度的 PFOS 可诱导 ESC-CMs 中线粒体肿胀和自噬体积累。PFOS 增加了 LC3-II、p62 和泛素化蛋白的水平。PFOS 还诱导 LC3 和 p62 定位到线粒体中，表明线粒体自噬降解受损。使用氯喹和 RFP-GFP-LC3 分析进行自噬流的结果表明，自噬体的积累不是由于形成而是由于降解受损。PFOS 能够阻止自噬体与溶酶体融合。PFOS 引起溶酶体功能障碍，因为它下调了 Lamp2a 和组织蛋白酶 D，但它不会诱导溶酶体膜通透性。同时，PFOS 介导的溶酶体功能和自噬流的抑制作用可以在 40 nM 浓度的 mTOR 抑制剂 PP242 作用下逆转。此外，PP242 恢复了 PFOS 诱导的 ATP 耗竭和线粒体膜电位。总之，PFOS 通过阻断自噬溶酶体降解导致心肌细胞毒性，从而诱导线粒体功能障碍。
