Chen Si, Zhang Zhuhong, Qing Tao, Ren Zhen, Yu Dianke, Couch Letha, Ning Baitang, Mei Nan, Shi Leming, Tolleson William H, Guo Lei
Division of Biochemical Toxicology, National Center for Toxicological Research (NCTR)/U.S. Food and Drug Administration (FDA), HFT-110, 3900 NCTR Road, Jefferson, AR, 72079, USA.
Division of Genetic and Molecular Toxicology, National Center for Toxicological Research/U.S. FDA, Jefferson, AR, 72079, USA.
Arch Toxicol. 2017 Mar;91(3):1293-1307. doi: 10.1007/s00204-016-1775-y. Epub 2016 Jul 1.
Many usnic acid-containing dietary supplements have been marketed as weight loss agents, although severe hepatotoxicity and acute liver failure have been associated with their overuse. Our previous mechanistic studies revealed that autophagy, disturbance of calcium homeostasis, and ER stress are involved in usnic acid-induced toxicity. In this study, we investigated the role of oxidative stress and the Nrf2 signaling pathway in usnic acid-induced toxicity in HepG2 cells. We found that a 24-h treatment with usnic acid caused DNA damage and S-phase cell cycle arrest in a concentration-dependent manner. Usnic acid also triggered oxidative stress as demonstrated by increased reactive oxygen species generation and glutathione depletion. Short-term treatment (6 h) with usnic acid significantly increased the protein level for Nrf2 (nuclear factor erythroid 2-related factor 2), promoted Nrf2 translocation to the nucleus, up-regulated antioxidant response element (ARE)-luciferase reporter activity, and induced the expression of Nrf2-regulated targets, including glutathione reductase, glutathione S-transferase, and NAD(P)H quinone oxidoreductase-1 (NQO1). Furthermore, knockdown of Nrf2 with shRNA potentiated usnic acid-induced DNA damage and cytotoxicity. Taken together, our results show that usnic acid causes cell cycle dysregulation, DNA damage, and oxidative stress and that the Nrf2 signaling pathway is activated in usnic acid-induced cytotoxicity.
许多含松萝酸的膳食补充剂已作为减肥产品推向市场，尽管过度使用它们会导致严重的肝毒性和急性肝衰竭。我们之前的机制研究表明，自噬、钙稳态紊乱和内质网应激与松萝酸诱导的毒性有关。在本研究中，我们调查了氧化应激和Nrf2信号通路在松萝酸诱导的HepG2细胞毒性中的作用。我们发现，用松萝酸处理24小时会导致DNA损伤和S期细胞周期阻滞，且呈浓度依赖性。松萝酸还引发了氧化应激，表现为活性氧生成增加和谷胱甘肽耗竭。用松萝酸短期处理（6小时）显著增加了Nrf2（核因子红细胞2相关因子2）的蛋白质水平，促进了Nrf2向细胞核的转位，上调了抗氧化反应元件（ARE）-荧光素酶报告基因活性，并诱导了Nrf2调节靶点的表达，包括谷胱甘肽还原酶、谷胱甘肽S-转移酶和NAD(P)H醌氧化还原酶-1（NQO1）。此外，用shRNA敲低Nrf2增强了松萝酸诱导的DNA损伤和细胞毒性。综上所述，我们的结果表明，松萝酸会导致细胞周期失调、DNA损伤和氧化应激，并且Nrf2信号通路在松萝酸诱导的细胞毒性中被激活。
