Lv Hongming, Liu Qinmei, Zhou Junfeng, Tan Guangyun, Deng Xuming, Ci Xinxin
Institute of Translational Medicine, The First Hospital, Jilin University, Changchun 130001, China; Key Laboratory of Zoonosis, Ministry of Education, College of Veterinary Medicine. Jilin University, Changchun 130061, China.
Institute of Translational Medicine, The First Hospital, Jilin University, Changchun 130001, China.
Free Radic Biol Med. 2017 May;106:38-52. doi: 10.1016/j.freeradbiomed.2017.02.016. Epub 2017 Feb 7.
Daphnetin (Daph), a natural coumarin derivative isolated from plants of the Genus Daphne, possesses abundant biological activities, such as anti-inflammatory, antioxidant and anticancer properties. In the present study, we focused on investigating the protective effect of Daph against tert-butyl hydroperoxide (t-BHP)-induced oxidative damage, mitochondrial dysfunction and the involvement of underlying molecular mechanisms. Our findings indicated that Daph effectively inhibited t-BHP-stimulated cytotoxicity, cell apoptosis, and mitochondrial dysfunction, which are associated with suppressed reactive oxygen species (ROS) generation, decreased malondialdehyde (MDA) formation, increased superoxide dismutase (SOD) levels and glutathione (GSH)/GSSG (oxidized GSH) ratio. Further investigation indicated that Daph significantly suppressed cytochrome c release and NLRP3 inflammasome activation and modulated apoptosis-related protein Bcl-2, Bax, and caspase-3 expression. Moreover, Daph dramatically induced the expression of the glutamate-cysteine ligase modifier (GCLM) subunit and the glutamate-cysteine ligase catalytic (GCLC) subunit, heme oxygenase-1 (HO-1), and NAD (P) H: quinone oxidoreductase (NQO1), which is largely dependent on upregulating the nuclear factor-erythroid 2-related factor 2 (Nrf2) nuclear translocation, reducing the Keap1 protein expression, and strengthening the antioxidant response element (ARE) promoter activity. Additionally, Daph remarkably activated a c-Jun NH2-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) phosphorylation, but ERK and JNK inhibitor pretreatment exhibited an evident decrease of the level of Daph-enhanced Nrf2 nuclear translocation. Furthermore, Daph exposure suppressed t-BHP-induced cytotoxicity and ROS overproduction, which are mostly blocked in Nrf2 knockout RAW 264.7 cells and peritoneal macrophages. Accordingly, Daph exhibited protective roles against t-BHP-triggered oxidative damage and mitochondrial dysfunction by the upregulation of Nrf2 antioxidant signaling pathways, which may be involved in the activation of JNK and ERK.
瑞香素（Daph）是从瑞香属植物中分离得到的一种天然香豆素衍生物，具有多种生物活性，如抗炎、抗氧化和抗癌特性。在本研究中，我们重点研究了瑞香素对叔丁基过氧化氢（t-BHP）诱导的氧化损伤、线粒体功能障碍的保护作用以及潜在分子机制的参与情况。我们的研究结果表明，瑞香素能有效抑制t-BHP刺激的细胞毒性、细胞凋亡和线粒体功能障碍，这与活性氧（ROS）生成受抑制、丙二醛（MDA）形成减少、超氧化物歧化酶（SOD）水平升高以及谷胱甘肽（GSH）/氧化型谷胱甘肽（GSSG）比值增加有关。进一步研究表明，瑞香素能显著抑制细胞色素c释放和NLRP3炎性小体激活，并调节凋亡相关蛋白Bcl-2、Bax和半胱天冬酶-3的表达。此外，瑞香素能显著诱导谷氨酸-半胱氨酸连接酶调节亚基（GCLM）和谷氨酸-半胱氨酸连接酶催化亚基（GCLC）、血红素加氧酶-1（HO-1）以及NAD（P）H：醌氧化还原酶（NQO1）的表达，这在很大程度上依赖于上调核因子红系2相关因子2（Nrf2）的核转位、降低Keap1蛋白表达以及增强抗氧化反应元件（ARE）启动子活性。此外，瑞香素能显著激活c-Jun氨基末端激酶（JNK）和细胞外信号调节激酶（ERK）的磷酸化，但ERK和JNK抑制剂预处理后，瑞香素增强的Nrf2核转位水平明显降低。此外，瑞香素处理能抑制t-BHP诱导的细胞毒性和ROS过度产生，而这在Nrf2基因敲除的RAW 264.7细胞和腹腔巨噬细胞中大多被阻断。因此，瑞香素通过上调Nrf2抗氧化信号通路对t-BHP引发的氧化损伤和线粒体功能障碍发挥保护作用，这可能与JNK和ERK的激活有关。
