Sun Ting, Zhang Yanmei, Zhong Shuping, Gao Fenfei, Chen Yicun, Wang Bin, Cai Wenfeng, Zhang Zhaojing, Li Weiqiu, Lu Shishi, Zheng Fuchun, Shi Ganggang
Department of Pharmacology, Shantou University Medical College, Shantou, China.
Department of Biochemistry and Molecular Biology, University of Southern California, Los Angeles, CA, United States.
Front Pharmacol. 2018 Jan 25;9:19. doi: 10.3389/fphar.2018.00019. eCollection 2018.
Early growth response-1 (Egr-1), a transcription factor which often underlies the molecular basis of myocardial ischemia/reperfusion (I/R) injury, and oxidative stress, is key to myocardial I/R injury. Silent information regulator of transcription 1(SIRT1) not only interacts with and is inhibited by Egr-1, but also downregulates reactive oxygen species (ROS) via the Forkhead box O1(FOXO1)/manganese superoxide dismutase (Mn-SOD) signaling pathway. -n-butyl haloperidol iodide (F), a new patented compound, protects the myocardium against myocardial I/R injury in various animal I/R models and various heart-derived cell hypoxia/reoxygenation (H/R) models . In addition, F can regulate the abnormal ROS/Egr-1 signaling pathway in cardiac microvascular endothelial cells (CMECs) and H9c2 cells after H/R. We studied whether there is an inverse Egr-1/ROS signaling pathway in H9c2 cells and whether the SIRT1/FOXO1/Mn-SOD signaling pathway mediates this. We verified a ROS/Egr-1 signaling loop in H9c2 cells during H/R and that F protects against myocardial H/R injury by affecting SIRT1-related signaling pathways. Knockdown of Egr-1, by siRNA interference, reduced ROS generation, and alleviated oxidative stress injury induced by H/R, as shown by upregulated mitochondrial membrane potential, increased glutathione peroxidase (GSH-px) and total SOD anti-oxidative enzyme activity, and downregulated MDA. Decreases in FOXO1 protein expression and Mn-SOD activity occurred after H/R, but could be blocked by Egr-1 siRNA. F treatment attenuated H/R-induced Egr-1 expression, ROS generation and other forms of oxidative stress injury such as MDA, and prevented H/R-induced decreases in FOXO1 and Mn-SOD activity Nuclear co-localization between Egr-1 and SIRT1 was increased by H/R and decreased by either Egr-1 siRNA or F. Therefore, our results suggest that Egr-1 inhibits the SIRT1/FOXO1/Mn-SOD antioxidant signaling pathway to increase ROS and perpetuate I/R injury. F inhibits induction of Egr-1 by H/R, thereby activating SIRT1/FOXO1/Mn-SOD antioxidant signaling and decreasing H/R-induced ROS, demonstrating an important mechanism by which F protects against myocardial H/R injury.
早期生长反应因子-1（Egr-1）是一种转录因子，常作为心肌缺血/再灌注（I/R）损伤和氧化应激的分子基础，是心肌I/R损伤的关键因素。沉默信息调节因子1（SIRT1）不仅与Egr-1相互作用并受其抑制，还通过叉头框O1（FOXO1）/锰超氧化物歧化酶（Mn-SOD）信号通路下调活性氧（ROS）。碘化N-丁基氟哌啶醇（F）是一种新的专利化合物，在各种动物I/R模型和各种心脏来源的细胞缺氧/复氧（H/R）模型中可保护心肌免受I/R损伤。此外，F可调节H/R后心脏微血管内皮细胞（CMECs）和H9c2细胞中异常的ROS/Egr-1信号通路。我们研究了H9c2细胞中是否存在Egr-1/ROS反向信号通路，以及SIRT1/FOXO1/Mn-SOD信号通路是否介导此过程。我们验证了H/R期间H9c2细胞中的ROS/Egr-1信号环路，且F通过影响SIRT1相关信号通路来保护心肌免受H/R损伤。通过小干扰RNA（siRNA）干扰敲低Egr-1可减少ROS生成，并减轻H/R诱导的氧化应激损伤，表现为线粒体膜电位上调、谷胱甘肽过氧化物酶（GSH-px）和总超氧化物歧化酶（SOD）抗氧化酶活性增加以及丙二醛（MDA）下调。H/R后FOXO1蛋白表达和Mn-SOD活性降低，但可被Egr-1 siRNA阻断。F处理可减轻H/R诱导Egr-1表达、ROS生成以及其他形式的氧化应激损伤如MDA，并防止H/R诱导的FOXO1和Mn-SOD活性降低。H/R增加了Egr-1与SIRT1之间的核共定位，而Egr-1 siRNA或F则降低了这种共定位。因此，我们的结果表明，Egr-1抑制SIRT1/FOXO1/Mn-SOD抗氧化信号通路以增加ROS并使I/R损伤持续存在。F抑制H/R诱导的Egr-1，从而激活SIRT1/FOXO1/Mn-SOD抗氧化信号并减少H/R诱导的ROS，这表明了F保护心肌免受H/R损伤的重要机制。
