Yue Liang, Zhao Lei, Liu Haixiao, Li Xia, Wang Bodong, Guo Hao, Gao Li, Feng Dayun, Qu Yan
Department of Neurosurgery, Tangdu Hospital, Fourth Military Medical University, Xi'an, China.
Oxid Med Cell Longev. 2016;2016:2957354. doi: 10.1155/2016/2957354. Epub 2016 Nov 30.
Glutamate- (Glu-) induced excitotoxicity plays a critical role in stroke. This study aimed to investigate the effects of APN on Glu-induced injury in HT22 neurons. HT22 neurons were treated with Glu in the absence or the presence of an APN peptide. Cell viability was assessed using the MTT assay, while cell apoptosis was evaluated using TUNEL staining. Levels of LDH, MDA, SOD, and GSH-Px were detected using the respective kits, and ROS levels were detected using dichlorofluorescein diacetate. Western blot was used to detect the expression levels of silent information regulator 1 (SIRT1), peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1), cleaved caspase-3, Bax, and Bcl-2. In addition to the western blot, immunofluorescence was used to investigate the expression levels of SIRT1 and PGC-1. Our results suggest that APN peptide increased cell viability, SOD, and GSH-Px levels and decreased LDH release, ROS and MDA levels, and cell apoptosis. APN peptide upregulated the expression of SIRT1, PGC-1, and Bcl-2 and downregulated the expression of cleaved caspase-3 and Bax. Furthermore, the protective effects of the APN peptide were abolished by SIRT1 siRNA. Our findings suggest that APN peptide protects HT22 neurons against Glu-induced injury by inhibiting neuronal apoptosis and activating SIRT1-dependent PGC-1 signaling.
谷氨酸（Glu）诱导的兴奋性毒性在中风中起关键作用。本研究旨在探讨脂联素（APN）对Glu诱导的HT22神经元损伤的影响。HT22神经元在不存在或存在APN肽的情况下用Glu处理。使用MTT法评估细胞活力，同时使用TUNEL染色评估细胞凋亡。使用相应试剂盒检测乳酸脱氢酶（LDH）、丙二醛（MDA）、超氧化物歧化酶（SOD）和谷胱甘肽过氧化物酶（GSH-Px）水平，使用二氯荧光素二乙酸酯检测活性氧（ROS）水平。采用蛋白质免疫印迹法检测沉默信息调节因子1（SIRT1）、过氧化物酶体增殖物激活受体γ共激活因子1α（PGC-1）、裂解的半胱天冬酶-3、Bax和Bcl-2的表达水平。除蛋白质免疫印迹法外，还采用免疫荧光法研究SIRT1和PGC-1的表达水平。我们的结果表明，APN肽可提高细胞活力、SOD和GSH-Px水平，并降低LDH释放、ROS和MDA水平以及细胞凋亡。APN肽上调SIRT1、PGC-1和Bcl-2的表达，下调裂解的半胱天冬酶-3和Bax的表达。此外，SIRT1小干扰RNA消除了APN肽的保护作用。我们的研究结果表明，APN肽通过抑制神经元凋亡和激活SIRT1依赖性PGC-1信号通路来保护HT22神经元免受Glu诱导的损伤。
