黄芩素通过激活正常人类肝细胞中的 Keap1/Nrf2 信号通路缓解砷诱导的氧化应激。
Baicalein Alleviates Arsenic-induced Oxidative Stress through Activation of the Keap1/Nrf2 Signalling Pathway in Normal Human Liver Cells.
机构信息
The Key Laboratory of Environmental Pollution Monitoring and Disease Control, Ministry of Education, School of Public Health, Guizhou Medical University, Guiyang, 550025, Guizhou, P.R. China.
出版信息
Curr Mol Med. 2024;24(3):355-365. doi: 10.2174/1566524023666230320163238.
BACKGROUND
Oxidative stress is a key mechanism underlying arsenicinduced liver injury, the Kelch-like epichlorohydrin-related protein 1 (Keap1)/nuclear factor E2 related factor 2 (Nrf2) pathway is the main regulatory pathway involved in antioxidant protein and phase II detoxification enzyme expression. The aim of the present study was to investigate the role and mechanism of baicalein in the alleviation of arsenic-induced oxidative stress in normal human liver cells.
METHODS
Normal human liver cells (MIHA cells) were treated with NaAsO (0, 5, 10, 20 μM) to observe the effect of different doses of NaAsO on MIHA cells. In addition, the cells were treated with DMSO (0.1%), NaAsO (20 μM), or a combination of NaAsO (20 μM) and Baicalein (25, 50 or 100 μM) for 24 h to observe the antagonistic effect of Baicalein on NaAsO. Cell viability was determined using a Cell Counting Kit- 8 (CCK-8 kit). The intervention doses of baicalein in subsequent experiments were determined to be 25, 50 and 100μM. The intracellular content of reactive oxygen species (ROS) was assessed using a 2',7'-dichlorodihydrofluorescein diacetate (DCFHDA) probe kit. The malonaldehyde (MDA), Cu-Zn superoxide dismutase (Cu-Zn SOD) and glutathione peroxidase (GSH-Px) activities were determined by a test kit. The expression levels of key genes and proteins were determined by real-time fluorescence quantitative polymerase chain reaction (qPCR) and Western blotting.
RESULTS
Baicalein upregulated the protein expression levels of phosphorylated Nrf2 (-Nrf2) and nuclear Nrf2, inhibited the downregulation of Nrf2 target genes induced by arsenic, and decreased the production of ROS and MDA. These results demonstrate that baicalein promotes Nrf2 nuclear translocation by upregulating p-Nrf2 and inhibiting the downregulation of Nrf2 target genes in arsenic-treated MIHA cells, thereby enhancing the antioxidant capacity of cells and reducing oxidative stress.
CONCLUSION
Baicalein alleviated arsenic-induced oxidative stress through activation of the Keap1/Nrf2 signalling pathway in normal human liver cells.
背景
氧化应激是砷诱导肝损伤的关键机制，Kelch 样环氧氯丙烷相关蛋白 1（Keap1）/核因子 E2 相关因子 2（Nrf2）途径是参与抗氧化蛋白和 II 相解毒酶表达的主要调节途径。本研究旨在探讨黄芩素在缓解正常人类肝细胞砷诱导氧化应激中的作用和机制。
方法
用 NaAsO（0、5、10、20 μM）处理正常人类肝细胞（MIHA 细胞），观察不同剂量 NaAsO 对 MIHA 细胞的影响。此外，用 DMSO（0.1%）、NaAsO（20 μM）或 NaAsO（20 μM）和黄芩素（25、50 或 100 μM）联合处理细胞 24 h，观察黄芩素对 NaAsO 的拮抗作用。用细胞计数试剂盒（CCK-8 试剂盒）测定细胞活力。后续实验中黄芩素的干预剂量确定为 25、50 和 100μM。用 2'，7'-二氯二氢荧光素二乙酸酯（DCFHDA）探针试剂盒评估细胞内活性氧（ROS）含量。用试剂盒测定丙二醛（MDA）、铜锌超氧化物歧化酶（Cu-Zn SOD）和谷胱甘肽过氧化物酶（GSH-Px）的活性。用实时荧光定量聚合酶链反应（qPCR）和 Western blot 测定关键基因和蛋白的表达水平。
结果
黄芩素上调了磷酸化 Nrf2（-Nrf2）和核 Nrf2 的蛋白表达水平，抑制了砷诱导的 Nrf2 靶基因的下调，并减少了 ROS 和 MDA 的产生。这些结果表明，黄芩素通过上调 p-Nrf2 并抑制砷处理的 MIHA 细胞中 Nrf2 靶基因的下调，促进 Nrf2 核易位，从而增强细胞的抗氧化能力并减轻氧化应激。
结论
黄芩素通过激活正常人类肝细胞中的 Keap1/Nrf2 信号通路缓解砷诱导的氧化应激。
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