α-Amanitin通过抑制过氧化物还氧蛋白6激活氧化应激和线粒体自噬,从而加重肝损伤。

IF 3.3 4区 医学 Q3 IMMUNOLOGY
Zhongfeng Cheng, Kerun Cheng, Yan Tang, Xueqiong Duan, Yangshan Fu, Hongdan Duan, Yong Ye
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引用次数: 0

摘要

蘑菇中毒主要由α-amanitin (α-AMA)引起,目前尚无治疗α-AMA中毒的有效药物。因此,寻找α-AMA损伤的早期诊断标志物显得尤为重要。建立了α-AMA诱导的肝细胞和小鼠肝损伤模型。采用3-(4,5-二甲基噻唑-2-基)-2,5-二苯基溴化四唑(MTT)法测定α-AMA处理后人正常肝细胞的细胞活力。采用酶联免疫吸附试验(ELISA)评估肝功能参数。2',7'-二氯荧光素双醋酸酯(DCFH-DA)和双氢乙啶(DHE)染色检测氧化应激。Western blot和免疫荧光染色检测细胞自噬和凋亡相关蛋白。采用苏木精和伊红(H&E)、末端脱氧核苷酸转移酶dUTP缺口末端标记(TUNEL)和油红O (ORO)染色观察细胞损伤程度和肝细胞凋亡情况。此外,采用JC-1免疫荧光染色和流式细胞术检测线粒体膜电位。结果表明,α-AMA呈剂量依赖性降低细胞活力。α- ama处理组的谷丙转氨酶(ALT)、天冬氨酸转氨酶(AST)和线粒体活性氧(mtROS)水平升高,抗氧化剂超氧化物歧化酶(SOD)水平降低。α-AMA可促进肝细胞自噬和凋亡,而PRDX6过表达可减轻这一作用。最后,发现PRDX6和Parkin在线粒体中蓄积,α-AMA通过沉默PRDX6激活线粒体自噬。综上所述,我们的研究结果表明α-AMA通过抑制PRDX6的表达激活氧化应激和线粒体自噬,导致肝损伤。这些来自体外和体内模型的研究结果为α-AMA的毒理学机制提供了新的见解,强调了PRDX6作为治疗α-AMA诱导的肝毒性的治疗靶点的潜力。重点:α-AMA导致ROS积累,激活氧化应激。α-AMA促进肝细胞自噬和凋亡。PRDX6可减轻α- ama诱导的肝损伤。PRDX6通过Parkin介导线粒体自噬。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
α-Amanitin aggravates hepatic injury by activating oxidative stress and mitophagy via peroxiredoxin 6 inhibition.

Mushroom poisoning is mainly caused by α-amanitin (α-AMA), and there is currently no effective drug to treat α-AMA poisoning. Therefore, it is particularly important to find early diagnostic markers for α-AMA injury. Hepatic injury models induced by α-AMA were established both in hepatic cells and mice. The cell viability of human normal hepatic cells after α-AMA treatment was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Liver function parameters was assessed by the Enzyme-Linked Immunosorbent Assay (ELISA). Furthermore, oxidative stress was detected by 2',7'-Dichlorofluorescin Diacetate (DCFH-DA) and Dihydroethidium (DHE) staining. Autophagy- and apoptosis-related proteins were assessed by Western blot and immunofluorescence staining. We applied Hematoxylin and Eosin (H&E), Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and Oil Red O (ORO) staining to observe the degree of cell damage and hepatocyte apoptosis. In addition, mitochondrial membrane potential was also determined by JC-1 immunofluorescence staining and flow cytometry. The results showed that α-AMA decreased cell viability in a dose-dependent manner. In addition, the levels of alanine aminotransferase (ALT), aspartate transaminase (AST) and mitochondrial reactive oxygen species (mtROS) were observed to increase in the α-AMA-treated groups, whereas antioxidants superoxide dismutase (SOD) levels were reduced. Moreover, α-AMA promoted hepatocyte mitophagy and apoptosis, which were alleviated by PRDX6 overexpression. Finally, PRDX6 and Parkin were found to accumulate in mitochondria and α-AMA activated mitophagy by silencing PRDX6. Collectively, our results demonstrated that α-AMA activates oxidative stress and mitophagy by inhibiting the expression of PRDX6, leading to hepatic injury. These findings from both in vitro and in vivo models provide insights into the toxicological mechanisms of α-AMA, underscoring the potential of PRDX6 as a therapeutic target for treating α-AMA-induced hepatotoxicity. HIGHLIGHTS: α-AMA leads to ROS accumulation and activates oxidative stress. α-AMA promotes hepatocyte mitophagy and apoptosis. PRDX6 alleviates α-AMA-induced hepatic injury. PRDX6 mediates mitophagy through Parkin.

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来源期刊
Immunologic Research
Immunologic Research 医学-免疫学
CiteScore
6.90
自引率
0.00%
发文量
83
审稿时长
6-12 weeks
期刊介绍: IMMUNOLOGIC RESEARCH represents a unique medium for the presentation, interpretation, and clarification of complex scientific data. Information is presented in the form of interpretive synthesis reviews, original research articles, symposia, editorials, and theoretical essays. The scope of coverage extends to cellular immunology, immunogenetics, molecular and structural immunology, immunoregulation and autoimmunity, immunopathology, tumor immunology, host defense and microbial immunity, including viral immunology, immunohematology, mucosal immunity, complement, transplantation immunology, clinical immunology, neuroimmunology, immunoendocrinology, immunotoxicology, translational immunology, and history of immunology.
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