抑制泛素特异性蛋白酶 7 可通过对比氧化应激改善铁蛋白沉积介导的心肌梗死:体外和体内分析

IF 4.4 2区 生物学 Q2 CELL BIOLOGY Cellular signalling Pub Date : 2024-09-18 DOI:10.1016/j.cellsig.2024.111423
Dong Yang , Tiling Zhang , Hai Qu, Shaolong Li, Jing Lu, Wanyan Cao, Zhipeng Chen, Han Zhang, Jing Yang, Jin Wang
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引用次数: 0

摘要

背景我们之前的研究确定,USP7 会加重心肌损伤。此外,现有研究表明 USP7 与铁蛋白沉积之间存在密切联系。然而,USP7 对铁蛋白沉积介导的心肌梗塞(MI)的影响仍不清楚。在这项研究中,我们建立了心肌梗死模型和慢病毒转染组,以抑制 USP7 在体内和体外的表达。用超声心动图检测心功能。TTC和HE染色用于评估心肌改变。通过 RT-qPCR 和 Western 印迹分析铁变态反应标志物(PTGS2、ACSL4、GPX4)的表达。流式细胞术和酶联免疫吸附法用于测量 Fe2+、脂质 ROS、GSH 和 GSSG 的水平。TEM 和普鲁士蓝染色用于观察线粒体的改变和铁沉积。采用 RT-qPCR、Western 印迹和免疫荧光技术分析 Keap1、Nrf2 和核 Nrf2 在体外和体内的表达情况。相反,在 USP7 抑制组,Keap1-Nrf2 信号通路的激活改善了铁蛋白沉积介导的 MI 结果。在体外,MI 模型表现出心肌细胞活力明显下降,线粒体损伤显著。然而,这些问题在 USP7 抑制组中得到了改善。在体内,USP7 加剧了 MI 模型组内的 MI 和铁沉积,导致 LVEF、LVFS、SV、LVAWd 和 LVPWs 值下降,而在 USP7 抑制组中,除了 LVPWd 和 LVPWs 没有明显变化外,其他值均有所改善。重要的是,体外和体内实验都显示了类似的结果:USP7 抑制后,Keap1 表达减少,Nrf2 和核 Nrf2 增加。此外,GPX4 的表达量减少,而 PTGS2 和 ACSL4 的表达量增加。值得注意的是,Fe2+、脂质 ROS、GSH 和 GSSG 的浓度明显降低。结论体外和体内研究发现,抑制 USP7 可减少铁沉积并抑制氧化应激,从而改善铁沉积诱导的 MI。
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Inhibition of ubiquitin-specific protease 7 ameliorates ferroptosis-mediated myocardial infarction by contrasting oxidative stress: An in vitro and in vivo analysis

Background

Our prior research determined that USP7 exacerbates myocardial injury. Additionally, existing studies indicate a strong connection between USP7 and ferroptosis. However, the influence of USP7 on ferroptosis-mediated myocardial infarction (MI) remains unclear. Given these findings, we are particularly interested in USP7's regulatory role in ferroptosis-mediated MI and its underlying mechanisms.

Methods

In this study, we established MI models and lentivirus-transfected groups to inhibit USP7 expression both in vivo and in vitro. Cardiac function was detected with Echocardiography. TTC and HE staining were employed to assess myocardial alterations. The expression of ferroptosis markers (PTGS2, ACSL4, GPX4) were analyzed by RT-qPCR and Western blotting. Flow cytometry and ELISA were used for measuring Fe2+, lipid ROS, GSH, and GSSG levels. TEM and Prussian blue staining were used to observe mitochondrial alterations and iron deposition. RT-qPCR, Western blotting, and immunofluorescence were conducted to analyze Keap1, Nrf2, and nuclear Nrf2 expression in vitro and in vivo.

Results

In the MI model group, USP7 expression significantly increased, worsening ferroptosis-mediated MI. Conversely, in the USP7-inhibited group, activation of the Keap1-Nrf2 signaling pathway improved ferroptosis-mediated MI outcomes. In vitro, the MI model exhibited a marked decline in cardiomyocyte viability and notable mitochondrial damage. However, these issues improved in the USP7-inhibited groups. In vivo, USP7 intensified MI and iron deposition within the MI model group, with decreased values of LVEF, LVFS, SV, LVAWd, and LVPWs, all of which showed improvement in the USP7-inhibited group, except for LVPWd and LVPWs, which showed no significant variation. Importantly, both the in vitro and in vivo experiments revealed analogous results: a reduction in Keap1 expression and an increase in both Nrf2 and nuclear Nrf2 post USP7 inhibition. Additionally, GPX4 expression decreased while PTGS2 and ACSL4 expressions increased. Notably, concentrations of Fe2+, lipid ROS, GSH, and GSSG significantly decreased.

Conclusion

In vitro and in vivo studies have found that inhibition of USP7 attenuates iron deposition and suppresses oxidative stress, resulting in amelioration of ferroptosis-induced MI.

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来源期刊
Cellular signalling
Cellular signalling 生物-细胞生物学
CiteScore
8.40
自引率
0.00%
发文量
250
审稿时长
27 days
期刊介绍: Cellular Signalling publishes original research describing fundamental and clinical findings on the mechanisms, actions and structural components of cellular signalling systems in vitro and in vivo. Cellular Signalling aims at full length research papers defining signalling systems ranging from microorganisms to cells, tissues and higher organisms.
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