Biliverdin modulates the Nrf2/A20/eEF1A2 axis to alleviate cerebral ischemia-reperfusion injury by inhibiting pyroptosis
Biliverdin modulates the Nrf2/A20/eEF1A2 axis to alleviate cerebral ischemia-reperfusion injury by inhibiting pyroptosis
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DOI：
10.1016/j.biopha.2023.115057
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发表时间：
2023-07-01
影响因子：
7.5
通讯作者：
Shao, Jianlin
中科院分区：
文献类型：
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作者：
Bai, Wenya;Huo, Siying;Shao, Jianlin
本研究旨在探讨血红素的代谢产物胆绿素是否能通过抑制细胞凋亡减轻脑缺血再灌注损伤。在此，通过大脑中动脉闭塞-再灌注（MCAO/R）在C57 BL/6 J小鼠中诱导CIRI，并通过氧和葡萄糖剥夺/再给氧（OGD/R）在HT 22细胞中建立CIRI模型，其用或不用胆绿素处理。GSDMD-N的时空表达和梗死体积分别通过免疫荧光染色和氯化三苯基四氮唑（TTC）进行评估。通过蛋白质印迹法测定了NLRP 3/Caspase-1/GSDMD途径，其是焦亡过程的中心，以及Nrf 2、A20和eEF 1A 2的表达。Nrf 2，A20，和eEF 1A 2相互作用进行了验证，使用双荧光素酶报告分析，染色质免疫沉淀，或免疫共沉淀。此外，使用A20或eEF 1A 2基因干扰（过表达和/或沉默）研究了Nrf 2/A20/eEF 1A 2轴在调节胆绿素的神经保护特性中的作用。胆绿素40 mg/kg可显著减轻体内外CIRI，促进Nrf 2活化，提高A20表达，降低eEF 1A 2表达。Nrf 2可以与A20的启动子结合，从而转录调节A20的表达。A20还可以通过其ZnF 4结构域与eEF 1A 2相互作用，使其泛素化并降解，从而导致eEF 1A 2的下调。我们的研究还表明，A20的敲除或eEF 1A 2的过表达都会削弱胆绿素的保护作用。拯救实验进一步证实胆绿素可通过Nrf 2/A20/eEF 1A 2轴调节NF-κ B通路。总之，我们的研究表明胆绿素通过抑制经由Nrf 2/A20/eEF 1A 2轴的NF-κ B途径来改善CIRI。我们的研究结果可以帮助确定治疗CIRI的新的治疗靶点。
This study aimed to examine whether Biliverdin, which is a common metabolite of haem, can alleviate cerebral ischemia reperfusion injury (CIRI) by inhibiting pyroptosis. Here, CIRI was induced by middle cerebral artery occlusion-reperfusion (MCAO/R) in C57BL/6 J mice and modelled by oxygen and glucose deprivation/reoxygenation (OGD/R) in HT22 cells, it was treated with or without Biliverdin. The spatiotemporal expression of GSDMD-N and infarction volumes were assessed by immunofluorescence staining and triphenyltetrazolium chloride (TTC), respectively. The NLRP3/Caspase-1/GSDMD pathway, which is central to the pyroptosis process, as well as the expression of Nrf2, A20, and eEF1A2 were determined by Western-blots. Nrf2, A20, and eEF1A2 interactions were verified using dual-luciferase reporter assays, chromatin immunoprecipitation, or coimmunoprecipitation. Additionally, the role of Nrf2/A20/eEF1A2 axis in modulating the neuroprotective properties of Biliverdin was investigated using A20 or eEF1A2 gene interference (overexpression and/or silencing). 40 mg/kg of Biliverdin could significantly alleviate CIRI both in vivo and in vitro, promoted the activation of Nrf2, elevated A20 expression, but decreased eEF1A2 expression. Nrf2 can bind to the promoter of A20, thereby transcriptionally regulating the expression of A20. A20 can furthermore interacted with eEF1A2 through its ZnF4 domain to ubiquitinate and degrade it, leading to the downregulation of eEF1A2. Our studies have also demonstrated that either the knock-down of A20 or over-expression of eEF1A2 blunted the protective effect of Biliverdin. Rescue experiments further confirmed that Biliverdin could regulate the NF-KB pathway via the Nrf2/A20/eEF1A2 axis. In summary, our study demonstrates that Biliverdin ameliorates CIRI by inhibiting the NF-KB pathway via the Nrf2/A20/eEF1A2 axis. Our findings can help identify novel therapeutic targets for the treatment of CIRI.