去泛素化酶 USP5 调节 RIPK1 驱动的细胞焦亡，以响应心肌缺血再灌注损伤。
Deubiquitinase USP5 regulates RIPK1 driven pyroptosis in response to myocardial ischemic reperfusion injury.
机构信息
Department of Cardiology, Henan Provincial People's Hospital, Zhengzhou University People's Hospital, No.7, Weiwu Road, Zhengzhou, 450000, Henan Province, China.
Department of Clinical Microbiology, Henan Provincial People's Hospital, Zhengzhou University People's Hospital, Zhengzhou, 450000, China.
出版信息
Cell Commun Signal. 2024 Sep 30;22(1):466. doi: 10.1186/s12964-024-01853-x.
BACKGROUND
Gasdermin D (GSDMD) mediated pyroptosis plays a significant role in the pathophysiology of myocardial ischemia/reperfusion (I/R) injury. However, the precise mechanisms regulating pyroptosis remain unclear. In the study, we aimed to investigate the underlying mechanism of pyroptosis in myocardial I/R injury.
METHODS
In the present study, we analyzed the effects of USP5 on the RIPK1 kinase activity mediated pyroptosis in vitro after H/R (hypoxia/reoxygenation) and in vivo in a MI/R mouse model. TTC and Evan's blue dye, Thioflavin S and immunohistochemistry staining were performed in wild-type, RIPK1 Cdh5-Cre and USP5 deficiency mice. CMEC cells were transfected with si-USP5. HEK293T cells were transfected with USP5 and RIPK1 overexpression plasmid or its mutants. The levels of USP5, RIPK1, Caspase-8, FADD and GSDMD were determined by Western blot. Protein interactions were evaluated by immunoprecipitation. The protein colocalization in cells was monitored using a confocal microscope.
RESULTS
In this study, our data demonstrate that RIPK1 is essential for limiting cardiac endothelial cell (CMEC) pyroptosis mediated by caspase-8 in response to myocardial I/R. Additionally, we investigate the role of ubiquitin-specific protease 5 (USP5) as a deubiquitinase for RIPK1. Mechanistically, USP5 interacts with RIPK1, leading to its deubiquitination and stabilization.
CONCLUSIONS
These findings offer new insights into the role of USP5 in regulating RIPK1-induced pyroptosis.
背景
Gasdermin D（GSDMD）介导的细胞焦亡在心肌缺血/再灌注（I/R）损伤的病理生理学中起着重要作用。然而，调节细胞焦亡的确切机制尚不清楚。在本研究中，我们旨在研究心肌 I/R 损伤中细胞焦亡的潜在机制。
方法
在本研究中，我们分析了 USP5 在体外 H/R（缺氧/复氧）后和体内 MI/R 小鼠模型中对 RIPK1 激酶活性介导的细胞焦亡的影响。在野生型、RIPK1 Cdh5-Cre 和 USP5 缺陷型小鼠中进行 TTC 和 Evans 蓝染料、硫黄素 S 和免疫组织化学染色。用 si-USP5 转染 CMEC 细胞。用 USP5 和 RIPK1 过表达质粒或其突变体转染 HEK293T 细胞。通过 Western blot 测定 USP5、RIPK1、Caspase-8、FADD 和 GSDMD 的水平。通过免疫沉淀评估蛋白相互作用。使用共聚焦显微镜监测细胞中蛋白质的共定位。
结果
在这项研究中，我们的数据表明 RIPK1 对于限制心肌 I/R 中 caspase-8 介导的心脏内皮细胞（CMEC）细胞焦亡是必不可少的。此外，我们研究了泛素特异性蛋白酶 5（USP5）作为 RIPK1 的去泛素化酶的作用。从机制上讲，USP5 与 RIPK1 相互作用，导致其去泛素化和稳定。
结论
这些发现为 USP5 在调节 RIPK1 诱导的细胞焦亡中的作用提供了新的见解。
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