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构建流感病毒感染小鼠中性粒细胞胞外诱捕网观察与评估体内模型的方案

Protocol for building an in vivo model for observation and evaluation of neutrophil extracellular trapping nets in influenza virus-infected mice.

作者信息

Dong Yingjie, Yang Qiuli, Wang Likun, Niu Ruiying, Xia Jingxuan, Bi Yujing, Liu Guangwei

机构信息

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, College of Life Sciences, Beijing Normal University, Beijing 100875, China.

Key Laboratory of Cell Proliferation and Regulation Biology, Ministry of Education, College of Life Sciences, Beijing Normal University, Beijing 100875, China.

出版信息

STAR Protoc. 2024 Dec 20;5(4):103436. doi: 10.1016/j.xpro.2024.103436. Epub 2024 Nov 4.

DOI:10.1016/j.xpro.2024.103436
PMID:39499615
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11568775/
Abstract

Neutrophils sense microbes and selectively release neutrophil extracellular traps (NETs) in response to pathogens. Here, we present a protocol for qualitative and quantitative analysis of NETs in PR8-infected mouse models. We describe steps for obtaining single-cell suspensions of infiltrated neutrophils from mouse bronchoalveolar lavage fluid (BALF). We then detail procedures to determine NET marker proteins using flow cytometry, confocal microscopes, and western blotting. This protocol may be applied to qualitative and quantitative analysis of NETs in influenza virus-infected mice. For complete details on the use and execution of this protocol, please refer to Zhang et al..

摘要

中性粒细胞能够感知微生物，并在病原体刺激下选择性地释放中性粒细胞胞外陷阱（NETs）。在此，我们展示了一种在PR8感染小鼠模型中对NETs进行定性和定量分析的方案。我们描述了从小鼠支气管肺泡灌洗液（BALF）中获取浸润中性粒细胞单细胞悬液的步骤。然后，我们详细说明了使用流式细胞术、共聚焦显微镜和蛋白质免疫印迹法来测定NET标记蛋白的程序。该方案可应用于流感病毒感染小鼠中NETs的定性和定量分析。有关本方案使用和实施的完整详细信息，请参考Zhang等人的研究。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/d68bea020a7a/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/20edcdf3b7de/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/cfe866a93ada/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/40251e37da80/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/f929202fc911/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/cdcd95bb7cde/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/d68bea020a7a/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/20edcdf3b7de/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/cfe866a93ada/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/40251e37da80/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/f929202fc911/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/cdcd95bb7cde/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/53ea/11568775/d68bea020a7a/gr5.jpg

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本文引用的文献

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Sirtuin 2 regulates neutrophil functions through NAD synthesis pathway in virus infection.沉默调节蛋白2在病毒感染中通过烟酰胺腺嘌呤二核苷酸合成途径调节中性粒细胞功能。
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Deacetylase sirtuin 2 negatively regulates myeloid-derived suppressor cell functions in allograft rejection.
去乙酰化酶 Sirtuin 2 负调控同种异体移植排斥中的髓源性抑制细胞功能。
Am J Transplant. 2023 Dec;23(12):1845-1857. doi: 10.1016/j.ajt.2023.08.017. Epub 2023 Aug 25.
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Protocol for influenza A virus infection of mice and viral load determination.甲型流感病毒感染小鼠的实验方案及病毒载量测定
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The Roles of Neutrophils in Cytokine Storms.中性粒细胞在细胞因子风暴中的作用。
Viruses. 2021 Nov 21;13(11):2318. doi: 10.3390/v13112318.
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Neutrophils and neutrophil extracellular traps in the liver and gastrointestinal system.中性粒细胞和中性粒细胞胞外诱捕网在肝脏和胃肠道系统中的作用。
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