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宿主-病原体相互作用过程中人类 M1 和 M2 单核细胞来源的巨噬细胞中脂质介质分析和表型分析的方案。

Protocol for lipid mediator profiling and phenotyping of human M1- and M2-monocyte-derived macrophages during host-pathogen interactions.

机构信息

Department of Pharmaceutical/Medicinal Chemistry, Institute of Pharmacy, Friedrich Schiller University Jena, Philosophenweg 14, 07743 Jena, Germany.

Section of Experimental Virology, Institute of Medical Microbiology, Center for Molecular Biomedicine (CMB), Jena University Hospital, Hans-Knoell-Str. 2, 07745 Jena, Germany.

出版信息

STAR Protoc. 2024 Sep 20;5(3):103142. doi: 10.1016/j.xpro.2024.103142. Epub 2024 Sep 17.

DOI:10.1016/j.xpro.2024.103142
PMID:39305487
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11424942/
Abstract

Here, we present a protocol for primary, human immune cell isolation and stimulation for lipid mediator profiling. We describe steps for the isolation of monocytes from human leukocyte concentrates via density centrifugation and differentiation/polarization toward M1- or M2-monocyte-derived macrophages (MDMs). We detail stimulation approaches of MDMs with live bacteria or influenza A virus for lipid mediator profiling and sample preparation for subsequent analysis, such as enzyme expression, mRNA analysis, or surface marker determination. For complete details on the use and execution of this protocol, please refer to Jordan et al..

摘要

在这里，我们提供了一种用于初级人免疫细胞分离和刺激以进行脂质介质分析的方案。我们描述了通过密度离心从人白细胞浓缩物中分离单核细胞的步骤，并将其分化/极化为 M1 或 M2 单核细胞衍生的巨噬细胞（MDM）。我们详细介绍了用活细菌或甲型流感病毒刺激 MDM 以进行脂质介质分析以及为随后的分析（如酶表达、mRNA 分析或表面标志物测定）准备样品的方法。有关此方案的使用和执行的完整详细信息，请参阅 Jordan 等人的研究。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/11013978ade3/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/6fb42ca11ded/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/3696afb1ead7/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/fc9b4511770c/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/d1f8807a2555/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/11013978ade3/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/6fb42ca11ded/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/3696afb1ead7/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/fc9b4511770c/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/d1f8807a2555/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/4374/11424942/11013978ade3/gr4.jpg

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本文引用的文献

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iScience. 2023 Dec 26;27(1):108775. doi: 10.1016/j.isci.2023.108775. eCollection 2024 Jan 19.
2
Cannabidiol acts as molecular switch in innate immune cells to promote the biosynthesis of inflammation-resolving lipid mediators.大麻二酚在先天免疫细胞中充当分子开关，促进抗炎脂质介质的生物合成。
Cell Chem Biol. 2023 Dec 21;30(12):1508-1524.e7. doi: 10.1016/j.chembiol.2023.08.001. Epub 2023 Aug 29.
3
Targeting biosynthetic networks of the proinflammatory and proresolving lipid metabolome.
靶向促炎和促修复脂质代谢组的生物合成网络。
FASEB J. 2019 May;33(5):6140-6153. doi: 10.1096/fj.201802509R. Epub 2019 Feb 8.
4
Propagation and Titration of Influenza Viruses.流感病毒的传播与滴定
Methods Mol Biol. 2018;1836:59-88. doi: 10.1007/978-1-4939-8678-1_4.