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成年小鼠受伤骨骼肌再生相关巨噬细胞体内谱系追踪方案。

Protocol for in vivo lineage tracing of regeneration-associated macrophages from injured skeletal muscle of adult mice.

作者信息

Sousa Neuza S, Sousa-Victor Pedro, Neves Joana

机构信息

GIMM- Gulbenkian Institute for Molecular Medicine, 1649-035 Lisbon, Portugal; Faculdade de Medicina, Universidade de Lisboa, 1649-028 Lisbon, Portugal.

出版信息

STAR Protoc. 2025 Jun 20;6(2):103844. doi: 10.1016/j.xpro.2025.103844. Epub 2025 May 24.

DOI:10.1016/j.xpro.2025.103844
PMID:40413749
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC12159898/
Abstract

Macrophages undergo phenotypic transitions that are essential for successful skeletal muscle (SkM) regeneration. Here, we present a protocol for in vivo lineage tracing of regeneration-associated macrophages, combining genetic labeling with transplantation of fluorescence-activated cell sorting (FACS)-isolated cells. Macrophages isolated from congenic CD45.1 donor mice are transplanted into pre-injured SkMs of CD45.2 mice and phenotyped by flow cytometry at designated time points of the regenerative process. We describe steps for muscle injury, SkM tissue processing, macrophage isolation, transplantation, and flow cytometry phenotyping. For complete details on the use and execution of this protocol, please refer to Sousa et al..

摘要

巨噬细胞会经历表型转变，这对于骨骼肌（SkM）的成功再生至关重要。在此，我们展示了一种用于再生相关巨噬细胞体内谱系追踪的方案，该方案将基因标记与荧光激活细胞分选（FACS）分离细胞的移植相结合。从同基因CD45.1供体小鼠分离的巨噬细胞被移植到CD45.2小鼠预先损伤的SkM中，并在再生过程的指定时间点通过流式细胞术进行表型分析。我们描述了肌肉损伤、SkM组织处理、巨噬细胞分离、移植和流式细胞术表型分析的步骤。有关此方案使用和执行的完整详细信息，请参考苏萨等人的研究。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/2556d41ad18f/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/fc5b9f51f7d9/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/60a589ac00d5/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/2cc628f6af60/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/d1984fd8c1bd/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/7fe84ca26296/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/2556d41ad18f/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/fc5b9f51f7d9/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/60a589ac00d5/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/2cc628f6af60/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/d1984fd8c1bd/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/7fe84ca26296/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/85c9/12159898/2556d41ad18f/gr5.jpg

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本文引用的文献

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The immune landscape of murine skeletal muscle regeneration and aging.鼠类骨骼肌再生和衰老的免疫图谱。
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Single-cell and spatial transcriptomics identify a macrophage population associated with skeletal muscle fibrosis.单细胞和空间转录组学鉴定出与骨骼肌纤维化相关的巨噬细胞群体。
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A growth factor-expressing macrophage subpopulation orchestrates regenerative inflammation via GDF-15.表达生长因子的巨噬细胞亚群通过 GDF-15 调控再生炎症。
J Exp Med. 2022 Jan 3;219(1). doi: 10.1084/jem.20210420. Epub 2021 Nov 30.
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Systemic and Local Phenotypes of Barium Chloride Induced Skeletal Muscle Injury in Mice.氯化钡诱导的小鼠骨骼肌损伤的全身和局部表型
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Inflammatory monocytes recruited after skeletal muscle injury switch into antiinflammatory macrophages to support myogenesis.骨骼肌损伤后募集的炎性单核细胞会转变为抗炎性巨噬细胞以支持肌生成。
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