Isolation and Identification of Interstitial Macrophages from the Lungs Using Different Digestion Enzymes and Staining Strategies.
Isolation and Identification of Interstitial Macrophages from the Lungs Using Different Digestion Enzymes and Staining Strategies.
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DOI：
10.1007/978-1-4939-7837-3_6
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发表时间：
2018
期刊：
影响因子：
--
通讯作者：
Jakubzick CV
中科院分区：
文献类型：
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作者：
Atif SM;Gibbings SL;Jakubzick CV
间质巨噬细胞（IM）存在于多个器官中。尽管对IM亚型发挥的独特功能作用的了解有限，但一般而言，巨噬细胞因其在稳态组织维持和炎症中的贡献而闻名，例如清除病原体和碎片以及分泌炎性介质和生长因子。IM亚型已在心脏、皮肤和肠道中被鉴定，最近我们在肺中鉴定了三种不同的IM。IM在其表面上表达高水平的MerTK、CD 64和CD 11b，在CD 11 c、CD 206和MHC II表达方面存在差异，并将三种肺IM亚型称为IM 1（CD 11 cloCD 206 +MHCIIlo）、IM 2（CD 11 cloCD 206 +MHCIIhi）和IM 3（CD 11 chiCD 206 loMHCIIhi）。在本章中，我们将重点介绍如何使用三种不同的消化酶：弹性蛋白酶、胶原酶D和Liberase TM从肺中提取IM。在这三种常用的酶中，Liberase TM在IM提取中是最有效的，特别是IM 3。此外，还检查了用于鉴定IM的替代染色策略，包括CD 64、MerTK、F4/80和Tim 4。因此，未来的研究强调IM亚型的功能作用将有助于我们进一步了解如何维持组织的稳态和炎症条件的诱导和解决。
Interstitial macrophages (IMs) are present in multiple organs. Although there is limited knowledge of the unique functional role IM subtypes play, macrophages, in general, are known for their contribution in homeostatic tissue maintenance and inflammation such as clearing pathogens and debris and secreting inflammatory mediators and growth factors. IM subtypes have been identified in the heart, skin, and gut, and more recently we identified three distinct IMs in the lung. IMs express on their surface high levels of MerTK, CD64, and CD11b, with differences in CD11c, CD206, and MHC II expression, and referred to the three pulmonary IM subtypes as IM1 (CD11cloCD206+MHCIIlo), IM2 (CD11cloCD206+MHCIIhi), and IM3 (CD11chiCD206loMHCIIhi). In this chapter, we highlight how to extract IMs from the lung using three different digestion enzymes: elastase, collagenase D, and Liberase TM. Of these three commonly used enzymes, Liberase TM was the most effective at IM extraction, particularly IM3. Furthermore, alternative staining strategies to identify IMs were examined, which included CD64, MerTK, F4/80, and Tim4. Thus, future studies highlighting the functional role of IM subtypes will help further our understanding of how tissue homeostasis is maintained and inflammatory conditions are induced and resolved.