Snyder Mark E, Sembrat John, Noda Kentaro, Myerburg Michael M, Craig Andrew, Mitash Nilay, Harano Takashi, Furukawa Masashi, Pilewski Joseph, McDyer John, Rojas Mauricio, Sanchez Pablo
Division of Pulmonary, Allergy, and Critical Care Medicine.
Department of Immunology.
Am J Respir Crit Care Med. 2021 May 15;203(10):1230-1244. doi: 10.1164/rccm.202006-2403OC.
Tissue-resident memory T cells (T) play a critical role in the defense against inhaled pathogens. The isolation and study of human lung tissue-resident memory T cells and lung-resident macrophages (M) are limited by experimental constraints. To characterize the spatial and functional relationship between M and human lung tissue-resident memory T cells using lung perfusion (EVLP). T and M were isolated using EVLP and intraperfusate-labeled CD45 antibody. Cells isolated after 6 hours of EVLP were analyzed using spectral flow cytometry. Spatial relationships between CD3 and CD68 cells were explored with multiplexed immunohistochemistry. Functional relationships were determined by using coculture and T-cell-receptor complex signal transduction. Lungs from 8 research-consenting organ donors underwent EVLP for 6 hours. We show that human lung T and M colocalize within the human lung, preferentially around the airways. Furthermore, we found that human lung CD8 T are composed of two functionally distinct populations on the basis of PD1 (programed cell death receptor 1) and ZNF683 (HOBIT) protein expression. We show that M provide costimulatory signaling to PD1 CD4 and CD8 lung T, augmenting the effector cytokine production and degranulation of T. EVLP provides an innovative technique to study resident immune populations in humans. Human M colocalize with and provide costimulation signaling to T, augmenting their effector function.
组织驻留记忆T细胞（T细胞）在抵御吸入性病原体方面发挥着关键作用。人类肺组织驻留记忆T细胞和肺驻留巨噬细胞（M细胞）的分离与研究受到实验限制。为了利用肺灌注（体外肺灌注，EVLP）来表征M细胞与人类肺组织驻留记忆T细胞之间的空间和功能关系。使用EVLP和灌注液标记的CD45抗体分离T细胞和M细胞。对EVLP 6小时后分离的细胞进行光谱流式细胞术分析。通过多重免疫组织化学探究CD3和CD68细胞之间的空间关系。通过共培养和T细胞受体复合物信号转导确定功能关系。来自8名同意参与研究的器官捐献者的肺进行了6小时的EVLP。我们发现人类肺T细胞和M细胞在人类肺内共定位，优先分布在气道周围。此外，我们发现基于程序性细胞死亡受体1（PD1）和锌指蛋白683（HOBIT）蛋白表达，人类肺CD8 T细胞由两个功能不同的群体组成。我们表明，M细胞为PD1阳性的CD4和CD8肺T细胞提供共刺激信号，增强T细胞的效应细胞因子产生和脱颗粒。EVLP提供了一种研究人类驻留免疫群体的创新技术。人类M细胞与T细胞共定位并为其提供共刺激信号，增强其效应功能。
