Trigger-happy resident memory CD4+ T cells inhabit the human lungs.

Resident memory T-cells (TRM) reside in the lung epithelium and mediate protective immunity against respiratory pathogens. While lung CD8+ TRM have been extensively characterized, the properties of CD4+ TRM remain unclear. Here we determined the transcriptional signature of CD4+ TRM, identified by the expression of CD103, retrieved from human lung resection material. Various tissue homing molecules were specifically upregulated on CD4+ TRM, while expression of tissue egress and lymph node homing molecules were low. CD103+ TRM expressed low levels of T-bet, only a small portion expressed Eomes, and while the mRNA levels for Hobit were increased, protein expression was absent. On the other hand, the CD103+ TRM showed a Notch signature. CD4+CD103+ TRM constitutively expressed high transcript levels of numerous cytotoxic mediators, which was functionally reflected by a fast recall response, magnitude of cytokine production, and a high degree of polyfunctionality. Interestingly, the superior cytokine production appears to be due to an accessible IFNγ locus and was partially due to rapid translation of preformed mRNA. Our studies provide a molecular understanding of the maintenance and potential function of CD4+ TRM in the human lung. Understanding the specific properties of CD4+ TRM is required to rationally improve vaccine design. Lung material was collected from non-cancerous lobectomy tissue of six patients with non-small cell lung carcinoma. Lung mononuclear cells where isolated after digestion of the partial or complete human lung resection material. Peripheral blood mononuclear cells were isolated from five healthy donors. Three populations were sorted from the lung, CD3+CD4+CD45RO+CD27-CD103+, CD3+CD4+CD45RO+CD27-CD103-, and CD3+CD4+CD45RO+CD27+CD103-. For the blood three populations were also sorted, CD3+CD4+CD45RA+CD45RO-CD27+ (memory), CD3+CD4+CD45RO+CD27+ (naive), CD3+CD4+CD45RO+CD27- (effector memory). RNA was isolated from the sorted cell samples. For the blood T cell naive, memory and effector memory subsets, RNA from five individually sorted samples was pooled. RNA from 24 samples (n=6 for the three lung T cell subsets; n=1 for the three blood T cell subsets, in duplicate) was hybridised on Illumina HumanHT-12 V4.0 microarrays. Four microarray samples were excluded after hybridization since their average signal was too low and one microarray was excluded after quality control using the arrayQualityMetrics R package.