Description

Exploration of Human Lung CD4⁺ T cell Heterogeneity

Anna E. Oja¹, Berber Piet², Martijn A. Nolte¹, René A.W. van Lier¹, Pleun Hombrink¹

¹Department of Hematopoiesis, Sanquin Research and Landsteiner Laboratory, Amsterdam, ²Department of Respiratory Medicine, OLVG, Amsterdam

T cells mediate adaptive immune responses. While circulating T cells protect against systemic infections, non-circulating resident memory T cells (T_RM) are critical for protection in barrier tissues such as the lungs, skin and intestine. While the mechanisms by which T_RM are recruited have become increasingly appreciated, major question remains regarding the heterogeneity and spatial organisation of these cells. While CD8⁺ T_RM are extensively investigated, the role of the more abundant CD4⁺ T_RM remains elusive. Single-cell analysis based on flow cytometry revealed three subsets of lung CD4⁺ T_RM: CD69⁺CD103⁺, CD69⁺CD103^- and CD69^-CD103^- cells. Using bulk-cell transcriptome analysis, we identified the transcriptional profile of human lung CD4⁺CD103⁺ T_RM and compared this to blood CD4⁺ T cell subsets. We demonstrated that lung CD4⁺CD103⁺ T_RM represent an unique subset of CD4⁺ T cells that is very different from circulating CD4⁺ T cells, but closely related to CD8⁺ T_RM. CD4⁺ T_RM expressed unique sets of chemokine receptors and adhesion molecules necessary for tissue homing. We revealed that resting CD4⁺CD103⁺T_RM constitutively express high mRNA levels of cytotoxic mediators, but no protein. Functionally this was reflected by a fast recall response, magnitude of cytokine production, and a high degree of polyfunctionality. Phenotypic validation of target genes revealed a previously unrecognized complexity of the CD4⁺ T_RM compartment in the lungs, as the expression of VLA-1, CXCR6 and the inhibitory 2B4 did not strictly associate to any of the conventional T_RM phenotypes. Future single-cell approaches may deal with this caveat of population-averaged measurements in conventional transcriptome analysis experiments. Better understanding of the molecular and functional characteristics of heterogeneous CD4⁺ T_RM populations may improve rational vaccine design and the efficacy of adoptive T cell therapy.