Immunity
ArticleLymphatic migration of unconventional T cells promotes site-specific immunity in distinct lymph nodes
Graphical abstract
Introduction
Lymph nodes (LNs) are specialized organs that bring together various cell types, such as dendritic cells, macrophages, natural killer (NK) cells, and B and T lymphocytes, to effectively initiate adaptive immune responses against pathogens (Qi et al., 2014). A key element for the function of LNs is their connection to the lymphatic system allowing them to integrate information from the peripheral tissues that they drain (Roozendaal et al., 2008). In principle, this information in the form of metabolites, cytokines, chemokines, microbial products, or pathogens can passively drain or can be actively transported by migratory dendritic cells via the lymph (Girard et al., 2012; Thomas et al., 2016). Importantly, this information is not only used to initiate and scale an immune response but also to shape its quality, which is adapted to the tissue that is drained by the respective LN. As such, it has been shown that conventional dendritic cells (cDCs) that migrate from distinct tissues optimize the polarization and expression of adhesion molecules in the lymphocytes they interact with. Thereby, they adjust the function of T and B cells and promote their recruitment to the target tissue from which these cDCs emigrated (Campbell and Butcher, 2002; Everson et al., 1996; Johansson-Lindbom et al., 2003; Mora et al., 2005). A specific challenge is the orchestration of immune responses in the LNs, which drain barrier tissues, because the inflammatory responses against pathogens need to be balanced against tolerance toward commensal bacteria and, in the intestine, against food antigens. In the intestine, specific requirements on local tissue immunity are solved by compartmentalizing the lymphatic drainage. Here, the lymph transports the regional information to LNs that harbor adapted stromal and immigrated dendritic cell populations that in turn depend on the local tissue environment and microbial communities (Cording et al., 2014; Esterházy et al., 2019; Hammerschmidt et al., 2008; Houston et al., 2016; Pezoldt et al., 2018; Worbs et al., 2006). Together these data established the paradigm that dendritic cells not only report the tissue status to lymphocytes in the draining LNs (dLNs) but additionally generate and fine-tune immune responses that are optimized to the tissue that they drain. Whether and how other immune cells have functions in determining the tissue status and transport this information to dLNs to generate site-specific immune responses is currently not known.
Here, we asked whether unconventional T cells (UTCs) could have such a role in linking tissue immunity to lymph node function given their tissue-specific distribution patterns, their capacity to exhibit early innate cytokine production, and their ability to sense cellular stress (Godfrey et al., 2015). These features would, in principle, qualify them as cells that can relay tissue-specific information and translate them into characteristic and tissue-adapted immune responses.
UTCs consist of three major lineages—gamma delta (γδ), MR1-restricted (e.g., mucosal-associated invariant T cells [MAIT]), and CD1d-restricted T cells (e.g., natural killer T cells [NKT]), which all display a T cell receptor (TCR) repertoire of limited diversity. These cells are selected by non-polymorphic antigen-presenting molecules, such as CD1 and MR1, and typically recognize endogenous lipids or microbial metabolites bound to these surface proteins (Legoux et al., 2019; Pellicci et al., 2020). They differentiate into polarized effector cells already in the thymus (Lee et al., 2020) with properties similar to those of specialized CD4 T cell subsets and seed organs in developmental waves (Bendelac et al., 2007; Ribot et al., 2021; Salou et al., 2019). In the tissues, they may be localized within the parenchyma, close to or within the epithelium, or reside within the microvasculature, as in the lungs or the liver (Geissmann et al., 2005; Scanlon et al., 2011; Thomas et al., 2011). UTCs that populate barrier tissues are, based on parabiosis experiments, considered to be tissue-resident lymphocytes (Scanlon et al., 2011). UTC populations that are found in the blood and secondary lymphoid organs recirculate similar to those found in the conventional T cells (Ugur et al., 2018). However, some UTCs in LNs also appear to be tissue resident (Audemard-Verger et al., 2017). Whether the populations present in secondary lymphoid organs and those in tissues are related is incompletely understood (Gray et al., 2012). On a functional level, UTCs interact with the commensal microbiome, maintain tissue homeostasis, and participate in tissue healing and pathogen defense (Ansaldo et al., 2021; Constantinides et al., 2019). To execute these functions, UTCs can be activated in two ways. One mechanism is dual cytokine stimulation, with one mediator being an IL-1 family member (IL-1β, IL-18, and IL-33) and the other signal being a STAT-activating cytokine (e.g., IL-2, IL-4, IL-12, IL-23, and IFNI) (McGinty and von Moltke, 2020). The second activation mode is executed via their specific TCR, and some effector functions may depend on this mode of activation and are not induced by cytokines alone, as demonstrated for the secretion of IL-4 (Brigl et al., 2011). However, only a limited number of exogenous pathogen-associated TCR ligands have been identified (Chien and Konigshofer, 2007; McWilliam and Villadangos, 2017).
When addressing whether UTCs could play a role in linking tissue immunity with LN function, we found that the local composition as well as the cytokine production capacity of UTCs diverged among LNs, which had a substantial impact on the immune responses that were generated. These local differences were a consequence of continuous migration of UTCs via the lymphatic route to locally draining LNs, establishing tissue-specific UTC composition in draining LNs. This migratory behavior was conserved across TCR-based lineages (NKT, MAIT, and γδ T cells) and effector states, suggesting shared, interconnected functions of these cells. Testing their function in further depth, we demonstrated their common cytokine-dependent innate response within shared homeostatic and microanatomical niches, supporting the concept that UTCs form locally diverse yet interconnected populations—functional units. Together, we show that UTCs continuously migrate via the lymphatic route to locally draining LNs where they form functional units that generate characteristic, site-specific immune responses and, thereby, link tissue immunity to lymph node function.
Section snippets
Different LNs generate characteristic UTC-dependent immune responses
To investigate whether lymph nodes (LNs) that drain distinct tissue sites generate characteristic immune responses we globally activated (PMA/ionomycin) single-cell suspensions from different LNs (skin-draining [sd], mediastinal [med], and mesenteric [mes]) and measured the resulting cytokines that were released in the culture supernatant. We focused on signature cytokines IFNγ, IL-4, IL-17 and GM-CSF that represent a polarized immune response against intracellular pathogens (Th1), parasites
Discussion
In this study, we showed that UTCs constitutively migrated from tissues to locally draining LNs, establishing tissue-specific UTC populations across distinct LNs. As these cells are equipped with immediate innate-like effector functions, their tissue-specific composition and imprinting determined the local cytokine environment in LNs early during infection in a site-specific manner. As we showed, local differences in UTC composition differentially affected adaptive humoral immune responses
Key resources table
REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Rat IgM 17D1 (unconjugated) Kind gift from Robert Tigelaar, Yale N/A Armenian Hamster IgG anti-mouse CCR6 (CD196) (PE/Dazzle™ 594 conjugated) 29-2L17
BioLegendCat# 129822;
RRID: AB_2687019Armenian Hamster IgG anti-mouse CCR6 (CD196) (APC conjugated) 29-2L17
BioLegendCat# 129814;
RRID: AB_1877147Armenian Hamster IgG anti-mouse CCR6 (CD196) (PE conjugated) 29-2L17
BioLegendCat# 129804;
RRID: AB_1279137Mouse anti-mouse CCR9 (CD199) (FITC conjugated) CW-1.2
Acknowledgments
We would like to thank Ronald N. Germain for critically reading the manuscript. The Core Unit for FACS and the Core Unit SysMed of the IZKF Würzburg have supported this study. The authors thank the Single-Cell Center Würzburg for technical support. The CD1d and MR1 tetramer were obtained through the NIH Tetramer Core Facility. This work was funded by grants through the German Research Foundation (DFG) under Germany’s Excellence Strategy—EXC 2155 “RESIST”—project ID 390874280 to J.H., GRK2581
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