Tissue-specific Contribution of Mucosal-Associated Invariant T Cells to Allergic Airway Inflammation

Mucosal-associated invariant T (MAIT) cells are major histocompatibility complex (MHC) class I-related protein 1 (MR1)-restricted T cells which exert anti-microbial as well as homeostatic functions, populate all mucosal tissues as well as the skin and have a microbiome-dependent development, making their contribution to allergic diseases likely.1 Whether that contribution is protective or pathogenic remains unresolved. Indeed, we recently described the disease promoting activity of MAIT cells in atopic dermatitis (AD), while others made opposing observations for asthma2, 3—the latter observation having since been validated and expanded using a transgenic mouse strain artificially enriched in MAIT cells.4 So far, the allergic potential of MAIT cells has only been studied in single tissues, each receiving distinct imprints from resident MAIT cells.5 To determine which imprint prevails during allergic propagation from the skin to the lung, we compared MR1−/− and WT mice in an experimental model of the allergic march, comprised of epicutaneous sensitization and airway challenge to house dust mite (HDM; Figure 1A). In WT mice, this model induced a robust HDM-specific IgE response and eosinophilic lung infiltration, detectable in both the bronchoalveolar lavage fluid and lung tissue (Figure 1B–E). Remarkably, MR1−/− mice were virtually protected from disease, as reflected by all disease parameters (Figure 1B–E). To circumvent the potentially confounded phenotype of MR1−/−, due to simultaneous removal of pathogenic and protective MAIT cells in the skin and lung, respectively, we next co-applied HDM and the MAIT cell antagonist Ac-6-FP (previously used as topical treatment against experimental AD2) to spatiotemporally block MAIT cells in the skin of WT mice. As shown in Figure 1B–E, Ac-6-FP-treated WT mice closely phenocopied the reduced eosinophilia and IgE production observed in MR1−/− mice, suggesting that the pathogenic contribution of MAIT cells is restricted to the skin and that MAIT cells may influence both innate and adaptive features of allergen sensitization. To start addressing the possibility that skin MAIT cells directly interact with dendritic cells,6 we determined the proportion of activated (CD86+) type-2 dendritic cells (DC2) in the skin draining lymph node of WT mice epicutaneously sensitized with HDM and observed a significant reduction in the presence of Ac-6-FP (Figure S1A). Skin MAIT cells also influenced the innate sensitization of the pulmonary microenvironment, as reflected by the significant inhibition of eosinophilic (pre-)activation (CD69+) (not observed in the blood) and the increase of alveolar macrophages in MR1−/− mice, conferring additionally protective effects7 (Figure S1B). To confirm the responsiveness of lung-resident MAIT cells in this model, we examined their number and activation state (CD25 and CD69 expression), which we found to be significantly elevated at endpoint (Figure 1F–H). While Ye et al.3 have previously shown that pulmonary MAIT cell numbers decrease upon repeated allergen administration, as early as Day 6 after the initial exposure, the pulmonary consequences of epicutaneous sensitization (Figure S1) may have qualitatively altered the response to HDM and avoided MAIT cell decline. We also observed that our baseline number of pulmonary MAIT cells was comparatively much lower, which may be related to different environmental conditions (i.e. lower environmental bioburden), as reported for cutaneous MAIT cells.3, 8 In such a setting, HDM, as a carrier of microbial metabolites, may provide sufficient antigenic stimulation to influence MAIT cell activation/expansion. Consistently, we found i.n. HDM to increase the percentage of Nur77-positive pulmonary MAIT cells—a surrogate for recent TCR signalling—albeit without reaching statistical significance, and modestly compared to the positive control 5-OP-RU (Figure 1I). Next, we compared WT and MR1−/− mice in allergen sensitization/challenge models which bypass the skin, including HDM and Alternaria alternata (Alternaria) lung-only models, and ovalbumin (OVA) lung challenge after intraperitoneal sensitization. In strong contrast to epicutaneously sensitized mice, WT and MR1−/− showed similar levels of pulmonary eosinophilia, in terms of both frequency and absolute numbers, when initially and repeatedly challenged with HDM or Alternaria via the airways, notwithstanding a significant decrease in the number of eosinophils in MR1−/− mice challenged with Alternaria (Figure 2A,B). To reconcile these observations with the reported protective function of MAIT cells in this setting, we crossed MR1−/− with IL-4/IL-13 dual reporter mice (4C13R) and indeed observed significantly higher IL-4+ and IL4+IL-13+ CD4+ T-cell frequencies in the lung-draining lymph node of 4C13R × MR1−/− mice after two successive i.n. HDM administrations (Figure 2C), consistent with the higher concentration of IL-13 in lung tissue observed by Ye et al.3 It is important to note that the extent of eosinophilic infiltration upon i.n. allergen administration, and its reproducibility or lack thereof across studies, may be related to qualitative batch differences between allergen extracts. Indeed, the dose of Alternaria that could be safely administered to our mice was 5 μg, while a single i.n. dose of 50 μg, as used by Ye et al. (sourced from the same manufacturer), proved lethal in our hands. Similarly, the varying levels of HDM-associated endotoxins have been shown to influence the transcriptomic signature of HDM-challenged airways.9 Lastly, intraperitoneal sensitization with OVA/alum and subsequent OVA challenge led again to almost identical pulmonary eosinophilia in WT and MR1−/− mice (Figure 2D), contrasting the moderately exacerbated phenotype observed in MR1−/− mice by Sasano et al.10 In summary, our observations indicate that the contribution of MAIT cells to allergic airway inflammation is dependent upon the route of sensitization. If sensitization occurs via the skin—of particular relevance for the allergic march—MAIT cells' contribution is pathogenic, including mechanistic aspects of both innate and adaptive nature. On the contrary, if allergen is first encountered via the airways, MAIT cells exert a protective function. It remains to be determined to what extent this dichotomy is related to intrinsic functional differences between skin- and lung-resident MAIT cells, tissue-specific combinations of T-cell receptor (TCR)-dependent and -independent activation and/or MAIT cells' local interaction partners (e.g. DC and epithelial cells). This work was supported by the New Zealand Health Research Council Independent Organization Fund (14/1003) and the High-Value Nutrition National Science Challenge, New Zealand. Conceptualization: AC, KW, KN, DOS, OG; Methodology and analysis: AC, KW, KN, KG, AS, RJ, AG; Writing—draft and editing: AC, KW, KN, DOS, OG; supervision, administration and funding: OG. The manufacture of 5-(2-oxopropylideneamino)-6-D-ribitylaminouracil (5-OP-RU) was carried out by Prof. Painter's laboratory financially supported by a programme grant from the New Zealand Ministry of Business Innovation and Employment (Award number RTVU1603). We are grateful to the Hugh Green Cytometry Centre, Malaghan Institute of Medical Research, for expert support with flow cytometry and confocal microscopy, staff of the Biomedical Research Unit, Malaghan Institute of Medical Research, for animal husbandry, and all colleagues at the Malaghan Institute for insightful discussion and advice. The authors declare no conflict of interest. The data that support the findings of this study are available from the corresponding author upon reasonable request. Appendix S1 Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.