Regulatory T cells (Tregs) are essential lymphocytes to maintain the immunological self-tolerance and suppress the excessive immune responses. We previously performed single-cell gene expression analysis of Tregs in draining lymph node of contact hyper sensitivity (CHS) model and revealed that a rare Treg subset which co-expresses multiple functional molecules (IL-10, granzyme B, TGF-β, and CTLA-4) and has capacity to remain in the inflamed skin is responsible for suppression of the inflammation. However, we did not investigate Tregs purified from skin and expression profiles of many other Treg functional molecules than the four molecules. In this study, to identify the novel functional Treg subsets and their tissue tropism in the inflamed skin, we investigated the single-cell gene and protein expression profiles of immunoinhibitory and migration-related molecules in skin Tregs of CHS-induced KikGR knock-in mouse by which cells remaining in the skin and coming to the skin are either discriminated.
We observed the functional and migratory heterogeneity in skin Tregs. Correlation analysis revealed that the expressions of some functional molecules (CD39, CD25, granzyme B, and so on) were positively correlated with each other and their expressions were negatively correlated with neuropilin-1 (Nrp1) expression. Skin-remaining Tregs included more CD39+ and CD25+ cells and less Nrp1+ cells than skin-coming Tregs. Moreover, expression profiles of migration-related molecules in CD39+ and Nrp1+ Tregs were distinct each other. Using these single-cell protein expression data, we discriminated skin-remaining and skin-coming Tregs by principal component analysis without the migratory information derived from KikGR signal profiles. Lag3- or Il10-expressing Tregs were rare and included more in CD39+ Tregs than in Nrp1+ Tregs. These data suggest that functional Treg subsets have distinct tissue tropism in the inflamed skin. Augmentation of skin-remaining high-functional Tregs could be novel therapy for the skin inflammation diseases.