Regulatory T Cell Transmigration and Intravascular Migration Undergo Mechanistically Distinct Regulation at Different Phases of the Inflammatory Response

Regulatory T cells (Tregs) play important roles in limiting inflammatory responses in the periphery. During these responses, Treg abundance in affected organs increases and interfering with their recruitment results in exacerbation of inflammation. However, the mechanisms whereby Tregs enter the skin remain poorly understood. The aim of this study was to use intravital microscopy to investigate adhesion and transmigration of Tregs in the dermal microvasculature in a two-challenge model of contact sensitivity. Using intravital confocal microscopy of Foxp3-GFP mice, we visualized endogenous Tregs and assessed their interactions in the dermal microvasculature. Four hours after hapten challenge, Tregs underwent adhesion with ∼25% of these cells proceeding to transmigration, a process dependent on CCR4. At 24 h, Tregs adhered but no longer underwent transmigration, instead remaining in prolonged contact with the endothelium, migrating over the endothelial surface. Four hours after a second challenge, Treg transmigration was restored, although in this case transmigration was CCR4 independent, instead involving the CCR6/CCL20 pathway. Notably, at 24 h but not 4 h after challenge, endothelial cells expressed MHC class II (MHC II). Moreover, at this time of peak MHC II expression, inhibition of MHC II reduced Treg adhesion, demonstrating an unexpected role for MHC II in Treg attachment to the endothelium. Together these data show that Treg adhesion and transmigration can be driven by different molecular mechanisms at different stages of an Ag-driven inflammatory response. In addition, Tregs can undergo prolonged migration on the inflamed endothelium.