Quantification of lymph node transit times reveals differences in the trafficking behavior of regulatory versus conventional CD4+ T cells

Abstract

Abstract Regulatory T cells (Tregs) express the canonical marker Foxp3 and are critical for suppressing the immune response in homeostatic and inflammatory conditions. Despite how widely studied CD4+Foxp3+ Tregs are in mice, their physiologic trafficking and scanning behavior of antigen-presenting cells in secondary lymphoid organs (SLOs) have yet to be quantitatively analyzed in the intravital setting. Our laboratory has previously described the dynamics of T cell trafficking through SLOs and scanning of dendritic cells (DCs), revealing that CD4+ T cells and CD8+ T cells exhibit different transit kinetics through lymph nodes (LNs). On average, CD4+ T cells spend less time scanning for self-peptide/MHC (self-pMHC), dwell shorter in LNs, and egress faster compared to CD8+ T cells. Here we use flow cytometry and 2-photon laser scanning microscopy (2-PLSM) to quantitate the homeostatic trafficking behavior and surveillance strategy of Tregs in LNs and to compare their behavior to conventional T cells (Tconvs). Our data reveal surprising differences in Treg versus Tconv trafficking in mesenteric versus other peripheral LNs. Furthermore, we find that Treg LN transit time is heavily dependent on recognition of self-pMHC. Finally, we use intravital 2-PLSM to quantify the contribution of self-pMHC recognition to Treg kinetics and scanning of DCs in peripheral and mesenteric LNs. These results add quantitative data to the emerging paradigm that self-pMHC interactions dynamically control homeostatic Treg retainment and localized suppressive function in peripheral LNs.