CXCR5/CXCL13 interaction is important for double-negative regulatory T cell homing to cardiac allografts

Accumulating evidence indicates that regulatory T (Treg) cells control development of various diseases both systemically and locally. However, molecular mechanisms involved in Treg cell homing remain elusive. We have shown previously that a beta TCR(+)CD3(+)CD4(-)CD8(-) double-negative (DN) Treg cells selectively accumulate in tolerant allografts to maintain localized immune regulation. However, the molecular mechanism leading to the accumulation of DN Treg cells in tolerant grafts was not known. Our cDNA microarray analysis revealed significant up-regulation of chemokine receptor CXCR5 mRNA in DN Treg clones compared with nonregulatory clones. In this study, we examined the importance of CXCR5 in mediating DN Treg migration. Compared with CD4 and CD8 T cells, both primary DN Treg cells and clones constitutively express high levels of CXCR5 protein, enabling them to migrate toward increasing CXCL13 gradients in vitro. After infusion into recipient mice, CXCR5(+) DN Treg clones, but not their CXCR5(-) mutants, preferentially accumulated in cardiac allografts and could prevent graft rejection. Furthermore, we found that allogeneic cardiac allografts express high levels of CXCL13 mRNA compared with either recipient native hearts or nontransplanted donor hearts. Ab neutralization of CXCL13 abrogated DN Treg cell migration in vitro and prevented in vivo homing of DN Treg clones into allografts. These data demonstrate that DN Treg cells preferentially express CXCR5, and interaction of this chemokine receptor with its ligand CXCL13 plays an important role in DN Treg cell migration both in vitro and in vivo.