Regulatory T cell (Treg) based therapies have been proposed as a therapy for graft-versus-host disease (GVHD) in recipients of allogeneic haematopoietic stem cell transplants (alloHSCT). However how Tregs suppress GVHD is not well understood. Our previous work in lymphopaenic Rag-KO mice demonstrated that infusing and expanding Tregs prior to T cell transfer promoted optimal T cell reconstitution. Tregs selectively inhibited the rapid oligoclonal expansion of transferred T cells, while permitting slower homeostatic repopulation of naïve T cell pool. This process was critically dependent upon Treg-mediated regulation of co-stimulatory molecules on dendritic cells. Based on these findings this protocol was then applied to a preclinical model of alloHSCT. Tregs were infused at the time of transplantation and expanded using IL-2/anti-IL-2 complexes, while the transfer of allogeneic Tconv cells was delayed until normalization of DC costimulation was achieved. Treg infusion prior to transfer of allogeneic T cells completely protected against disease, which was a marked improvement over a more conventional Treg-Tconv co-transfer protocol.

Our data characterising the effect of Tregs on allogeneic T cells strongly parallels our results in lymphopaenic Rag-KO mice. In mice that did not receive Tregs allogeneic T cells underwent rapid proliferation, acquired an effector memory phenotype and expressed high levels of inflammatory cytokines. Minimal homeostatic repopulation of naïve T cells took place in these mice. In contrast Tregs inhibited the rapid proliferative response of allogeneic T cells while promoting homeostatic repopulation of the naïve T cell pool. As a result, the T cell pool in these mice had an increased frequency of naïve T cells, a reduction in effector/memory T cells, and reduced expression of inflammatory cytokines. These results indicate that Treg infusion prior to transfer of allogeneic T cells not only potently protects against GVHD, but also promotes optimal reconstitution of the T cell pool.