Treg suppressor function within allografts is required for tolerance

Abstract The role of regulatory CD4+Foxp3+ T cells (Treg) suppressor function within allografts in tolerance remain unclear. To directly address this, we first used a model where immune reactions are restrained to the graft only. Effector T cells (Teff) alone, or with Treg (2–3×106 each) were transferred to B6 mice lacking secondary lymphoid organs (SLO−; splenectomized LTβR−/−) 5 days after islet allograft (Balb/c). Adoptive transfer of Teff alone induced acute rejection (10d MST). In contrast, Teff + Treg co-transfers significantly prolonged graft survival, with 65% of recipients surviving long-term (>100d MST). Interestingly, Treg did not affect Teff proliferation or accumulation within allografts, but significantly reduced MHC-II expression on DCs and IFN-γ in Teff. Secondly, given that S1PR1 signaling is necessary for lymphocyte exit from SLO, we prevented Treg migration to allografts during tolerance by removing S1PR1 expression in endogenous Treg specifically. Tamoxifen-fed B6.Foxp3EGFP-Cre/Ert2.S1PR1fl/fl (S1PR1KO-Treg) or B6.S1PR1fl/fl (WT-Treg) mice received Balb/c islets. Untreated mice rejected allografts acutely (18d MST). Interestingly, while anti-CD45RB treatment induced tolerance in WT-Treg control mice (95d MST), all S1PR1KO-Treg mice rejected their graft (28d MST; p<0.05 vs WT-Treg). Also, these results correlated with a 25% decrease in Treg infiltrating islet allografts and with a 22% increase in Treg in lymph nodes on day 10 in S1PR1KO-Treg + anti-CD45RB compared to WT-Treg + anti-CD45RB recipients. Overall, our data demonstrate that Treg can suppress rejection by Teff within allografts without prior activation in SLO, and suggest that Treg suppressor function within allografts is required for tolerance.