Background: The clinical application of xenotransplantation often leads to T cell-mediated graft rejection. Immunosuppressive agents including polyclonal regulatory T cells (poly-Tregs) promote global immunosuppression, resulting in serious infection and malignancy in patients. Xenoantigen-expanded Tregs (xeno-Tregs) has become a promising immune therapy strategy that can protect xenografts with less side effects. In this study, we aimed to uncover a more efficient and stabler species of xeno-Tregs.

Methods: We enriched the CD27+ xeno-Tregs by cell sorting and evaluated their suppressive functions and stability in vitro via mixed lymphocyte reaction (MLR), RT-PCR, an inflammatory induction assay and western blotting. A humanized xenotransplanted mouse model was used to evaluate the function of CD27+ xeno-Tregs in vivo.

Results: Our results showed that CD27+ xeno-Tregs expressed higher levels of Foxp3, CTLA-4, and Helios and lower IL-17 than their CD27- counterparts. In addition, CD27+ xeno-Tregs showed significantly enhanced potency in suppressing the proliferation of xenoantigen-specific responder T cells at ratios of 1:4 and 1:16. Under inflammatory conditions, CD27+ xeno-Tregs displayed lower levels of IL-17 and IFN-gamma and less converted to IL17+ cells. Mice received porcine grafts showed normal tissue phenotype and less leukocyte infiltration after the injection of CD27+ xeno-Tregs.

Conclusion: Taken together, these data indicated that CD27+ xeno-Tregs could suppress immune responses in a xenoantigen specific manner, effectively protecting xenografts from tissue damage.