FTY720 treatment in skin allograft rejection Commodaro AG et al. Sewgobind et al. (35) showed that the relative mRNA expression of FOXP3 decreased in the blood of kidney- transplanted patients treated with rabbit anti-thymocyte globulin (rATG) or rATG + tacrolimus, whereas the levels of IL-10 and IL-27p28 increased compared with the levels in non-transplanted blood donors. Battaglia et al. (36) showed that the administration of IL- 10 in combination with the immunosuppressive drug rapamycin promoted tolerance in type 1 diabetic mice after pancreatic islet transplantation by the induction of Tr1 cells. In addition, IL-27 secretion from dendritic cells has been shown to enhance the development of Tr1 cells and, as observed by Yoshimoto et al. (37), inhibit not only Th2 commitment but also Th2 cytokine production from already polarized Th2 cells (down-regulation of GATA-3 and up- regulation of T-bet). Moreover, IL-27 was shown by Stumhofer et al. (38) to directly antagonize the development of the Th17-cell responses in experimental autoimmune models. In our model, even though statistical significance could not be reached among groups, the prolonged skin allograft survival promoted by FTY720 treatment could be explained by the increased expression of IL-10 and IL-27 in the skin graft, suggesting the development of the Tr1 cells. However, we also found an increase in the IL-23 expression in the skin graft, confirming the presence of Th17 cells. In agreement with our data, Li et al. (39) found both Treg and Th17 cells in mouse liver transplants. They showed a time-dependent and significant increase in Th17-related cytokines (IL-17, IL- 6 and IL-23), whereas the expression of Treg-related cytokines (IL-10, TGF-b1) was markedly low in the liver of transplanted mice prior to graft rejection. Our results show that FTY720 treatment causes a different modulation of IL-10 and IL-23 in the spleen and skin graft; namely, the IL-10 and IL-23 expression decreased in the spleen but increased in the graft. In contrast, IL-27 and Foxp3 exhibited the same modulation in the spleen and skin, with an increase in IL-27 and decrease in Foxp3. These findings suggest that, at early time points (day +5) after transplantation, FTY720 controls the balance between Tr1 (IL-10 and IL-27) and Th17 (IL-23) in the graft, which prevents rejection for a definite period of time. In conclusion, we demonstrated that skin allograft survival was prolonged but not indefinite in FTY720-treated mice, indicating that the drug did not prevent the long-term imbalance between the Treg and Th17 cells in the graft, which thus led to rejection. ACKNOWLEDGMENTS This study was supported by grants from Fundac¸a˜o de Amparo a Pesquisa do Estado de Sa˜ o Paulo (FAPESP), Conselho Nacional de Desenvolvimento Cient´ıfico e Tecnolo´gico (CNPq) and CAPES. AUTHOR CONTRIBUTIONS Commodaro AG performed the skin transplantation, real-time PCR, Bio- Plex cytokine assay, FTY720 treatment, tissue harvesting and flow cytometry; conducted the statistical analyses; made the figures; and wrote the manuscript. Pedregosa JF performed the skin transplantation, FTY720 treatment, tissue harvesting and flow cytometry. Bueno V performed the skin transplantation, FTY720 treatment, tissue harvesting and flow cytometry; conducted the statistical analyses; made the figures; and wrote the manuscript. Peron JP and Branda˜o WN performed the real-time PCR and the Bio-Plex cytokine assay. Rizzo LV conducted the statistical analyses, made the figures, and wrote the manuscript. 812 CLINICS 2012;67(7):805-813 REFERENCES 1. Rosenberg AS, Singer A. 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