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Background: Deciphering the mechanisms of tolerance represents a crucial aim of research in transplantation. We previously identified by DNA chip interleukin (IL)-27 p28 and transforming growth factor (TGF)-β1 as overexpressed in a model of rat cardiac allograft tolerance mediated by regulatory CD4CD25 T cells. The role of these two molecules on the control of the inflammatory response remains controversial. However, both are involved in the regulation of the T helper 17/Treg axis, suggesting their involvement in tolerance., Methods: We analyzed regulation of IL-27 and TGF-β1 expression in allograft response and their role in tolerance by using blocking anti-TGF-β antibody and by generating an adeno-associated virus encoding IL-27., Results: Here, we confirmed the overexpression of IL-27 and TGF-β1 in tolerated cardiac allografts in two different rodent models. We observed that their expression correlates with inhibition of T helper 17 differentiation and with expansion of regulatory CD4CD25 T cells. We showed in a rat model that anti-TGF-β treatment abrogates infectious tolerance mediated by the transfer of regulatory CD4CD25 T cells. Moreover, overexpression of IL-27 by adeno-associated virus administration in combination with a short-term immunosuppression allows prolongation of cardiac allograft survival and one tolerant recipient. We found that IL-27 overexpression did not induce Foxp3CD4CD25 T-cell expansion but rather IL-10-expressing CD4 T cells in the tolerant recipient., Conclusions: Taken together, these data suggest that both TGF-β1 and IL-27 play a role in the mechanisms of tolerance. However, in contrast to TGF-β1, IL-27 seems not to be involved in regulatory CD4CD25 T-cell expansion but rather in their mode of action.

Background: Leukocyte depletion at the time of transplantation with alemtuzumab (Campath-1H) has been demonstrated to be a potential strategy for reducing long-term exposure to immunosuppressive drugs. Although the impact of alemtuzumab treatment on the immune system has been explored, the effects of long-term immunosuppressive therapy in alemtuzumab-treated patients still need to be elucidated., Methods: T-regulatory cells and Th1/Th17 responses were assessed by flow cytometry and real-time polymerase chain reaction more than 4 years after transplantation in 10 kidney recipients treated with alemtuzumab induction. Seven patients were converted to sirolimus monotherapy at 12 months posttransplant, whereas the remaining three patients with history of graft rejection were treated with sirolimus and mycophenolate mofetil. In addition, we sorted and expanded interleukin (IL)-17A-producing CCR6CD4 T cells and assessed their susceptibility to suppression by regulatory T (Treg) cells in in vitro suppression tests., Results: Three years of mammalian target of rapamycin inhibitor monotherapy correlates with an increase in the number of IL-17A producing cells, compared with patients treated with sirolimus and mycophenolate mofetil. In these patients, IL-17A expression was compensated for by an increase in Treg cell frequency and number. In addition, we demonstrated that both proliferation and cytokine production by Th17 cells can be effectively regulated by Treg cells., Conclusions: Our results demonstrate that history of rejection and long-term maintenance immunosuppression has an impact on the number of circulating Treg and Th17 cells. However, more importantly, we have shown that Treg cells can effectively regulate Th17cells both in vitro and in vivo.

Background: In this study, we have investigated the mechanisms involved in both the induction of suppressive anergy, the stability of the anergy induced, and the possible mechanisms by which the response of immunocompetent CD4+ T cells are suppressed., Methods: We used immobilized anti-CD3 monoclonal antibody (mAb) to induce anergy in T helper (Th) 1 and Th0 cells reactive with MHC class II molecule H2 I-Ab., Results: We observed that suppressive anergy was induced independently of costimulation in Th0 but not Th1 cells. Although the anergic and suppressive states of Th0 cells were stable in the presence of exogenous interleukin-2, this was not the case for Th1 cells. No evidence for linked epitope suppression was observed for any of the I-Ab reactive cells investigated. Neither anergy nor suppression was observed in Th0 cells upon restimulation with anti-CD3 in the presence of syngeneic antigen-presenting cells (APCs). However, anergy but not suppression was observed in co-cultures restimulated with anti-T-cell antigen receptor (TCR) mAbs/syngeneic APCs and suppression could be restored by the addition of I-Ab+ APCs., Conclusions: Overall, these data suggested that the MHC-peptide complex recognized by the Th0 cells was required for suppression of the response of immunocompetent cells. We propose that suppression is mediated either by down-modulation of the MHC-peptide complex recognized by the anergic T cells or that a molecule specific to the MHC-peptide/TCR interaction facilitates negative regulation by APC:T or T:T interactions.