Intravenous immunoglobulins promote skin allograft acceptance by triggering functional activation of CD4+Foxp3+ T cells.

BACKGROUND: Intravenous immunoglobulins (IVIg) therapy is effective as a treatment for T-cell-mediated immune diseases, but whether and how IVIg suppress allogeneic T-cell responses is largely unknown. METHODS: In vitro, human CD4(+), CD4(+)CD25(-), or CD4(+)CD25(+) T cells were stimulated with allogeneic antigen-presenting cells (APCs), and mouse CBA/Ca (H2(k)) CD4(+) or CD4(+)CD25(-) T cells were stimulated with C57BL/10 (H2(b)) splenocytes, in the presence or absence of IVIg. Proliferation, binding of IVIg, expression of activation markers, and ZAP70-phosphorylation were determined. In vivo, 1x10(5) CD4(+) or CD4(+)CD25(-) T cells of CBA/Ca mice were adoptively transferred into CBA/RAG1(-/-) mice, which were 1 day later transplanted with skin grafts of C57BL/10 mice. IVIg was administered intravenously and skin graft survival was determined. RESULTS: IVIg bound to the surface of human and mouse CD4(+)Foxp3(+) regulatory T cells (Tregs). IVIg binding resulted in functional activation of Tregs, as detected by increased expression of surface activation markers, enhanced ZAP70-phosphorylation, and increased capacity to suppress allogeneic T-cell proliferation. IVIg inhibited allogeneic T-cell proliferation in the presence of Tregs, but this effect was abrogated on selective depletion of CD25(+) cells from responder T cells. IVIg prevented T-cell-mediated rejection of fully mismatched skin grafts in CBA/RAG1(-/-) mice reconstituted with CD4(+) T cells, but this effect was lost on selective depletion of CD4(+)CD25(+) cells from transferred T cells, indicating that IVIg induced dominant allograft protection mediated by Tregs. CONCLUSIONS: Our data show that IVIg suppress allogeneic T-cell responses by direct activation of Tregs. IVIg treatment, which has been proven safe, may have therapeutic potential in tolerance-inducing strategies in transplant medicine.

Interferon gamma: friend or foe?

Interferon gamma (IFN-gamma) can elicit an inflammatory TH1-driven immune response but has also been found to be necessary for long-term allograft survival induced by costimulation blockade. Recently, we have found that regulatory T cells rapidly and transiently produce IFN-gamma creating a microenvironment that can influence the function of antigen presenting cells (APCs), T-cell proliferation and activation as well as T-cell effector mechanisms, thereby controlling immune responses locally. Moreover, addition of IFN-gamma to cocultures of T cells and APCs can drive the generation of T cells with regulatory activity. Thus, the influence of IFN-gamma on the immune response to a transplant is likely to be context dependent.

Functional dichotomy of NK cells in organ transplantation.

Evaluation of: Yu G, Xu X, Vu MD, Kilpatrick ED, Li XC. NK cells promote transplant tolerance by killing donor antigen-presenting cells. J. Exp. Med. 203, 1851-1858 (2006). Natural killer (NK) cells have the potential to display different functional activities after transplantation. The traditional view is that NK cells have the capacity to contribute to rejection by facilitating the activation/differentiation of leukocytes that destroy the graft. By contrast, in the article under review, a novel role for NK cells was identified in the setting of costimulation blockade where alloreactive NK cells of recipient origin were found to have the capacity to kill donor-derived antigen-presenting cells, thereby reducing T-cell priming and promoting long-term skin graft acceptance.

CD25+CD4+ regulatory T cells develop in mice not only during spontaneous acceptance of liver allografts but also after acute allograft rejection.

BACKGROUND: Liver grafts transplanted across a major histocompatibility barrier are accepted spontaneously and induce donor specific tolerance in some species. Here, we investigated whether liver allograft acceptance is characterized by, and depends upon, the presence of donor reactive CD25CD4 regulatory T cells. METHODS: CD25 and CD25CD4 T cells, isolated from CBA. Ca (H2) recipients of C57BL/10 (B10; H2) liver and heart allografts 10 days after transplantation, were transferred into CBA. Rag1 mice to investigate their influence on skin allograft rejection mediated by CD45RBCD4 effector T Cells. RESULTS: Fully allogeneic B10 liver allografts were spontaneously accepted by naive CBA.Ca recipient mice, whereas B10 cardiac allografts were acutely rejected (mean survival time=7 days). Strikingly, however, CD25CD4 T cells isolated from both liver and cardiac allograft recipients were able to prevent skin allograft rejection in this adoptive transfer model. Interestingly, CD25CD4 T cells isolated from liver graft recipients also showed suppressive potency upon adoptive transfer. Furthermore, depletion of CD25CD4 T cells in primary liver allograft recipients did not prevent the acceptance of a secondary donor-specific skin graft. CONCLUSIONS: Our data provide evidence that the presence of CD25CD4 regulatory T cells is not a unique feature of allograft acceptance and is more likely the result of sustained exposure to donor alloantigens in vivo.

CD25+CD4+ regulatory T cells generated by exposure to a model protein antigen prevent allograft rejection: antigen-specific reactivation in vivo is critical for bystander regulation.

The importance of CD25(+)CD4(+) regulatory T (Treg) cells in the control of immune responses is established, but their antigen specificity in vivo remains unclear. Understanding Treg-cell specificity requirements will be important if their potential is to be developed for immunotherapy. Pretreatment of recipient mice with donor alloantigen plus anti-CD4 antibody generates CD25(+)CD4(+) Treg cells with the capacity to prevent skin allograft rejection in adoptive transfer recipients. Here we demonstrate that, although this regulation can be antigen-specific, reactivation with the original tolerizing alloantigen allows the Treg cells to suppress rejection of third-party allografts. Aware of the limitations of alloantigen pretreatment, we asked whether graft-protective Treg cells could be generated against unrelated, nongraft antigens. We demonstrate that bystander regulation also extends to CD25(+)CD4(+) Treg cells generated in vivo by exposure to nominal antigens under anti-CD4 antibody cover. Providing these Treg cells are reexposed to the tolerizing antigens before adoptive transfer, they prevent the rejection of fully allogeneic skin grafts. That this might form the basis of a clinically relevant tolerance induction strategy is demonstrated by the fact that, when combined with subtherapeutic anti-CD8 antibody, Treg cells generated in response to nongraft antigens facilitate the acceptance of cardiac allografts in primary recipients.

Alloantigen-induced CD25+CD4+ regulatory T cells can develop in vivo from CD25-CD4+ precursors in a thymus-independent process.

The capacity of naturally occurring autoreactive CD25+CD4+ regulatory T cells (Treg) to control immune responses both in vivo and in vitro is now well established. It has been demonstrated that these cells undergo positive selection within the thymus and appear to enter the periphery as committed CD25+CD4+ Treg. We have shown previously that CD25+CD4+ Treg with the capacity to prevent skin allograft rejection can be generated by pretreatment with donor alloantigen under the cover of anti-CD4 therapy. Here we demonstrate that this process does not require an intact thymus. Furthermore, generation of these Treg is not dependent on the expansion of CD25+CD4+ thymic emigrants, because depletion of CD25+ cells before pretreatment does not prevent Treg development, and Treg can be generated from CD25-CD4+ precursors. Taken together, these results clearly demonstrate that CD25+CD4+ Treg can be generated in the periphery from CD25-CD4+ precursors in a pathway distinct to that by which naturally occurring autoreactive CD25+CD4+ Treg develop. These observations may have important implications for the design of protocols, both experimental and clinical, for the induction of tolerance to autoantigens or alloantigens in adults with limited thymic function.

Regulatory T cells in transplantation.

There has recently been an explosion of renewed interest in regulatory T cells, particularly those within the CD4(+)CD25(+) population. It is becoming increasingly apparent that these cells exist not only as naturally occurring cells that may contribute to the maintenance of self-tolerance, but they also have the potential to prevent rejection of allografts in experimental models. Such cells have now been identified in humans as well as in rodents.

The role of the graft in establishing tolerance.

At the present time, clinical solid organ transplantation continues to rely on the use of non-specific immunosuppressive protocols in order to prevent graft rejection. However, these regimens bring with them complications related both to the global immunosuppression that they cause, and to toxicity related to individual drugs. The pursuit of protocols that will allow graft-specific tolerance thus remains a major goal of research both in animal models and in clinical practice. There is evidence that the graft itself may play an active part in establishing and maintaining donor-specific hyporesponsiveness and ultimately tolerance; the aim of this review is to analyze this role in more detail.

CD25+CD4+ regulatory T cells prevent graft rejection: CTLA-4- and IL-10-dependent immunoregulation of alloresponses.

Specific and selective immunological unresponsiveness to donor alloantigens can be induced in vivo. We have shown previously that CD25+CD4+ T cells from mice exhibiting long-term operational tolerance to donor alloantigens can regulate rejection of allogeneic skin grafts mediated by CD45RB(high)CD4+ T cells. In this study, we wished to determine whether donor-specific regulatory cells can be generated during the induction phase of unresponsiveness, i.e., before transplantation. We provide evidence that pretreatment with anti-CD4 Ab plus a donor-specific transfusion generates donor-specific regulatory CD25+CD4+ T cells that can suppress rejection of skin grafts mediated by naive CD45RB(high)CD4+ T cells. Regulatory cells were contained only in the CD25+ fraction, as equivalent numbers of CD25-CD4+ T cells were unable to regulate rejection. This pretreatment strategy led to increased expression of CD122 by the CD25+CD4+ T cells. Blockade of both the IL-10 and CTLA-4 pathways abrogated immunoregulation mediated by CD25+ T cells, suggesting that IL-10 and CTLA-4 are required for the functional activity of this population of immunoregulatory T cells. In clinical transplantation, the generation of regulatory T cells that could provide dynamic control of rejection responses is a possible route to permanent graft survival without the need for long-term immunosuppression.