Impact of infection on transplantation tolerance

Allograft tolerance is the ultimate goal of organ transplantation. Current strategies for tolerance induction mainly focus on inhibiting alloreactive T cells while promoting regulatory immune cells. Pathogenic infections may have direct impact on both effector and regulatory cell populations, therefore can alter host susceptibility to transplantation tolerance induction as well as impair the quality and stability of tolerance once induced. In this review, we will discuss existing data demonstrating the effect of infections on transplantation tolerance, with particular emphasis on the role of the stage of infection (acute, chronic, or latent) and the stage of tolerance (induction or maintenance) in this infection‐tolerance interaction. While the deleterious effect of acute infection on tolerance is mainly driven by proinflammatory cytokines induced shortly after the infection, chronic infection may generate exhausted T cells that could in fact facilitate transplantation tolerance. In addition to pathogenic infections, commensal intestinal microbiota also has numerous significant immunomodulatory effects that can shape the host alloimmunity following transplantation. A comprehensive understanding of these mechanisms is crucial for the development of therapeutic strategies for robustly inducing and stably maintaining transplantation tolerance while preserving host anti‐pathogen immunity in clinically relevant scenarios.     

Tweaking the immune system: Gene therapy-assisted autologous haematopoietic stem cell transplantation as a treatment for autoimmune disease

Autoimmune diseases represent a major challenge for medical research. The aberrant self-recognition by the immune system leads to a range of pathologies for which cures have not been forthcoming. Treatments are commonly non-specific and often lead to unwanted side-effects. A number of strategies are currently being explored to tackle autoimmunity; aimed at eliminating existing pathogenic clones and the induction of immune tolerance through resetting or regulating the immune system. Autologous haematopoietic stem cell transplantation (HSCT) is one such strategy and is being trailed in a number of autoimmune diseases. However, a common feature of this strategy is disease relapse and may indicate incomplete tolerance mechanisms. It is well known that bone marrow derived cells have a major influence on immune tolerance. It is also well documented that ectopic expression of antigens within the immune system can promote robust tolerance. This review considers these observations in the context of promoting a strategy involving genetic manipulation of haematopoietic stem cells together with HSCT to induce immune tolerance and tackle autoimmunity.     

Tweaking the immune system: gene therapy-assisted autologous haematopoietic stem cell transplantation as a treatment for autoimmune disease

Autoimmune diseases represent a major challenge for medical research. The aberrant self-recognition by the immune system leads to a range of pathologies for which cures have not been forthcoming. Treatments are commonly non-specific and often lead to unwanted side-effects. A number of strategies are currently being explored to tackle autoimmunity; aimed at eliminating existing pathogenic clones and the induction of immune tolerance through resetting or regulating the immune system. Autologous haematopoietic stem cell transplantation (HSCT) is one such strategy and is being trailed in a number of autoimmune diseases. However, a common feature of this strategy is disease relapse and may indicate incomplete tolerance mechanisms. It is well known that bone marrow derived cells have a major influence on immune tolerance. It is also well documented that ectopic expression of antigens within the immune system can promote robust tolerance. This review considers these observations in the context of promoting a strategy involving genetic manipulation of haematopoietic stem cells together with HSCT to induce immune tolerance and tackle autoimmunity.     

Host-Derived CD8+ Dendritic Cells Protect Against Acute Graft-versus-Host Disease after Experimental Allogeneic Bone Marrow Transplantation

Graft-versus-host disease (GVHD) is a frequent life-threatening complication after allogeneic hematopoietic stem cell transplantation (HSCT) and induced by donor-derived T cells that become activated by host antigen-presenting cells. To address the relevance of host dendritic cell (DC) populations in this disease, we used mouse strains deficient in CD11c⁺ or CD8α⁺ DC populations in a model of acute GVHD where bone marrow and T cells from BALB/c donors were transplanted into C57BL/6 hosts. Surprisingly, a strong increase in GVHD-related mortality was observed in the absence of CD11c⁺ cells. Likewise, Batf3-deficient (Batf3^-/-) mice that lack CD8α⁺ DCs also displayed a strongly increased GVHD-related mortality. In the absence of CD8α⁺ DCs, we detected an increased activation of the remaining DC populations after HSCT, leading to an enhanced priming of allogeneic T cells. Importantly, this was associated with reduced numbers of regulatory T cells and transforming growth factor-β levels, indicating an aggravated failure of peripheral tolerance mechanisms after HSCT in the absence of CD8α⁺ DCs. In summary, our results indicate a critical role of CD8α⁺ DCs as important inducers of regulatory T cell–mediated tolerance to control DC activation and T cell priming in the initiation phase of GVHD.     

Tolerance, suppression and the fetal allograft

In solid organ transplantation the recipient immune system recognises foreign alloantigens expressed by the graft. This results in an immune attack of the transplanted organ leading to rejection, which can be prevented only by therapeutic immunosuppression. During pregnancy the fetus should also be rejected by the maternal immune system, since it expresses antigens derived from the father. Whilst the immune system retains the ability to respond to foreign antigen, tolerance mechanisms ensure that inappropriate responses against self-antigen are prevented. Maternal immune aggression directed against the fetus is partly inhibited by peripheral tolerance mechanisms that act locally to deplete cells capable of attacking the fetus. Other local mechanisms inhibit the pathways that cause tissue damage after immune activation. Recent studies in mice and humans indicate that the maternal immune system undergoes a more systemic change that promotes materno-fetal tolerance. Naturally occurring regulatory T cells, which are commonly associated with maintaining tolerance to self-antigens, can also suppress maternal allo-responses targeted against the fetus. We review the mechanisms that mediate materno-fetal tolerance, with particular emphasis on changes in regulatory T cell function during pregnancy and discuss their implications.     

Murine bone marrow stromal progenitor cells elicit an in vivo cellular and humoral alloimmune response.

Stromal progenitor cells (SPC) exhibit immunosuppressive effects in vitro that have led to speculation regarding their capacity to evade host immune recognition and to treat autoimmune diseases and gravt-versus-host disease. However, there is little in vivo experimental data to support these immunologic claims. To assess immune recognition of SPC in vivo, we evaluated the immune response of animals transplanted with SPC. C57BL/6 (B6) or Balb/c adult, murine, bone marrow-derived SPC (AmSPC) were administered by intraperitoneal injection into B6 recipients. T cell proliferation and alloantibody response was assessed from spleens and peripheral blood harvested from transplanted animals and analyzed by cell proliferation assay and flow cytometry. To assess tolerance induction, transplanted animals also received allogeneic skin grafts. Animals injected with allogeneic AmSPC mounted an accelerated CD4 response to alloantigen compared to syngeneic AmSPC injected and uninjected controls. Allogeneic AmSPC-injected animals also demonstrated high titers (> or =1:1000) of antibody directed against allogeneic AmSPC targets. Animals primed with donor or host-matched AmSPC also failed to induce tolerance, and all animals exhibited rejection of allogeneic skin grafts (n = 7, P < .0001). In contrast to their in vitro behavior, our data demonstrate that AmSPC are recognized by the host immune system in vivo, elicit a cellular and humoral immune response, and fail to induce tolerance. These findings have significant implications for all allogeneic SPC-based therapeutic strategies.     

Regulatory T cells after organ transplantation: where does their action take place?

Regulatory T cells are considered to be pivotal for the induction of tolerance to donor antigens. In the past decades, several regulatory T-cell subsets have been identified, such as CD4(+)CD25(+) regulatory T cells and the CD8(+)CD28(-) suppressor T cells. Although many studies have investigated the role of these regulators in transplant tolerance, relatively little attention has focused on the exact place where these cells suppress immune responses directed to donor antigens. The localization of regulatory T cells may influence their effect on allogeneic immune responses. More insight into the localization and migration of regulatory T cells in transplant recipients is therefore important, especially when these cells are to be used for monitoring purposes and for cellular immune therapy. In the present review we summarize current knowledge about the presence of functional donor-directed regulatory T cells in the secondary lymphoid organs, peripheral blood, and the transplanted organ itself. In addition, we discuss the importance of the appropriate localization for the control of anti-donor immune reactivity.     

Parenchymal cells in immune and tolerance induction

There is increasing evidence that dendritic cells are capable of inducing T cell tolerance to tissue-specific antigens by presenting these antigens to CD4 and CD8 T cells in the respective draining lymph nodes. In contrast, parenchymal cells are often seen as immunologically inert. This lecture summarizes studies showing that tissue cells like sinusoidal endothelial cells and hepatocytes in the liver as well as keratinocytes in the skin can induce peripheral T cell tolerance. This tolerance induction is often preceded by T cell activation. Thus, several tolerance mechanisms are operating in parallel. We still have no learn which mechanism is most suitable for therapeutic intervention in autoimmune diseases and organ transplantation.     

Gene therapy in the transplantation of allogeneic organs and stem cells

Most of the current hematopoietic stem cell (HSC) -directed gene therapy applications have focused on the replacement of defective or deficient genes in an autologous setting. More recently HSC gene therapy applications have also included the enhancement or improvement of HSC features. Allogeneic HSCs have been used to facilitate and improve allogeneic transplantation and to achieve tolerance to transplanted cells, tissues or organs. Different gene transfer approaches addressing a variety of immunomodulatory mediators contributing to graft tolerance or immunological ignorance may have a critical role in improving long-term graft survival. Allogeneic tissues are frequently recognized by allospecific T cells as foreign and are rapidly rejected in the absence of immunosuppression. The higher susceptibility to cancer and infectious diseases of immunosuppressed patients led to investigation of new therapies to induce graft-specific tolerance. Peripheral tolerance to allogeneic grafts can be achieved by a variety of mechanisms including clonal deletion, suppression caused by regulatory T cells and anergy induction associated with microchimerism effect. In the last decades, potential candidates to confer allograft protection were identified. In this review, we summarize ongoing strategies and developments in genetic manipulation of cells, tissues and organs for allogeneic transplantation including modulating the effector arm of the immune response.     

T-cell-based immunotherapy in multiple sclerosis: induction of regulatory immune networks by T-cell vaccination

Multiple sclerosis (MS) is a chronic inflammatory disease of the CNS with presumed autoimmune origin. Pathogenic autoimmune responses in MS are thought to be the result of a breakdown of self tolerance. Several mechanisms account for the natural state of immunological tolerance to self antigens, including clonal deletion of self-reactive T cells in the thymus. However, autoimmune T cells are also part of the normal T-cell repertoire, supporting the existence of peripheral regulatory mechanisms that keep these potentially pathogenic T cells under control. One such mechanism involves active suppression by regulatory T cells. It has been indicated that regulatory T cells do not function properly in autoimmune disease. Immunization with attenuated autoreactive T cells, T-cell vaccination, may enhance or restore the regulatory immune networks to specifically suppress autoreactive T cells, as shown in experimental autoimmune encephalomyelitis, an animal model for MS. In the past decade, T-cell vaccination has been tested for MS in several clinical trials. This review summarizes these clinical trials and updates our current knowledge on the induction of regulatory immune networks by T cell vaccination.     

Treg cell therapy: How cell heterogeneity can make the difference

CD4⁺CD25^highCD127^low/−FOXP3⁺ T regulatory cells are responsible for maintaining immune tolerance and controlling excessive immune responses. Treg cell use in pre-clinical animal models showed the huge therapeutic potential of these cells in immune-mediated diseases and laid the foundations for their applications in therapy in humans. Currently, there are several clinical trials utilizing the adoptive transfer of Treg cells to reduce the morbidity in autoimmune disorders, allogeneic HSC transplantation, and solid organ transplantation. However, a large part of them utilizes total Treg cells without distinction of their biological variability. Many studies on the heterogeneity of Treg cell population revealed distinct subsets with different functions in the control of the immune response and induction of peripheral tolerance. Some of these subsets also showed a role in controlling the general homeostasis of non-lymphoid tissues. All these Treg cell subsets and their peculiar properties can be therefore exploited to develop novel therapeutic approaches. This review describes these functionally distinct subsets, their phenotype, homing properties and functions in lymphoid and non-lymphoid tissues. In addition, we also discuss the limitations in using Treg cells as a cellular therapy and the strategies to enhance their efficacy.     

The transplantation of hematopoietic stem cells after non-myeloablative conditioning: a cellular therapeutic approach to hematologic and genetic diseases

Originally, allogeneic hematopoietic stem cell transplantation (HSCT) was viewed as a form of rescue from the marrow lethal effects of high doses of chemo-radiotherapy used to both eradicate malignancy and to provide sufficient immunosuppression to ensure allogeneic engraftment. Clear evience of a therapeutic graft-versus-tumor (GVT) effect mediated by allogeneic affector cells (T cells) has prompted the exploration of HSCT regimens that rely solely upon host immunosuppression (non-myeloblative) to facilitate allogenic donor engraftment. The engrafted donor effector cells are then used to accomplish the task of eradicating host malignant cells. The non-myeloblative regimen developed in Seattle uses 2 Gy total body irradiation (TBI) before transplant followed by postgrafting cyclosporine (CSP) and mycophenolate mofetil (MMF). This regimen resulted in initial mixed donor-host chimerism in all patients with hematologic malignancies and genetic disorders who received HLA-matched sibling allografts. The 17% incidence of graft rejection was reduced to 3% with the addition of fludarabine, 30 mg/m²/day on d-4,-3, and-2. The non-myeloblative combination of fludarabine/TBI has also been successful at achieving high engraftment rates in recipients of 10 of 10 HLA antigen matched unrelated donor HSCTs in patients with hematologic malignancies. By reducing acute toxicities relative to conventional HSCT, most patients have received their pre- and post-HSCT therapy almost exclusively as outpatients. Acute and chronic GVHD occur after non-myeloablative HSCT, but the incidence and severity appear less compared to conventional HSCT. As in conventional transplants, immune dysregulation from GVHD and its treatment and delayed reconstitution of immune function continue to present risks to patients who have otherwise undergone successful non-myeloablative HSCT. Cellular therapeutic effects have been nobserved after non-myeloblative HSCT such as correction of inherited genetic disorders, and eradication of hematologic malignant diseases and renal cell carcinoma via GVT responses.     

Induction of transplantation tolerance to fully mismatched cardiac allografts by T cell mediated delivery of alloantigen

Induction of transplantation tolerance has the potential to allow for allograft acceptance without the need for life-long immunosuppression. Here we describe a novel approach that uses delivery of alloantigen by mature T cells to induce tolerance to fully allogeneic cardiac grafts. Adoptive transfer of mature alloantigen-expressing T cells into myeloablatively conditioned mice results in long-term acceptance of fully allogeneic heart transplants without evidence of chronic rejection. Since myeloablative conditioning is clinically undesirable we further demonstrated that adoptive transfer of mature alloantigen-expressing T cells alone into mice receiving non-myeloablative conditioning resulted in long-term acceptance of fully allogeneic heart allografts with minimal evidence of chronic rejection. Mechanistically, tolerance induction involved both deletion of donor-reactive host T cells and the development of regulatory T cells. Thus, delivery of alloantigen by mature T cells induces tolerance to fully allogeneic organ allografts in non-myeloablatively conditioned recipients, representing a novel approach for tolerance induction in transplantation.     

Intrathymic Tolerance Induction: Determination of Tolerance to Class II Major Histocompatibility Complex Antigens in Maturing T Lymphocytes by a Bone Marrow-Derived Non-Lymphoid Thymus Cell

Two new experimental approaches were established to analyse the influence of the thymus on tolerance induction to major histocompatibility complex (MHC) antigens: The aim of the first experiment was to perform successful transplantation of adult allogeneic thymus tissue into nude mice, an attempt that has been unsuccessful in the past. Tolerance for the MHC genotype of a prospective thymus graft recipient (A) was induced in mice of strain B by injection of (A X B) splenocytes during the neonatal period. Adult thymic tissue obtained from these allogeneic donors (B) were grafted into the nude mice of strain A. The allogeneic thymus was accepted by the nude mice and immunoreconstitution was achieved. Subsequently the recipients developed tolerance to the MHC antigens of the allogeneic thymus donor as proved by mixed lymphocyte cultures and the acceptance of skin grafts. The second experiment was designed to determine which Ia-positive thymic compartment participates in conferring tolerance to MHC antigens in maturing T lymphocytes. Chimaeric thymus grafts were created by transplantation of neonatal thymus (A) into allogeneic nude mice (B) for a period of 8 weeks. The graft was populated with host bone marrow-derived Ia antigen-positive cells. The chimaeric thymuses consisting of type A epithelium but populated with both type A and B lymphocytes and non-lymphoid cells (i.e. Ia-positive macrophages and dendritic cells), were newly transplanted into nude mice of strain A. The engraftment lead to immunological reconstitution and the nude mice acquired tolerance to the MHC antigens expressed by the allogeneic Ia-positive cells populating the chimaeric graft. Irradiation of the chimaeric thymus prior to transplantation allowed transplantation of chimaeric thymus devoid of living thymocytes but still populated with functionally intact Ia-positive non-lymphoid cells. Transplantation of irradiated chimaeric thymuses resulted in immunoreconstitution and induced exactly the same allotolerance pattern as described above. The results demonstrate that not thymus epithelial cells but a bone-marrow-derived non-lymphoid thymus cell, most likely the Ia-antigen-positive thymic macrophage of dendritic cell, is responsible for the induction of tolerance to MHC antigens in developing T lymphocytes.     

Tolerogenic dendritic cells and their role in transplantation

The pursuit of clinical transplant tolerance has led to enhanced understanding of mechanisms underlying immune regulation, including the characterization of immune regulatory cells, in particular antigen-presenting cells (APC) and regulatory T cells (Treg), that may play key roles in promoting operational tolerance. Dendritic cells (DC) are highly efficient APC that have been studied extensively in rodents and humans, and more recently in non-human primates. Owing to their ability to regulate both innate and adaptive immune responses, DC are considered to play crucial roles in directing the alloimmune response towards transplant tolerance or rejection. Mechanisms via which they can promote central and peripheral tolerance include clonal deletion, the induction of Treg, and inhibition of memory T cell responses. These properties have led to the use of tolerogenic DC as a therapeutic strategy to promote organ transplant tolerance. In rodents, infusion of donor- or recipient-derived tolerogenic DC can extensively prolong donor-specific allograft survival, in association with regulation of the host T cell response. In clinical transplantation, progress has been made in monitoring DC in relation to graft outcome, including studies in operational liver transplant tolerance. Although clinical trials involving immunotherapeutic DC for patients with cancer are ongoing, implementation of human DC therapy in clinical transplantation will require assessment of various critical issues. These include cell isolation and purification techniques, source, route and timing of administration, and combination immunosuppressive therapy. With ongoing non-human primate studies focused on DC therapy, these logistics can be investigated seeking the optimal approaches. The scientific rationale for implementation of tolerogenic DC therapy to promote clinical transplant tolerance is strong. Evaluation of technical and therapeutic logistic issues is an important next step prior to the application of tolerogenic DC in clinical organ transplantation.     

Mechanisms regulating the development and function of natural regulatory T cells

The key of the immune system is to protect the host from foreign threat posed by pathogens and from the internal threat posed by self-attacking lymphocytes. The ability to discriminate self versus non-self ensures that only “non-self” pathogens, but not the self antigens, are attacked. Such tolerance to “self” arises from the central tolerance mechanisms that include the deletion of thymocytes with high reactivity to self antigens and also the induction of unresponsiveness of autoreactive T cells in the periphery. Natural regulatory T cells (nTregs) directly inhibit effector T cells, and keep their proliferation in control. Apart from preventing autoimmune reactions, Tregs also contribute to peripheral immune homeostasis as evidenced by the excessive lymphocyte accumulation in peripheral lymphoid organs and intestinal inflammation in the absence of nTregs. Here we discuss the molecular aspects of the development and suppressive function of naturally occurring Tregs. Accumulating evidence shows the importance of these Tregs in autoimmunity, tumor immunity, organ transplantation, allergy, and microbial immunity.     

Fetal tolerance in human pregnancy--a crucial balance between acceptance and limitation of trophoblast invasion

During human pregnancy the semi-allogeneic/allogeneic fetal graft is normally accepted by the mother's immune system. Initially the contact between maternal and fetal cells is restricted to the decidua but during the 2nd trimester it is extended to the entire body. Two contrary requirements influence the extent of invasion of extravillous fetal trophoblast cells (EVT) in the maternal decidua: anchorage of the placenta to ensure fetal nutrition and protection of the uterine wall against over-invasion. To establish the crucial balance between tolerance of the EVT and its limitation, recognition of the semi-allogeneic/allogeneic fetal cell by maternal leukocytes is prerequisite. A key mechanism to limit EVT invasion is induction of EVT apoptosis. Apoptotic bodies are phagocytosed by antigen-presenting cells (APC). Peptides from apoptotic cells are presented by APC cells and induce an antigen-specific tolerance against the foreign antigens on EVT cells. These pathways, including up-regulation of the expression of IDO, IFNgamma and CTLA-4 as well as the induction of T(regulatory) cells, are general immunological mechanisms which have developed to maintain peripheral tolerance to self-antigens. Together these data suggest that the mother extends her "definition of self" for 9 months on the foreign antigens of the fetus.     

Allogeneic hematopoietic transplantation and natural killer cell recognition of missing self

Summary:  Although the optimal donor for allogeneic hematopoietic stem cell transplantation (HSCT) is a human leukocyte antigen-matched sibling, 75% of patients do not have a match, and alternatives are matched unrelated volunteers, unrelated umbilical cord blood units, and full-haplotype-mismatched family members. To cure leukemia, allogeneic HSCT relies on donor T cells in the allograft, which promote engraftment, eradicate malignant cells, and reconstitute immunity. Here, we focus on the open issues of rejection, graft-versus-host disease (GVHD), and infections and the benefits of natural killer (NK) cell alloreactivity and its underlying mechanisms. Donor-versus-recipient NK cell alloreactivity derives from a mismatch between inhibitory receptors for self-major histocompatibility complex (MHC) class I molecules on donor NK clones and the MHC class I ligands on recipient cells. These NK clones sense the missing expression of the self-MHC class I allele on the allogeneic targets and mediate alloreactions. HSCT from 'NK alloreactive' donors controls acute myeloid relapse without causing GVHD. We review the translation of NK cell recognition of missing self into the clinical practice of allogeneic hematopoietic transplantation and discuss how it has opened innovative perspectives in the cure of leukemia.     

A20 deletion in T cells modulates acute graft‐versus‐host disease in mice

The NF‐κB regulator A20 limits inflammation by providing negative feedback in myeloid cells and B cells. Functional lack of A20 has been linked to several inflammatory and autoimmune diseases. To define how A20 affects the functionality of T effector cells in a highly inflammatory environment, we performed conventional allogeneic hematopoietic stem cell transplantation (allo‐HSCT) with A20‐deficient CD4⁺ and CD8⁺ donor T cells in mice. Severity and mortality of graft‐versus‐host disease (GVHD) after allo‐HSCT was drastically reduced in recipients transplanted with conventional doses of A20‐deficient T cells. Consistently, we found that the A20‐deficient donor T‐cell compartment was strongly diminished at various timepoints after allo‐HSCT. However, proportionally more A20‐deficient donor T cells produced IFN‐γ and systemic inflammation was elevated early after allo‐HSCT. Consequently, increasing the dose of transplanted A20‐deficient T cells reversed the original phenotype and resulted in enhanced GVHD mortality compared to recipients that received A20^+/+ T cells. Still, A20‐deficient T cells, activated either through T cell receptor‐dependent or ‐independent mechanisms, were less viable than control A20^+/+ T cells, highlighting that A20 balances both, T‐cell activation and survival. Thus, our findings suggest that targeting A20 in T cells may allow to modulate T‐cell‐mediated inflammatory diseases like GVHD.     

Mechanisms of peripheral tolerance to allergens

The immune system is regulated to protect the host from exaggerated stimulatory signals establishing a state of tolerance in healthy individuals. The disequilibrium in immune regulatory vs effector mechanisms results in allergic or autoimmune disorders in genetically predisposed subjects under certain environmental conditions. As demonstrated in allergen‐specific immunotherapy and in the healthy immune response to high‐dose allergen exposure models in humans, T regulatory cells are essential in the suppression of Th2‐mediated inflammation, maintenance of immune tolerance, induction of the two suppressive cytokines interleukin‐10 and transforming growth factor‐β, inhibition of allergen‐specific IgE, and enhancement of IgG4 and IgA. Also, suppression of dendritic cells, mast cells, and eosinophils contributes to the construction of peripheral tolerance to allergens. This review focuses on mechanisms of peripheral tolerance to allergens with special emphasis on recent developments in the area of immune regulation.