Evidence for immune responses to a self-antigen in lung transplantation: role of type V collagen-specific T cells in the pathogenesis of lung allograft rejection

We have reported that lung allograft rejection involves an immune response to a native protein in the lung, type V collagen (col(V)), and that col(V)-induced oral tolerance prevented acute and chronic rejection. In support of these findings col(V) fragments were detected in allografts during rejection, but not in normal lungs. The purpose of the current study was to isolate and characterize col(V)-specific allograft-infiltrating T cells and to determine their contribution to the rejection response in vivo. Two col(V)-specific T cell lines, LT1 and LT3, were isolated from F344 (RT1(lv1)) rat lung allografts during rejection that occurred after transplantation into WKY (RT1(l)) recipients. Both cell lines, but not normal lung lymphocytes, proliferated in response to col(V). Neither LT1 nor LT3 proliferated in response to alloantigens. LT1 and LT3 were CD4(+)CD25(-) and produced IFN-gamma in response to col(V). Compared with normal CD4(+) T cells, both cell lines expressed a limited V-beta TCR repertoire. Each cell strongly expressed V-beta 9 and 16, but differed in expression of other V-betas. Adoptive transfer of each cell line did not induce pathology in lungs of normal WKY rats. In contrast, adoptive transfer of LT1, but not LT3, caused marked peribronchiolar and perivascular inflammation in isograft (WKY) lungs and abrogated col(V)-induced oral tolerance to allograft (F344) lungs. Collectively, these data show that lung allograft rejection involves both allo- and autoimmune responses, and graft destruction that occurs during the rejection response may expose allograft-infiltrating T cells to potentially antigenic epitopes in col(V).     

Epitope Analysis of the Collagen Type V-Specific T Cell Response in Lung Transplantation Reveals an HLA-DRB1*15 Bias in Both Recipient and Donor

Background

IL-17-dependent cellular immune responses to the α1 chain of collagen type V are associated with development of bronchiolitis obliterans syndrome after lung transplantation, and with idiopathic pulmonary fibrosis and coronary artery disease, primary indications for lung or heart transplantation, respectively.

Methodology/Principal Findings

We found that 30% of the patients awaiting lung transplantation exhibited a strong cell-mediated immune response to col(V). Of these, 53% expressed HLA-DR15, compared to a 28% HLA-DR15 frequency in col(V) low-responders (p=0.02). After transplantation, patients with HLA-DR1 and -DR17, not -DR15, developed anti-col(V) responses most frequently (p=0.04 and 0.01 vs. controls, respectively). However, recipients of a lung from an HLA-DR15⁺ donor were at significantly elevated risk of developing anti-col(V) responses (p=0.02) and BOS (p=0.03). To determine the molecular basis of this unusual pattern of DR allele bias, a peptide library comprising the collagenous region of the α1(V) protein was screened for binding to HLA-DR0101, -DR1501, -DR0301 (DR17) or to HLA-DQ2 (DQA1*0501: DQB1*0201; in linkage disequilibrium with -DR17) and -DQ6 (DQA1*0102: DQB1*0602; linked to -DR15). Eight 15-mer peptides, six DR-binding and two DQ-binding, were identified. HLA-DR15 binding to two peptides yielded the highest binding scores: 650 (where 100 = positive control) for p799 (GIRGLKGTKGEKGED), and 193 for p1439 (LRGIPGPVGEQGLPG). These peptides, which also bound weakly to HLA-DR1, elicited responses in both HLA-DR1⁺ and -DR15⁺ col(V) reactive hosts, whereas binding and immunoreactivity of p1049 (KDGPPGLRGFPGDRG) was DR15-specific. Remarkably, a col(V)-reactive HLA-DR1⁺DR15^neg lung transplant patient, whose donor was HLA-DR15⁺, responded not only to p799 and p1439, but also to p1049.

Conclusions/Significance

HLA-DR15 and IPF disease were independently associated with pre-transplant col(V) autoimmunity. The increased risk of de novo immunity to col(V) and BOS, associated with receiving a lung transplant from an HLA-DR15⁺ donor, may result from presentation by donor-derived HLA- DR15, of novel self-peptides to recipient T cells.     

Aging Impairs Recipient T Cell Intrinsic and Extrinsic Factors in Response to Transplantation

Background

As increasing numbers of older people are listed for solid organ transplantation, there is an urgent need to better understand how aging modifies alloimmune responses. Here, we investigated whether aging impairs the ability of donor dendritic cells or recipient immunity to prime alloimmune responses to organ transplantation.

Principal Findings

Using murine experimental models, we found that aging impaired the host environment to expand and activate antigen specific CD8⁺ T cells. Additionally, aging impaired the ability of polyclonal T cells to induce acute allograft rejection. However, the alloimmune priming capability of donor dendritic cells was preserved with aging.

Conclusion

Aging impairs recipient responses, both T cell intrinsic and extrinsic, in response to organ transplantation.     

Mechanism of thyroid allograft rejection.

Balb/c thyroids, held in organ culture for 26 days, survive and function as well as isografts for greater than 100 days in CBA recipients. Uncultured allografts are totally rejected by 20 days after transplantation. Prolonged allograft survival can also be achieved by the treatment of donor animals with cyclophosphamide prior to harvesting tissues for transplantation. These allografts do not survive as well as 26 day cultured allografts, but cyclophosphamide pretreatment reduces the culture time required to achieve indefinite survival to 7 days. The provision of an allogeneic (LD) stimulus by thyroid tissue that is I-region incompatible with the host does not facilitate the rejection of a tolerated cultured allograft. However, activation of the host immune system by an uncultured graft syngeneic to a tolerated cultured allograft leads to the chronic rejection of the cultured transplant. The transfer of a tolerated cultured allograft back to its strain of origin induces an acute inflammatory reaction that causes tissue damage within the transplant but does not lead to the total destruction of the tissue.     

Manipulating IL-2 Availability Amid Presentation of Donor MHC Antigens Suppresses Murine Alloimmune Responses by Inducing Regulatory T Cells

Background

Major histocompatibility complex (MHC) antigens are important for alloimmune responses as well as immune tolerance. Previous studies have shown that presentation of donor MHC antigens by donor-specific transfusion prior to or upon transplantation promotes transplant tolerance induced by other agents. However, it is unclear whether presentation of donor MHC antigens by DNA vaccination induces long-term allograft survival.

Methodology/Principal Findings

We investigated whether presentation of MHC class-II and/or class-I donor antigens by DNA vaccination suppresses alloimmune responses and promotes long-term allograft acceptance. We initially found that presentation of both MHC donor antigens by DNA vaccination itself prior to transplantation fails to significantly prolong islet allograft survival in otherwise untreated mice. However, islet allograft survival was significantly prolonged when MHC class-II DNA vaccination was accompanied with IL-2 administration (MHCII + IL-2) while MHC class-I DNA vaccination was followed by IL-2 and subsequent neutralizing anti-IL-2 treatments (MHCI + IL-2/anti-IL-2). Especially, this protocol promoted long-term allograft survival in the majority of recipients (57%) when combined with low doses of rapamycin post-transplantation. Importantly, MHCII + IL-2 induced FoxP3+ Treg cells in both spleens and grafts and suppressed graft-infiltrating CD4+ cell proliferation, whereas MHCI + IL-2/anti-IL-2 mainly inhibited graft-infiltrating CD8+ cell proliferation and donor-specific CTL activity. The combined protocol plus rapamycin treatment further reduced both CD4+ and CD8+ T cell proliferation as well as donor-specific CTL activity but spared FoxP3+ Treg cells. Depleting CD25+ Treg cells or adoptive transfer of pre-sensitized CD8+ T cells abolished this long-term allograft survival.

Conclusions/Significance

Manipulating IL-2 availability during presentation of MHC class-II and class-I donor antigens by DNA vaccination pre-transplantation induces Treg cells, suppresses alloimmune responses and promotes long-term allograft survival.     

Phenotype frequencies of autosomal minor histocompatibility antigens display significant differences among populations

Minor histocompatibility (H) antigens are allogeneic target molecules having significant roles in alloimmune responses after human leukocyte antigen–matched solid organ and stem cell transplantation (SCT). Minor H antigens are instrumental in the processes of transplant rejection, graft-versus-host disease, and in the curative graft-versus-tumor effect of SCT. The latter characteristic enabled the current application of selected minor H antigens in clinical immunotherapeutic SCT protocols. No information exists on the global phenotypic distribution of the currently identified minor H antigens. Therefore, an estimation of their overall impact in human leukocyte antigen–matched solid organ and SCT in the major ethnic populations is still lacking. For the first time, a worldwide phenotype frequency analysis of ten autosomal minor H antigens was executed by 31 laboratories and comprised 2,685 randomly selected individuals from six major ethnic populations. Significant differences in minor H antigen frequencies were observed between the ethnic populations, some of which appeared to be geographically correlated.     

Thioredoxin Priming Prolongs Lung Allograft Survival by Promoting Immune Tolerance

Tolerance to allograft antigen is the major challenge and final goal of transplant medicine. Our previous study demonstrated that thioredoxin-1 (Trx) priming of donor lung significantly protected allogeneic lung graft. To determine whether Trx priming of donor lung inhibits allograft rejection, extends allograft survival and induces immune tolerance, orthotopic left lung transplantation was performed from Lewis to Sprague-Dawley rats without immunosuppression. Donor lungs were primed with Trx at 4°C for 4 hr prior to transplantation. After up to 37 days post-transplantation, allograft lung morphology, recipient T cell and humoral alloantigen-specific immune responses were examined. We found that Trx-primed lungs exhibited much reduced acute rejection and associated lung injuries resulting in loss of graft functional area at 5-37 days post-transplant in contrast to the control groups. CD4⁺ T cells from the recipients with Trx-primed grafts responded to the stimulation of dendritic cells (DCs) of donor origin, in contrast to DCs from the third party, with significantly reduced proliferation. Consistent with above findings, we observed that CD4⁺Foxp3⁺ regulatory T cells in spleen cells from the recipients with Trx-primed grafts were significantly increased compared to controls, and CD4⁺ T cells from the recipients with Trx-primed grafts produced much higher levels of immunosuppressive cytokine, IL-10 when stimulated with allogeneic donor DCs. In addition, humoral immune tolerance was also induced as there was no significant increase levels of serum antibodies against donor antigens in Trx-lung recipients when re-challenged with allogeneic donor antigens. Our results demonstrate that one-time Trx-priming of donor lung grafts prior to transplantation significantly prolongs the survival of the grafts through inducing or promoting cellular and humoral alloantigen-specific immune tolerance, which might be associated with the induction of immunosuppressive regulatory T cells.     

Treatment with cyclosporin and risks of graft rejection in male kidney and heart transplant recipients with non-O blood.

In a consecutive series of 146 kidney transplant recipients treated with cyclosporin A a strong correlation between matching for the HLA-A, HLA-B, and HLA-DR loci specificities and outcome of the grafts was observed in male recipients with non-O blood groups. Such a beneficial effect of matching was not found in female patients or male patients with blood group O. In these patients survival of the grafts at one year was good irrespective of the number of HLA-A, B, and DR mismatches. Also in 47 male heart transplant recipients immune responsiveness against mismatched HLA antigens was related to blood group. A significantly higher incidence of rejection episodes was observed in male patients with non-O blood groups (n = 32) than in those with blood group O (n = 15). Matching for HLA-DR reduced the number of acute rejection episodes in male patients with non-O blood. These findings may help explain the controversial reports about the importance of HLA matching in organ transplantation. Furthermore, as most candidates for heart transplantation are male and not of blood group O, the higher incidence of graft rejection in these patients underscores the need for an exchange strategy of donor hearts.     

Analysis of chronic lung transplant rejection by MALDI-TOF profiles of bronchoalveolar lavage fluid

While lung transplant is an effective therapy for advanced lung disease, chronic allograph rejection remains a primary basis for lower survival rates than those for other solid organ transplants. This study used carefully controlled Zip-Tip extraction of bronchoalveolar lavage fluid (BALF) followed by MALDI-TOF MS to identify biomarkers of chronic lung transplant rejection. Many differences were observed between controls, those who did not develop chronic rejection within 100 months, and patients who had developed chronic rejection, diagnosed as bronchiolitis obliterans syndrome (BOS). Intensity ratios of peaks within the same MALDI-TOF profile were used to quantify the result. One of the best identifiers of BOS was a lowered ratio of clara cell protein (CCP m/z = 15,835) to lysozyme (m/z = 14,700), which gave 94% specificity and 74% sensitivity for diagnosis. Furthermore, low values for CCP/Lysozyme (<0.3) were observed in 66% of samples taken at 1 to 15 months prior to the diagnosis of BOS. Many other components of the profile gave similar or better outcomes for diagnosis but tended to be less valuable for the prediction of future disease. Overall, this study demonstrated the feasibility of this approach for the detection of disease biomarkers.     

Immature and mature CD8alpha+ dendritic cells prolong the survival of vascularized heart allografts.

CD8alpha+ and CD8alpha- dendritic cells (DCs) arise from committed bone marrow progenitors and can induce or regulate immune reactivity. Previously, the maturational status of CD8alpha-(myeloid) DCs has been shown to influence allogeneic T cell responses and allograft survival. Although CD8alpha+ DCs have been implicated in central tolerance and found to modulate peripheral T cell function, their influence on the outcome of organ transplantation has not been examined. Consistent with their equivalent high surface expression of MHC and costimulatory molecules, sorted mature C57BL/10J (B10; H2(b)) DCs of either subset primed naive, allogeneic C3H/HeJ (C3H; H2(k)) recipients for Th1 responses. Paradoxically and in contrast to their CD8alpha- counterparts, mature CD8alpha+ B10 DCs given systemically 7 days before transplant markedly prolonged B10 heart graft survival in C3H recipients. This effect was associated with specific impairment of ex vivo antidonor T cell proliferative responses, which was not reversed by exogenous IL-2. Further analyses of possible underlying mechanisms indicated that neither immune deviation nor induction of regulatory cells was a significant contributory factor. In contrast to the differential capacity of the mature DC subsets to affect graft outcome, immature CD8alpha+ and CD8alpha- DCs administered under the same experimental conditions significantly prolonged transplant survival. These observations demonstrate for the first time the innate capacity of CD8alpha+ DCs to regulate alloimmune reactivity and transplant survival, independent of their maturation status. Mobilization of such a donor DC subset with capacity to modulate antidonor immunity may have significant implications for the therapy of allograft rejection.     

Differential expression of Smad7 transcripts identifies the CD4+CD45RChigh regulatory T cells that mediate type V collagen-induced tolerance to lung allografts

Regulatory T cells (Tregs) induced by oral tolerance may suppress immunity by production of TGF-beta that could also enhance Treg activity. However, all cells that are phenotypically Tregs in rats (CD4(+)CD45RC(high)-RC(high)) may not have regulatory function. Because Smad7 expression in T cells is associated with inflammation and autoimmunity, then lack of Smad7 may identify those cells that function as Tregs. We reported that feeding type V collagen (col(V)) to WKY rats (RT1(l)) induces oral tolerance to lung allografts (F344-RT1(lvl)) by T cells that produce TGF-beta. The purpose of the current study was to identify the Tregs that mediate col(V)-induced tolerance, and determine Smad7 expression in these cells. RC(high) cells from tolerant rats were unresponsive to allogeneic stimulation and abrogated rejection after adoptive transfer. In contrast, CD4(+)CD45RC(low) (RC(low)) cells from tolerant rats and RC(high) or RC(low) cells from normal rats or untreated allograft recipients proliferated vigorously in response to donor Ags, and did not suppress rejection after adoptive transfer. TGF-beta enhanced proliferation in response to col(V) presented to tolerant RC(high), but not other cells. In contrast to other cells, only RC(high) cells from tolerant rats did not express Smad7. Collectively, these data show that the Tregs that mediate col(V)-induced tolerance to lung allografts do not express SMAD7 and, therefore, are permissive to TGF-beta-mediated signaling.     

The generation of CD25+ CD4+ regulatory T cells that prevent allograft rejection does not compromise immunity to a viral pathogen

In all but a small minority of cases, continued survival of solid organ grafts after transplantation depends on lifelong, nonselective immunosuppression that, although effective, results in increased rates of infection, cancer, and vascular disease. Therapeutic strategies that engage or mimic self-tolerance may allow prolonged allograft survival without the disadvantages of nonspecific immunotherapy. Pretreatment of recipient mice with donor alloantigen combined with transient modulation of the peripheral T cell pool with anti-CD4 Ab leads to the indefinite survival of MHC-incompatible cardiac allografts without further therapy. Tolerance is dependent on CD25+ CD4+ regulatory T cells that arise from naive CD25- precursors and regulate rejection via both IL-10 and CTLA-4. Although these cells are clearly effective at controlling rejection, the proven ability of recently activated CD25+ cells to mediate bystander regulation raises the possibility that tolerized individuals might also have a reduced capacity to respond to environmental pathogens. We have examined anti-influenza responses in tolerized primary heart recipients, secondary recipients following adoptive transfer of regulatory populations, and tolerized mice in which bystander regulation has been deliberately induced. Neither virus-specific CTL activity in vitro nor the clearance of virus in vivo was significantly diminished in any of these treatment groups compared with infected unmanipulated controls. The data suggest that the induction of dominant allograft tolerance dependent on regulatory T cells does not necessarily result in attenuated responses to pathogens providing further support for the development of tolerance induction protocols in clinical transplantation.     

On the acceptability of xenografts

Transplantation represents a major advance in modern medicine with a major impact on the interactions between individuals and society. The numbers of patients undergoing organ transplantation increased steadily over the years and around 250,000 individuals are living nowadays in Europe with a transplanted organ. On the other hand, the numbers of cadaveric (brain-dead) donors used for organ transplantation remains stable, at around 5,000 each year, and the numbers of transplantation from living donors only slowly increase in Europe. Therefore, a gap is growing between the numbers of patients in need of a transplant and the numbers of organs available for transplantation. About 45,000 patients are currently on renal transplant waiting lists in Europe and, depending on the countries considered, 15 to 30 % of candidates for liver or heart transplantation die before a life-saving transplant becomes available to them. There is therefore an urgent need to implement innovative research and to take full advantage of recent biotechnological advances to explore new avenues in xenotransplantation, and to simultaneously address the ethical, societal and public health issues related to organ replacement. Much progresses have been accomplished in the understanding of xenograft rejection processes that include hyperacute, acute vascular and cellular rejection mechanisms. Strategies to promote xenograft survival that are currently under evaluation include genetic engineering of donor pigs, adapted immunosuppressive treatments and tolerance induction. Also, the psychological acceptance has been evaluated.     

Molecular mechanisms of immunity in corneal allotransplantation and xenotransplantation

Corneal allotransplantation is the most common and successful form of solid organ transplantation in humans. In uncomplicated cases, the two-year graft survival rate is over 90%. This extraordinary success can be attributed in part to various features of the normal cornea and anterior segment that together account for their 'immune-privileged' status. However, despite this success, a significant number of corneal grafts fail and immunological rejection remains by far the leading cause of graft failure. Studies on animal models of corneal transplantation have yielded a wealth of information on the molecular and cellular features of graft rejection, and have established that this process is mediated primarily by CD4+ T cells of the T helper 1 (Th1) phenotype. In addition, studies have elucidated that certain facets of allosensitisation differ between corneal and other solid organ transplants. On the basis of these findings, novel experimental strategies selectively targeting the afferent or efferent arms of corneal alloimmunity have provided promising results in preventing corneal allograft rejection in the laboratory. Finally, because of the global shortage of human donor corneas, there is currently renewed interest in the possibility of using corneas from other species for transplantation into human eyes (xenotransplantation). Preliminary studies on animal models of corneal xenotransplantation have documented both antibody-mediated and cell-mediated responses that might play important roles in the accelerated rejection observed in corneal xenotransplants. This review synthesises the principal concepts emerging from studies of the molecular mechanisms in corneal transplant immunology.     

The role of the CD134-CD134 ligand costimulatory pathway in alloimmune responses in vivo

The CD134-CD134 ligand (CD134L) costimulatory pathway has been shown to be critical for both T and B cell activation; however, its role in regulating the alloimmune response remains unexplored. Furthermore, its interactions with other costimulatory pathways and immunosuppressive agents are unclear. We investigated the effect of CD134-CD134L pathway blockade on allograft rejection in fully MHC-mismatched rat cardiac and skin transplantation models. CD134L blockade alone did not prolong graft survival compared with that of untreated recipients, and in combination with donor-specific transfusion, cyclosporine, or rapamycin, was less effective than B7 blockade in prolonging allograft survival. However, in combination with B7 blockade, long-term allograft survival was achieved in all recipients (>200 days). Moreover, this was synergistic in reducing the frequency of IFN-gamma-producing alloreactive lymphocytes and inhibiting the generation of activated/effector lymphocytes. Most impressively, this combination prevented rejection in a presensitized model using adoptive transfer of primed lymphocytes into athymic heart transplant recipients. In comparison to untreated recipients (mean survival time (MST): 5.3 +/- 0.5 days), anti-CD134L mAb alone modestly prolonged allograft survival (MST: 14 +/- 2.8 days) as did CTLA4Ig (MST: 21.5 +/- 1.7 days), but all grafts were rejected within 24 days. Importantly, combined blockade further and significantly prolonged allograft survival (MST: 75.3 +/- 12.7 days) and prevented the expansion and/or persistence of primed/effector alloreactive T cells. Our data suggest that CD134-CD134L is a critical pathway in alloimmune responses, especially recall/primed responses, and is synergistic with CD28-B7 in mediating T cell effector responses during allograft rejection. Understanding the mechanisms of collaboration between these different pathways is important for the development of novel strategies to promote long-term allograft survival.     

HLA-DRBl and susceptibility to kidney allograft rejection in Southern Iranian patients

Kidney transplantation is the best treatment option for the patients with end-stage renal disease. Viral infections and genetic factors such as HLA-II antigens may affect the kidney transplant outcome. The compatibility of HLA-DRB1 molecules in the survival of kidney transplant is important. Also, the correlation between these molecules and viral infections is significant. The current study investigates the allele frequency of HLA-DRB1 in 41 recipient kidney transplant and 203 normal healthy controls by polymerase chain reaction using sequence specific primers. Moreover the relation between HLA-DRB1 allelic groups and hepatitis B, hepatitis C and cytomegalovirus viral infections was also studied. However statistical analysis of the allele frequencies didn’t show any significant association between HLA-DRB1 allelic group distributions or sharing and susceptibility to acute kidney transplant rejection (P > 0.05). Comparing the allele frequencies between HLA-DRB1*14 and DRB1*04 allelic showed a significant difference in controls and patients (P = 0.03 and P = 0.05 respectively). The results of the present study also showed a significant association between possession of HLA-DRB1*07 allele in kidney transplant recipients and hepatitis C virus infection (P = 0.009). In conclusion however the results of the present study did not showed relation between HLA-DRB1 allele’s frequencies or sharing and kidney transplantation outcome, the results indicated that HLA-DRB1 alleles may susceptible individuals to renal disease or play a role in susceptibility to viral infection in kidney transplant patients.     

Early chemokine cascades in murine cardiac grafts regulate T cell recruitment and progression of acute allograft rejection

The identification of early inflammatory events after transplant in solid tissue organ grafts that may direct T cell recruitment and promote acute allograft rejection remain largely unknown. To better understand temporal aspects of early inflammatory events in vascularized organ grafts, we tested the intragraft expression of four different chemokines in heterotopically transplanted A/J (H-2(a)) and syngeneic heart grafts in C57BL/6 (H-2(b)) recipient mice from 1.5 to 48 h after transplant. Similar temporal expression patterns and equivalent levels of chemokine expression were observed in both syngeneic and allogeneic cardiac allografts during this time period. Expression of the neutrophil chemoattractant growth-related oncogene alpha (KC) was observed first and reached peak levels by 6 h after transplant and was followed by the monocyte/macrophage chemoattractant protein-1 (JE) and then macrophage inflammatory proteins 1beta and 1alpha. Administration of rabbit KC antiserum to allograft recipients within 30 min of cardiac transplantation attenuated downstream events including intra-allograft expression of the T cell chemoattractants IFN-gamma-inducible protein-10 and monokine induced by IFN-gamma, cellular infiltration into the allograft, and graft rejection. Similarly, depletion of recipient neutrophils at the time of transplantation significantly extended allograft survival from day 8 to 10 in control-treated recipients up to day 21 after transplant. These results indicate the induction of highly organized cascades of neutrophil and macrophage chemoattractants in cardiac grafts and support the proposal that early inflammatory events are required for optimal recruitment of T cells into allografts during the progression of acute rejection of cardiac allografts.     

Contributions of direct and indirect alloresponses to chronic rejection of kidney allografts in nonhuman primates

The relative contribution of direct and indirect allorecognition pathways to chronic rejection of allogeneic organ transplants in primates remains unclear. In this study, we evaluated T and B cell alloresponses in cynomolgus monkeys that had received combined kidney/bone marrow allografts and myeloablative immunosuppressive treatments. We measured donor-specific direct and indirect T cell responses and alloantibody production in monkeys (n = 5) that did not reject their transplant acutely but developed chronic humoral rejection (CHR) and in tolerant recipients (n = 4) that never displayed signs of CHR. All CHR recipients exhibited high levels of anti-donor Abs and mounted potent direct T cell alloresponses in vitro. Such direct alloreactivity could be detected for more than 1 y after transplantation. In contrast, only two of five monkeys with CHR had a detectable indirect alloresponse. No indirect alloresponse by T cells and no alloantibody responses were found in any of the tolerant monkeys. Only one of four tolerant monkeys displayed a direct T cell alloresponse. These observations indicate that direct T cell alloresponses can be sustained for prolonged periods posttransplantation and result in alloantibody production and chronic rejection of kidney transplants, even in the absence of detectable indirect alloreactivity.     

Inhibiting CXCR3-dependent CD8+ T cell trafficking enhances tolerance induction in a mouse model of lung rejection

Lung transplantation remains the only effective therapy for patients with end-stage pulmonary diseases. Unfortunately, acute rejection of the lung remains a frequent complication and is an important cause of morbidity and mortality. The induction of transplant tolerance is thought to be dependent, in part, on the balance between allograft effector mechanisms mediated by effector T lymphocytes (Teff), and regulatory mechanisms mediated by FOXP3(+) regulatory T cells (Treg). In this study, we explored an approach to tip the balance in favor of regulatory mechanisms by modulating chemokine activity. We demonstrate in an adoptive transfer model of lung rejection that CXCR3-deficient CD8(+) Teff have impaired migration into the lungs compared with wild-type Teff, which results in a dramatic reduction in fatal pulmonary inflammation. The lungs of surviving mice contained tolerized CXCR3-deficient Teff, as well as a large increase in Treg. We confirmed that Treg were needed for tolerance and that their ability to induce tolerance was dependent on their numbers in the lung relative to the numbers of Teff. These data suggest that transplantation tolerance can be achieved by reducing the recruitment of some, but not necessarily all, CD8(+) Teff into the target organ and suggest a novel approach to achieve transplant tolerance.     

PDL1 is required for peripheral transplantation tolerance and protection from chronic allograft rejection

The PD-1:PDL pathway plays an important role in regulating alloimmune responses but its role in transplantation tolerance is unknown. We investigated the role of PD-1:PDL costimulatory pathway in peripheral and a well established model of central transplantation tolerance. Early as well as delayed blockade of PDL1 but not PDL2 abrogated tolerance induced by CTLA4Ig in a fully MHC-mismatched cardiac allograft model. Accelerated rejection was associated with a significant increase in the frequency of IFN-gamma-producing alloreactive T cells and expansion of effector CD8(+) T cells in the periphery, and a decline in the percentage of Foxp3(+) graft infiltrating cells. Similarly, studies using PDL1/L2-deficient recipients confirmed the results with Ab blockade. Interestingly, while PDL1-deficient donor allografts were accepted by wild-type recipients treated with CTLA4Ig, the grafts developed severe chronic rejection and vasculopathy when compared with wild-type grafts. Finally, in a model of central tolerance induced by mixed allogeneic chimerism, engraftment was not abrogated by PDL1/L2 blockade. These novel data demonstrate the critical role of PDL1 for induction and maintenance of peripheral transplantation tolerance by its ability to alter the balance between pathogenic and regulatory T cells. Expression of PDL1 in donor tissue is critical for prevention of in situ graft pathology and chronic rejection.