In many experimental models, heart, pancreas and kidney allografts are accepted long-term following costimulation-targeting therapies, whereas skin, lung and intestine resist the induction of tolerance under the same regimens. We noted that a common feature of the resistant organs is their constant exposure to commensal microbes and hypothesized that these microorganisms may stimulate Toll-like receptors (TLRs), promote alloresponses and prevent tolerance induction. This hypothesis prompts the predictions that TLR engagement at the time of transplantation should avert tolerance to heart allografts in animals treated with costimulation-targeting therapies, whereas inhibition of TLR signaling should promote tolerance to skin allografts under the same conditions. Indeed, engagement of a single TLR was sufficient to prevent anti-CD154-mediated long-term cardiac allograft acceptance and correlated with abolished intragraft recruitment of CD4 + /FoxP3 + regulatory T cells and the development of linked-suppression. Conversely, a lack of donor and recipient MyD88-dependent signaling led to successful skin allograft acceptance in anti-CD154-treated animals. Thus, the status of TLR signaling contributes to the resistance versus susceptibility of organs to transplantation tolerance.

Exposure to certain viruses and parasites has been shown to prevent the induction of transplantation tolerance in mice via the generation of cross-reactive memory T cell responses or the induction of bystander activation. Bacterial infections are common in the perioperative period of solid organ allograft recipients in the clinic, and correlations between bacterial infections and acute allograft rejection have been reported. However, whether bacterial infections at the time of transplantation have any effect on the generation of transplantation tolerance remains to be established. We used the Gram-positive intracellular bacterium Listeria monocytogenes (LM) as a model pathogen because its effects on immune responses are well described. Perioperative LM infection prevented cardiac and skin allograft acceptance induced by anti-CD154 and donor-specific transfusion in mice. LM-mediated rejection was not due to the generation of cross-reactive T cells and was largely independent of signaling via MyD88, an adaptor for most TLRs, IL-1, and IL-18. Instead, transplant rejection following LM infection was dependent on the expression of the phagosome-lysing pore former listeriolysin O and on type I IFN receptor signaling. Our results indicate that bacterial exposure at the time of transplantation can antagonize tolerogenic regimens by enhancing alloantigen-specific immune responses independently of the generation of cross-reactive memory T cells.

Classic hyperacute rejection is dependent on the activation of the terminal components of complement. Recently, xenoantibodies with limited abilities to activate the classical pathway of complement in vitro have been implicated in the acute vascular rejection of xenografts. It is unclear how these Abs affect their pathogenic activities in vivo. In this study, we demonstrate the ability of an anti-Gal-α1,3Gal (Gal) IgG1, with modest complement-activating abilities in vitro, to induce xenograft rejection. This rejection was dependent on the activation of complement, on FcγR-mediated interactions, and on the presence of NK cells. Inhibition of any one of these factors resulted in the abrogation of IgG1-mediated rejection. In contrast, an anti-Gal IgG3 mAb induced classic, hyperacute rejection that was solely dependent on complement activation. Our observations implicate two types of IgG-mediated rejection; one that is dependent on complement activation, and a second that is uniquely dependent on complement, FcγR, and NK cells.

The impact of memory B cells and alloantibodies on the ability to induce transplantation tolerance has not been elucidated. We have developed a murine heart transplant model that isolates the contributions of functional memory B cells from memory T cells in allograft rejection. Memory 3-83 B cells with dual specificity for H-2Kk and H-2Kb were generated in 3-83 Igi BCR knockin (BALB/c background) mice by the transplantation of C3H (H-2Kk) hearts in the absence of immunosuppression. To test the effect of functional memory 3-83 B cells, C3H-primed 3-83 Igi recipients were challenged with C57BL/6 hearts (H-2Kb) at 60–90 days post-C3H heart transplant and treated with anti-CD154 mAbs. Despite immunosuppression, the C57BL/6 hearts were acutely rejected within 10–13 days and graft rejection was associated with increased frequencies of C57BL/6-specific IFN-γ-producing T cells. Histology revealed significant numbers of infiltrating T cells, consistent with acute T cell-mediated rejection. The resistance to tolerance induction was dependent on the synergistic effects of memory 3-83 B cells and alloantibodies, whereas memory T cells are not necessary. We conclude that the combined effects of functional memory B cells and alloantibodies prevent anti-CD154-mediated graft acceptance by facilitating the CD40-CD154-independent activation of alloreactive T cells. This study provides insight into the potential ability of memory B cells and alloantibodies to prevent anti-CD154-mediated graft acceptance.

No abstract

FK778 has both immune suppressive and antiviral activities, neither of which is entirely dependent on inhibition of pyrimidine synthesis. These, and other published observations, suggest that the antiviral activity and a considerable part of the efficacy of the malononitrilamide family of drugs is attributable to activities other than drug induced pyrimidine deficiency.

Anti-graft antibodies are often associated with graft rejection. Under special conditions, grafts continue to function normally even in the presence of anti-graft antibodies and complement. This condition is termed accommodation. We developed a xenograft accommodation model in which baby Lewis rat hearts are transplanted into Rag/GT-deficient mice, and accommodation is induced by repeated i.v. injections of lowdose anti-a -Gal IgG 1 . The accommodated grafts survived a bolus dose of anti-a -Gal IgG 1 , while freshly transplanted second grafts were rejected. To study the mechanism of anti-a -Gal IgG 1 -mediated accommodation, both real-time PCR and immunohistochemical staining revealed elevated expression of DAF, Crry and CD59 in the accommodated grafts. In vitro exposure of rat endothelial cells to anti-a -Gal IgG 1 also induced the up-regulation of DAF, Crry and CD59, as revealed by Western blot analyses, and was associated with an acquired resistance to antibody and complementmediated lysis in vitro. Collectively, these studies suggest that the up-regulation of complement regulatory proteins may abrogate complement-mediated rejection and permit the development of xenograft accommodation.

Contrary to previous reports of the ability of bone marrow cells to induce central deletional tolerance, our data suggest that the regimen involving co-transplantation of IAB on the day of heart allograft transplantation and transient anti-CD40L therapy induces a robust donor-specific peripheral tolerance.