Andrea Deiwick

The functional relevance of passenger leukocytes and microchimerism for heart allograft acceptance in the rat.

With an organ transplant, hematopoietic donor cells are transferred to the recipient. To study the relevance of the resulting microchimerism for allograft acceptance, we analyzed a rat model of cyclosporine-induced tolerance for strongly incompatible heart allografts. Using a monoclonal antibody that detects a donor-specific CD45 allotype (RT7a), we selectively depleted donor leukocytes at different times after transplantation (days 0 or 18). Depletion was similarly effective at both times. However, only depletion on day 0 prevented tolerance induction and was associated with severe acute or chronic graft rejection. This indicates that passenger leukocytes have an essential immunomodulatory effect on the induction phase of allograft acceptance.

Background and prognostic implications of perireperfusion tissue injuries in human liver transplants: A panel histochemical study

BACKGROUND Hepatic graft reperfusion is associated with inflammatory processes of unknown relevance to the fate of graft. This study aimed to clarify this relevance by histochemical analyses of human hepatic grafts. METHODS Paired tissue samples were taken at the end of cold preservation and 2 hr after reperfusion (n=39). From six additional grafts, biopsies were performed at the end of cold preservation only. Injury or inflammatory markers of sinusoidal endothelium (von Willebrand factor-related antigen [vWF]), Kupffer cells (25F9), platelets (CD62), neutrophil leukocytes (CD11b), interleukin (IL)-1beta, intercellular adhesion molecule (ICAM)-1, and HLA-DR were evaluated semiquantitatively by indirect immunoperoxidase staining. Steatosis was also evaluated by hematoxylin and eosin staining. RESULTS vWF, CD62+ platelet aggregation, CD11b+ leukocytes, and IL-1beta levels increased after reperfusion, and these levels correlated with prereperfusion levels. Not only vWF, CD62+ platelets, CD11b+ leukocytes, IL-1beta, ICAM-1, and steatosis after reperfusion, but also IL-1beta, ICAM-1, and steatosis before reperfusion correlated with postoperative peak transaminase. Furthermore, vWF, CD11b+ leukocytes, 25F9+ macrophages, and ICAM-1 after reperfusion were associated with primary graft nonfunction and strong expressions of ICAM-1 or HLA-DR with early acute rejection. Although some markers (IL-1beta, CD62+ platelets, and CD11b+ leukocytes) correlated with preharvesting parameters (donor age or length of intensive care unit stay), none showed any significant correlation with cold preservation. CONCLUSION Synergistic inflammatory events in the hepatic graft at reperfusion, which have a significant impact on the later clinical course, are largely defined and precipitated by injury or activation of nonparenchymal cells preceding reperfusion or even graft harvesting.

Specific immunosuppression by postoperative infusion of allogeneic spleen cells: requirement of donor major histocompatibility complex expression and graft-versus-host reactivity.

BACKGROUND Donor leukocytes may exert positive immunoregulatory effects on allograft acceptance. Most recent studies have focused on pretreatment protocols. In this study, the effect of postoperative infusion of donor leukocytes on graft survival and the phenotypic and functional requirements for infused cells were investigated in fully major histocompatibility complex (MHC)-mismatched rat heart transplant models. METHODS LEW (RT1l) heart grafts were implanted heterotopically into abdomens of LEW.1W (RT1u), and different types of cells were infused postoperatively. Immunohistochemistry was used to evaluate histopathological changes of grafts. RESULTS In the absence of any immunosuppressive agents, a single dose of viable donor spleen cells (SC), but not bone marrow cells, was able to prolong heart allograft survival to about 21 days, while they were rejected promptly at day 7 in controls. Infusion of T cell-depleted donor SC, irradiated donor SC or third-party (BN) SC showed no effect on graft survival. Compared with resting cells, neither in vitro nor in vivo prestimulation of infused donor SC improved graft survival. Clinical signs of graft-versus-host reaction were not observed in all above groups. Histology showed remarkable reduction in the severity of graft infiltrate and interleukin-2 receptor-positive cells in grafts of cell-treated animals. Postoperative infusion of SC of F1 generation between different strain combinations showed two requirements for infused cells to be effective: (1) expression of donor-type MHC antigens and (2) strong alloreactivity against the host MHC antigens. CONCLUSION Postoperative infusion of viable donor SC can lead to allospecific down-regulation of alloreactivity by a graft-versus-host-associated effect.

Delayed low-level calcineurin inhibition promotes allospecific tolerance induction by posttransplantation donor leukocyte infusion

Background. Donor lymphocytes infused after organ transplantation can have strong immunoregulatory effects. Application of such protocols for transplant tolerance induction in a clinical setting will, however, require combination of specific immunomodulatory strategies with nonspecific immunosuppressive medication for safety reasons. The aim of this study was to analyze the effects of immunosuppressive treatment on tolerance induction protocols by posttransplantation donor lymphocyte infusion. Methods. The interaction of postoperative donor leukocyte infusion with different types, dosage, and timing of immunosuppressive drugs were studied in a rat model of heart transplantation. Results. Tolerance could be achieved if donor cell infusion was combined with delayed, but not immediate, low-dose cyclosporine treatment, and this was associated with activation and apoptosis of host lymphocytes. In contrast, combinations with an antibody against the interleukin 2 receptor led to long-term graft survival but severe chronic rejection, and combinations with high-dose cyclosporine or sirolimus led to acute rejection. Conclusions. Postoperative donor leukocyte infusion is a potential way for tolerance induction, but the type, dose, and timing of medication are highly critical for its efficacy.

Long-term allograft acceptance induced by single dose anti-leukocyte common antigen (RT7) antibody in the rat.

Background. In clinical organ transplantation monoclonal antibodies (mAb) to different surface molecules of immunocompetent cells become integral parts of the immunosuppressive therapy. In this study, a mAb against the rat leukocyte common antigen CD45 (RT7) was tested for its immunosuppressive potency after a single perioperative injection. Methods. Binding and depleting properties of the anti-RT7 mAb were investigated by flow cytometry. In the fully major histocompatibility complex–disparate heart and skin transplantation models (LEW [RT1l] → LEW.1W [RT1u]), a single dose of anti-RT7 mAb (10 mg/kg) was administered intravenously (day −1). To characterize the long-term acceptance of heart allografts second set skin transplantation (day 100), mixed lymphocyte reaction studies (day 100) and reverse transcriptase-polymerase chain reaction analysis for intragraft cytokine expression (day 200) were performed. Results. The anti-RT7 mAb bound to nearly all hematopoietic lineage cells, but particularly T and NK cells, and profoundly depleted these cells in circulation and lymphoid tissues. Anti-RT7 mAb-treated rats showed long-term acceptance of heart allografts (>200 days; n=12), whereas untreated recipients rejected allografts by day 8 (n=6). In contrast to hearts, primary skin allograft survival was only moderately prolonged. Animals with stable heart allograft acceptance showed normal in vitro lymphocyte proliferation responses to donor and third party antigen. These recipients also acutely rejected second set donor-strain skin grafts without inducing rejection of persisting heart allografts. Reverse transcriptase-polymerase chain reaction analysis of intragraft cytokines showed up-regulation of Fas-ligand and IL-4 mRNA in long-surviving heart allografts. Conclusions. The findings demonstrate that a single injection of an anti-RT7 mAb in the rat can induce stable long-term acceptance of heart allografts by transient but profound T-cell depletion. Local immunoregulatory mechanisms seem to play a role for maintenance of long-term graft acceptance.

Bone marrow aplasia induced by passenger leukocytes from heart allografts

OBJECTIVE Organ allografts contain passenger leukocytes that are transferred to the recipient with the transplantation, but their functional relevance to the recipients immune system is still controversial. MATERIALS AND METHODS To clarify the functional capacity of passenger leukocytes, we attempted to enhance their effect in rat heart allograft recipients by selective depletion of recipient leukocytes using a monoclonal antibody (mAb) against a recipient-specific allotype of CD45 (RT7(a)). RESULTS Although antibody treatment of the recipient alone led to profound lymphopenia and reversible myelosuppression, additional transplantation of an major histocompatibility complex-incompatible heart graft from an RT7(b) donor led to lethal aplastic anemia in the recipients. This lethal effect was completely abrogated by postoperative anti-CD3 treatment of the recipient and was partially abrogated or delayed by depletion of passenger leukocytes through additional anti-RT7(b) antibody treatment of the recipient or gamma-irradiation of the graft. CONCLUSIONS The results suggest a role for both donor and recipient-type T cells for the induction of aplastic anemia in this model. The study shows that, under defined conditions, allogeneic passenger leukocytes in a heart graft can have a profound effect on the recipients immune system and bone marrow.