Wilson Wong

Transplant renal artery stenosis in 77 patients--does it have an immunological cause?

Transplant renal artery stenosis (TRAS) is a common complication after transplantation and is an important cause of graft dysfunction. Damage from graft rejection, trauma, and atherosclerosis have been implicated as possible causes. We reviewed all 917 patients transplanted in our unit since 1978 to study the prevalence, clinical features, and possible causes of TRAS. Seventy-seven patients with TRAS were identified. The detected incidence was 2.4% before the introduction of color doppler ultrasonography (CDU) and rose to 12.4% after CDU was introduced in 1985, giving an overall incidence of 8.4% during a mean follow-up period of 6.9 years. The TRAS group was compared with a control group of 77 transplanted patients matched for age, year of transplant, sex, and number of previous grafts. Mean ages for the study and control groups were 43.6 +/- 15 and 44.8 +/- 13.7 yr. A total of 25% of cases of TRAS were diagnosed within the first 8 wk of transplantation and in 60% within the first 30 wk (median = 23 wk). All patients were treated with angioplasty, 28 patients had recurrence of TRAS requiring multiple angioplasties (maximum 5) and 1 went on to have surgery. Angioplasty resulted in a significant fall in plasma creatinine. Patient and graft survival were significantly worse in the TRAS group: 69% vs. 83% (P < 0.05) and 56% vs. 74% (P < 0.05) (TRAS vs. Control), respectively. There was a significantly higher incidence of rejection, especially cellular rejection in the TRAS group, 0.67 vs. 0.35 episodes per patient (P < 0.01) (TRAS vs. Control). Recurrence but not occurrence of TRAS was associated with the use of cyclosporine.

Pivotal role of CD4+ T cells in renal fibrosis following ureteric obstruction

Tubulointerstitial fibrosis is a common consequence of a diverse range of kidney diseases that lead to end-stage renal failure. The degree of fibrosis is related to leukocyte infiltration. Here, we determined the role of different T cell populations on renal fibrosis in the well-characterized mouse model of unilateral ureteric obstruction. Depletion of CD4(+) T cells in wild-type mice with a monoclonal antibody significantly reduced the amount of interstitial expansion and collagen deposition after 2 weeks of obstruction. Reconstitution of lymphopenic RAG knockout mice with purified CD4(+) but not CD8(+) T cells, prior to ureteric obstruction, resulted in a significant increase in interstitial expansion and collagen deposition. Wild-type mice had significantly greater interstitial expansion and collagen deposition compared with lymphopenic RAG(-/-) mice, following ureteric obstruction; however, macrophage infiltration was equivalent in all groups. Thus, our results suggest that renal injury with subsequent fibrosis is likely to be a multifactorial process, with different arms of the immune system involved at different stages. In this ureteric obstruction model, we found a critical role for CD4(+) T cells in kidney fibrosis. These cells could be a potential target of therapeutic intervention to prevent excessive fibrosis and loss of function due to renal injury.

Pretransplant administration of a single donor class I major histocompatibility complex molecule is sufficient for the indefinite survival of fully allogeneic cardiac allografts: evidence for linked epitope suppression.

BACKGROUND Allograft survival can be prolonged by the administration of alloantigen(s) before transplantation. Blood transfusion is the commonest form of alloantigen pretreatment currently used in clinical practice. However, for recipients of organs from cadaver donors, it is not possible to predict the identity of the organ donor in advance. Therefore, it is highly unlikely that all the alloantigens expressed by a cadaver organ donor will be represented in the alloantigen pretreatment inoculum. We have previously shown that it is not necessary to expose the recipient to the full complement of donor alloantigens to induce long-term survival of a subsequent cardiac allograft. Here, we investigated the in vivo mechanism responsible for this phenomena. METHODS Unresponsiveness to the mouse MHC class I molecule Kb was induced in CBA.Ca (H2k) recipients by administration of bone marrow cells from transgenic CBA mice, CBK (H2k + Kb) before transplantation of fully allogeneic and F1 vascularized cardiac allografts. RESULTS Pretreatment with CBK bone marrow cells resulted in the long-term survival of all cardiac allografts expressing H2-Kb. For example, C57BL/10 (H2b) and (CBKxBALB/c) F1 (H2k,d + Kb) cardiac allografts were accepted by recipients treated with CBK bone marrow cells before transplantation. In contrast, allografts that did not express Kb, such as BALB/c (H2d) or (CBAxBALB/c) F1 (H2k,d), were rejected acutely, with a median survival time (MST) of 7 and 6 days, respectively, in recipients treated with CBK bone marrow cells. Furthermore, when recipients pretreated with CBK bone marrow cells were grafted with a BALB/c heart and a CBK heart simultaneously, the BALB/c hearts were rejected (MST=10 days), whereas the CBK hearts were accepted. By contrast, in the maintenance phase (i.e., after transplantation), recipients with long-term surviving (CBKxBALB/c) F1 hearts (> 100 days) were found to accept BALB/c hearts indefinitely, whereas fourth-party B10.S (H2s) grafts were rejected (MST=7.5 days). This indicated that the allografts bearing Kb could tolerize recipients to other alloantigens expressed by the transplanted heart. CONCLUSIONS These data provide clear evidence for linked epitope suppression in the induction of operational tolerance in vivo.

Homeostatic Proliferation of Lymphocytes Results in Augmented Memory-Like Function and Accelerated Allograft Rejection

Homeostatic proliferation is a normal physiological process triggered by lymphopenia to maintain a constant level of T cells. It becomes the predominant source of new T cells in adulthood after thymus regression. T cells that have undergone homeostatic proliferation acquire the memory phenotype, cause autoimmune disease, and are resistant to tolerance induction protocols. Transplantation is a rare example in which lymphopenia is deliberately induced for its immunosuppressive effect. However, it is not known whether the homeostatic proliferation that follows will have the opposite effect and accelerate rejection. We show that T cells that have undergone homeostatic proliferation acquire a memory phenotype, spontaneously skews toward the Th1 phenotype, even in the absence of antigenic stimulus. Interestingly, in contrast, the percentage of Foxp3+ regulatory T cells increased by 28-fold following homeostatic proliferation. Using a mouse life-sustaining kidney transplant model, we showed that T cells that have gone through homeostatic proliferation in lymphopenic hosts transformed chronic rejection to acute rejection of a single MHC class II-mismatched kidney allograft. T cells that have undergone homeostatic proliferation consistently cause reliable rejection even when bona fide memory T cells cannot. These functional changes are long-lasting and not restricted to the acute phase of homeostatic proliferation. Our findings have important implications for tolerance induction or graft-prolonging protocols involving leukocyte depletion such as irradiation bone marrow chimera, T cell-depleting Abs, and lymphopenia induced by infections such as CMV and HIV.

Synthesis of complement protein C3 in the kidney is an important mediator of local tissue injury

Increased exposure of the tubular epithelium to filtered protein is a proposed mechanism of progressive renal failure associated with glomerular disease, but how this protein overload translates into tubular damage remains unclear. We have examined a model of adriamycin‐induced proteinuria to determine the effect of locally synthesized C3, the central proinflammatory protein of the complement cascade. C3—/— kidney isografts placed in wild‐type C3+/+ mice were protected from proteinuria‐associated complement activation, tubular damage, and progressive renal failure despite the presence of abundant circulating C3. The quantity of urinary protein was unaffected by the absence of C3, and thus the influence of C3 was not explained by alteration in the filtered protein load. These results suggest that local synthesis of complement from renal epithelial cells is a critical mediator of tubular damage in proteinuria‐associated renal disease. Our results concur with previous findings of increased synthesis of C3 in human tubular epithelium exposed to high concentrations of protein in vitro. Because progressive renal damage in humans associates with proteinuria regardless of cause, our findings have implications for the pathogenesis and treatment of renal failure from many common causes, immunological and nonimmunological. Sheerin, N. S., Risley, P., Abe, K., Tang, Z., Wong, W., Lin, T., Sacks, S. H. Synthesis of complement protein C3 in the kidney is an important mediator of local tissue injury. FASEB J. 22, 1065–1072 (2008)

Natural killer-cell activity after human renal transplantation in relation to killer immunoglobulin-like receptors and human leukocyte antigen mismatch

Background. Natural killer (NK) cells use killer immunoglobulin-like receptors (KIR) that bind to self-class I major histocompatibility complex (MHC) molecules to prevent killing of autologous cells. Mismatched allografts, which do not express recipient MHC class I molecules, can therefore be potential targets for NK-cell killing. In our living related-unrelated renal transplantation program, donor-recipient pairs vary in the amount of both HLA and KIR genes they share. This provides us with a unique opportunity to dissect the influence of KIR on NK-cell function after transplantation. Methods. Recipient NK cells were used in a cytotoxicity assay against donor peripheral blood mononuclear cells 2 days before, on the day of, and 3 days after transplantation. Results were correlated to HLA-KIR compatibility between donor and recipient. Results. NK killing, in a direct ex vivo setting, was demonstrated to be HLA mismatch dependent. Recipient NK antidonor cytotoxicity was unaltered despite having received 2 days’ treatment with cyclosporine A before transplantation. However, cytotoxicity increased 3 days after transplantation in 71% of recipients. Recipients exhibiting increased NK cytotoxicity against their donors after transplantation were found to possess more activating KIR genes specific for donor class I MHC molecules than those in whom killing activity did not increase (P <0.04). Conclusions. NK cells are activated after transplantation despite quadruple immunosuppression, suggesting that recipient NK-cell cytotoxicity against the donor may be a previously unrecognized area of the rejection process, especially in poorly matched donor-recipient pairs where the recipient may not express the correct repertoire of inhibitory receptors to prevent killing of donor cells.

Syngeneic bone marrow expressing a single donor class I MHC molecule permits acceptance of a fully allogeneic cardiac allograft

Bone marrow cells may be a useful vehicle for pretransplant delivery of alloantigen to induce tolerance in vivo. However, infusion of fully allogeneic bone marrow cells carries the risk of graft-versus-host disease. In order to reduce this risk while retaining the tolerogenic potential of the bone marrow infusion, we have investigated the ability of recipient bone marrow cells expressing a single donor MHC class I antigen to induce specific unresponsiveness in vivo. We show that 5x10(7) and 5x10(6) bone marrow cells from a transgenic strain of CBA mice, CBK, that express a single donor class I MHC gene, H2Kb (H2k + H2Kb), were able to induce long term survival of a fully allogeneic C57BL/10 (H2b) cardiac allograft in 80% and 20% of unmanipulated CBA (H2k) recipients, respectively, when administered intravenousely on the day of transplantation. In contrast, the same doses of fully allogeneic C57BL/10 donor bone marrow were completely ineffective at prolonging graft survival. When the interval between bone marrow infusion and transplantation was increased to 14 days, CBK bone marrow at either dose (5x10(6) and 5x10(7)) induced long term survival of C57BL/10 cardiac allografts in all recipients (MST>100 days) while fully allogeneic donor bone marrow was ineffective (MST=7, 5x10(6) cells; MST=6, 5x10(7)). Only when 27 or 42 days had elapsed between bone marrow infusion and transplantation did fully allogeneic bone marrow exert a beneficial effect on graft survival. Administration of 5x10(6) C57BL/10 bone marrow cells 27 and 42 days before transplantation resulted in long term survival of C57BL/10 hearts in 67% and 75% of CBA recipients. Next, we investigated whether manipulating the periphery of the recipient with a depleting anti-Cd4(4) monoclonal antibody before bone marrow infusion would facilitate the induction of unresponsiveness. When pretreatment with bone marrow cells was combined with anti-Cd4 monoclonal antibody 28 days before transplantation, a 10-fold reduction in the number of either C57BL/10 or CBK bone marrow cells required to induce tolerance was observed. These data confirm that bone marrow is a suitable vehicle for alloantigen delivery at the time of, or before, transplantation, on its own or in combination with anti-Cd4. The use of recipient type bone marrow cells expressing one or more donor MHC genes may be more effective than fully allogeneic, donor bone marrow cells in inducing tolerance in vivo. This difference may have important clinical implications for the current trials of donor bone marrow given at the time of transplantation in order to augment chimerism and to prolong graft survival.

Tertiary lymphoid organs in renal allografts can be associated with donor-specific tolerance rather than rejection

Tertiary lymphoid organs can form at sites of chronic inflammation. Their presence has been mainly associated with tissue destruction. In transplantation, there is a dynamic immune response as in chronic inflammation. Indeed, the presence of tertiary lymphoid organs has been associated with chronic rejection. In addition to a destructive alloimmune response, secondary lymphoid organs are also important in transplant tolerance. We hypothesised that tertiary lymphoid organs may also form during transplantation tolerance as this process also requires an active local immune response. If so, their presence may enhance tolerance, resulting in better graft function rather than chronic rejection. Using a mouse kidney allograft model of tolerance, we demonstrate the formation of tertiary lymphoid organs within tolerated allografts. Tertiary lymphoid organs are supplied by high endothelial venules, and contain T and B cells, macrophages, DC, Foxp3+ T cells, donor MHC class II+ cells and recipient cells presenting donor‐derived allopeptides. Formation of tertiary lymphoid organs and the presence of immune cells within them are associated with superior graft function, suggesting that tertiary lymphoid organs act to amplify the prevailing immune response, be it a tolerant and beneficial immune response or the previously described destructive alloimmunity.

Extensive and bidirectional transfer of major histocompatibility complex class II molecules between donor and recipient cells in vivo following solid organ transplantation

Intercellular transfer of surface molecules has been demonstrated in vitro, or in vivo under artificial situations. Transplantation is a unique clinical situation in which foreign major histocompatibility complex (MHC) molecules are deliberately introduced. This provides a model to study intercellular MHC transfer because donor MHC molecules can easily be tracked. Here we describe the bidirectional transfer of MHC class II molecules between donor and recipient cells after transplantation of vascularized kidney and cardiac allografts in mice. Cells that are positive for both donor and recipient MHC class II accounted for up to 30% of the donor MHC class II+ population, suggesting that they play a significant role in the antigen presentation process. The majority of these cells were dendritic cells, but macrophages and B cells were also able to acquire foreign MHC molecules. Most doublepositive cells were also positive for costimulatory molecules, indicating a capability to elicit a T‐cell response. This transfer of MHC molecules between donor and recipient cells provides a link between the direct and indirect pathways of alloantigen presentation and suggests that MHC transfer is also likely to occur under normal physiological conditions, which has implications in the fields of infection, vaccination, and tumor immunology.— Brown, K., Sacks, S. H., Wong, W. Extensive and bidirectional transfer of major histocompatibility complex class II molecules between donor and recipient cells in vivo following solid organ transplantation. FASEB J. 22, 3776–3784 (2008)

The role of complement in regulating the alloresponse

Purpose of reviewThis review emphasizes new information concerning the role of anaphylatoxins in the regulation of the immune response to allografts. Its timeliness relates the growing concept of the innate immune response as a regulator of the adaptive immune system and to how this concept lends itself to therapeutic advance. Recent findingsRecent work has extended our understanding of the role of local complement synthesis and how this facilitates the interaction between antigen-presenting cells and alloreactive T cells, resulting in a potent effector response. In particular, this work has identified new roles for anaphylatoxins as regulators of antigen presentation, T-cell proliferation and T-cell longevity. SummaryStrategies for comprehensive blockade of complement at the site of action, or for more selective blockade of specific complement components, are not only possible but merit further exploration based on these results.