Vascularized Organ Transplantation Research Articles

Engineering therapeutic T cells to suppress alloimmune responses using TCRs, CARs, or BARs.

Adoptive cell therapy with therapeutic T cells has become one of the most promising strategies to stimulate or suppress immune responses. Using virus-mediated genetic manipulation, the antigen specificity of T cells can now be precisely redirected. Tailored specificity has not only overcome technical limitations and safety concerns but also considerably broadened the spectrum of therapeutic applications. Different T cell-engineering strategies have now become available to suppress alloimmune responses. We first provide an overview of the allorecognition pathways and effector mechanisms that are responsible for alloimmune injuries in the setting of vascularized organ transplantation. We then discuss the potential to use different T cell-engineering approaches to suppress alloimmune responses. Specifically, expression of allospecific T cell receptors, single-chain chimeric antigen receptors, or antigen domains recognized by B cell receptors (B cell antibody receptors) in regulatory or cytotoxic T cells are considered. The ability of these strategies to control the direct or indirect pathways of allorecognition and the cellular or humoral alloimmune responses is discussed. An intimate understanding of the complex interplay that occurs between the engineered T cells and the alloimmune players is a necessary prerequisite for the design of safe and successful strategies for precise immunomodulation in transplantation.

Endothelial Cells in Antibody-Mediated Rejection of Kidney Transplantation: Pathogenesis Mechanisms and Therapeutic Implications.

Antibody-mediated rejection (AMR) has been identified as a main obstacle for stable immune tolerance and long survival of kidney allografts. In spite of new insights into the underlying mechanisms of AMR, accurate diagnosis and efficient treatment are still challenges in clinical practice. Endothelium is the first barrier between recipients' immune systems and grafts in vascularized organ transplants. Considering that endothelial cells express a number of antigens that can be attacked by various allo- and autoantibodies, endothelial cells act as main targets for the recipients' humoral immune responses. Importantly, emerging evidence has shown that endothelial cells in transplants could also initiate protective mechanisms in response to immune injuries. A better understanding of the role of endothelial cells during the pathogenesis of AMR might provide novel therapeutic targets. In the present review, we summarize the antigens expressed by endothelial cells and also discuss the activation and accommodation of endothelial cells as well as their clinical implications. Collectively, the progress discussed in this review indicates endothelial cells as promising targets to improve current diagnosis and therapeutic regimens for AMR.

The Influence of Donor and Recipient Gender Incompatibility on Corneal Transplant Rejection and Failure.

In vascularized organ transplants, gender mismatches have higher rates of immunological rejection. We investigated the influence of gender incompatibility, including H-Y incompatibility, on corneal transplant graft rejection and failure. Patients were included who had undergone a first corneal transplant for keratoconus (KC), Fuchs endothelial dystrophy (FED), pseudophakic bullous keratopathy (PBK), infection and other indications. A Cox regression model was fitted for each indication to determine factors affecting graft failure and rejection at 5 years. The impact of gender, including H-Y, matching was analyzed after accounting for other factors, including known risk factors. Of 18 171 patients, 4314 had undergone a transplant for FED, 4783 for KC, 3669 for PBK, 1903 for infection and 3502 for other disorders. H-Y mismatched (male [M]→female [F]) corneas were at greater risk of graft failure or rejection. For FED, F→F were 40% less likely to fail (p < 0.0001) and 30% less likely to reject (p = 0.01); M→M were 20% less likely to fail (p = 0.04) and 30% less likely to reject (p = 0.01). For KC, M→M matched corneas were 30% less likely to fail (p = 0.05) and 20% less likely to reject (p = 0.01) compared with H-Y mismatches. H-Y antigen mismatched (M→F) patients were at greater risk of rejection or graft failure.

The effect of pregnancy on humoral rejection in patients after vascularized organ transplantation

The aim of the study was to evaluate the effect of pregnancy on the production of donor- and nondonor-specific anti-human leukocyte antigen antibodies (anti-HLA Abs) in organ allograft recipients. The study group included four pregnant kidney (RT) and four liver (LT) transplant recipients. The genotype of HLA class I (A, B) and class II (DR) antigens was assessed. Anti-HLA antibodies class I and II were evaluated between 36 and 40 weeks’ gestation. Two different control groups consisted of the following: group I (n=8) with nonpregnant RT (n=6) and LT recipients (n=2), and group II with healthy pregnant women (n=10) with anti-HLA Abs detected between 38 and 41 weeks’ gestation. The HLA genotype was determined in fathers of the fetuses from the study group and group II controls. Half of group II controls had donor-specific anti-HLA (A, B, and/or DR) Abs, while nondonor-specific anti-HLA Abs were detected in all subjects from that group. Anti-HLA Abs were found in all group II controls. In the study group, anti-HLA Abs were found in only two LT recipients and one RT recipient, but they were not confirmed as donor-specific. Anti-HLA antibodies were not detected in the study group, whereas six out of ten group II controls had anti-HLA Abs against the HLA of the child's father. Pregnancy in vascularized organ recipients does not trigger the mechanism of humoral rejection involving anti-HLA class I and II antibodies with a potentially adverse impact on graft function.

Effector T cell migration to pancreatic islet allografts is dependent on donor antigen recognition and not Gai chemokine signaling. (TRAN1P.945)

Abstract Host effector T cell graft infiltration is a key step in the pathogenesis of both acute and chronic rejection. Recent data indicate effector T cell migration into primary vascularized organ transplants such as heart and kidney is driven by donor antigen recognition, not via Gai-coupled chemokine receptor signaling. Whether it is true for neovascularized pancreatic islet allografts is unknown. We co-transferred pertussis toxin (Ptx)-treated and untreated TCR-tg (OT-1) effector T cells into B6 recipients 7-10 days after transplanting B6-Act-OVA or B6 islet grafts under the kidney capsule. Ptx irreversibly blocks Gai-dependent signaling. Grafts were imaged 20-24 hours after T cell transfer by two-photon intravital microscopy. We observed B6-Act-OVA grafts were heavily infiltrated with OT-1 effector T cells irrespective of pre-treatment with Ptx or not (453.7+/-170.0 vs. 632.8+/-221.1 per imaging volume, n = 10, p = 0.0522 ), while B6 grafts, which lack the cognate antigen recognized by OT-1 T cells, were free of infiltrate (&amp;lt;5 cells, n = 7). These results indicate that, similar to heart and kidney transplants, islet allograft infiltration with host effector T cells is driven by donor antigen, not by Gai signaling. The paradigm of antigen-driven effector T cell migration is therefore generalizable to both primary vascularized grafts, in which donor antigens are abundantly expressed on the endothelium, and neovascularized cellular grafts whereby the endothelium is of host origin.

Patient care after heart and kidney transplantation – case report

A patient after vascularized organ transplantation requires especially attentive medical care because of the risk of rejection of the transplant, the necessity for monitoring immunosuppressive therapy, and increased susceptibility to infection during this treatment. Due to the progress in transplantation medicine the number of patients after organ transplantation (heart, kidneys, liver, lungs) increases every year, and becomes a great challenge for medical specialists, and increasingly more often for specialists in family medicine. It is important that physicians should be adequately prepared to cover this group of patients with care.

Origin and Function of Host-Derived Dendritic Cells in Vascularized Organ Transplants.

Donor DC exit organ grafts after transplantation and are replaced by host DC. The origin and function of these tissue-resident, host DC however is unknown. Using flow cytometry and 2-photon intravital microscopy, we observed rapid replacement of donor DC by host DC in both syngeneic and allogeneic heart and kidney grafts within 3 days after transplantation, and demonstrated that the vast majority of host DC were monocyte-derived (Mono-DC). By day 7, allografts harbored >5-fold more DC than syn grafts and >45-fold more than native tissue. Enhanced DC accumulation in allografts was inhibited by treating recipients with cyclosporine A, indicating a crucial role for the host alloimmune response in DC influx. DC accumulation was only partially dependent on fractalkine receptor (CX3CR1) expression on host monocytes. Host DC that infiltrated grafts localized to perivascular areas, extended dendrites into the vessel lumens, mediated the transmigration of cognate effector T cells (Teff) into the graft, and continued to make stable contacts with the Teff after they had transmigrated. Depletion of host Mono-DC by administering DT to CD11b-DTR mice significantly reduced T cell infiltration and IFN-γproduction in the graft. Hence, host Mono-DC amass rapidly in transplanted organs, replacing donor DC, and accelerate rejection by enhancing Teff migration, retention, and activation. Our results provide novel insights into the pathogenesis and treatment of allograft rejection.

Origin and function of host-derived dendritic cells in vascularized organ transplants (TRAN3P.889)

Abstract Donor DC exit organ grafts after transplantation and are replaced by host DC. The origin and function of these tissue-resident, host DC however is unknown. Using flow cytometry and 2-photon intravital microscopy, we observed rapid replacement of donor DC by host DC in both syngeneic and allogeneic heart and kidney grafts within 3 days after transplantation, and demonstrated that the vast majority of host DC were monocyte-derived (Mono-DC). By day 7, allografts harbored &amp;gt;5-fold more DC than syn grafts and &amp;gt;45-fold more than native tissue. Enhanced DC accumulation in allografts was inhibited by treating recipients with cyclosporine A, indicating a crucial role for the host alloimmune response in DC influx. DC accumulation was only partially dependent on fractalkine receptor (CX3CR1) expression on host monocytes. Host DC that infiltrated grafts localized to perivascular areas, extended dendrites into the vessel lumens, mediated the transmigration of cognate effector T cells (Teff) into the graft, and continued to make stable contacts with the Teff after they had transmigrated. Depletion of host Mono-DC by administering DT to CD11b-DTR mice significantly reduced T cell infiltration and IFN-γ production in the graft. Hence, host Mono-DC amass rapidly in transplanted organs, replacing donor DC, and accelerate rejection by enhancing Teff migration, retention, and activation. Our results provide novel insights into the pathogenesis and treatment of allograft rejection.

Primary Vascularization of Allografts Governs Their Immunogenicity and Susceptibility to Tolerogenesis

We investigated the influence of allograft primary vascularization on alloimmunity, rejection, and tolerance in mice. First, we showed that fully allogeneic primarily vascularized and conventional skin transplants were rejected at the same pace. Remarkably, however, short-term treatment of mice with anti-CD40L Abs achieved long-term survival of vascularized skin and cardiac transplants but not conventional skin grafts. Nonvascularized skin transplants triggered vigorous direct and indirect proinflammatory type 1 T cell responses (IL-2 and IFN-γ), whereas primarily vascularized skin allografts failed to trigger a significant indirect alloresponse. A similar lack of indirect alloreactivity was also observed after placement of different vascularized organ transplants, including hearts and kidneys, whereas hearts placed under the skin (nonvascularized) triggered potent indirect alloresponses. Altogether, these results suggest that primary vascularization of allografts is associated with a lack of indirect T cell alloreactivity. Finally, we show that long-term survival of vascularized skin allografts induced by anti-CD40L Abs was associated with a combined lack of indirect alloresponse and a shift of the direct alloresponse toward a type 2 cytokine (IL-4, IL-10)-secretion pattern but no activation/expansion of Foxp3(+) regulatory T cells. Therefore, primary vascularization of allografts governs their immunogenicity and tolerogenicity.

SPECT/CT Lymphoscintigraphy of Heterotopic Cardiac Grafts Reveals Novel Sites of Lymphatic Drainage and T Cell Priming

The normal function of lymphatic vessels is to facilitate the trafficking of antigen presenting cells to draining lymph nodes where they evoke an immune response. Donor lymphatic vessels are not connected to that of recipients' during organ transplantation. The pathophysiology of this disruption has received little attention. Murine heterotopic cardiac transplantation has been used extensively in transplantation research. Following vascularized organ transplantation, the main site of allosensitization is thought to be in the spleen of the recipient as a result of migration of donor passenger leukocytes via blood. Here, using Single Photon Emission Computed Tomography/Computerized Tomography (SPECT/CT) lymphoscintigraphy, we studied the pattern of lymphatic flow from mouse heterotopic abdominal cardiac grafts and identified mediastinal lymph nodes as the draining nodes for the donor graft. Staining with HY tetramer after transplantation of HY mismatched heart grafts and ELISPOT following allogeneic grafts to detect donor specific T cells revealed them as important sites for allosensitization. Our data indicates that mediastinal lymph nodes play a crucial role in the alloimmune response in this model, and should be used for ex vivo and adoptive transfer studies after transplantation in addition to the spleen.

Progress in abdominal organ transplantation

SummaryThe excellent results of vascularized organ transplantation have resulted in an increasing number of end-stage organ failure patients seeking such treatment. The results of organ transplantation depend on a number of factors – the quality of the donor (and an organ), living vs. deceased donation, magnitude of ischemic injury (and its prevention), and recipient-dependent factors. Ischemia/reperfusion injury in organ transplantation is a multifactorial process, which may lead to delayed graft function. In addition, surgical and preservation techniques, type of immunosuppressive regimens, complications after transplantation and post-transplant management may also have a significant impact on short- and long-term results of transplantation. In this paper we describe advances in transplantation in recent years, with particular emphasis on kidney, liver, intestines, whole pancreas and pancreatic islets.

Extracellular purines serve as an innate trigger to allograft rejection (145.39)

Abstract Innate immune triggers are well studied in infectious models, yet they remain unclear in solid organ transplantation. Extracellular purines (e.g. ATP, adenosine), which signal through receptors on immune cells, modulate the extent of ischemia reperfusion injury. To test the effect of purinergic signaling on graft rejection, we used a cardiac transplant model in outbred mice in which accelerated graft rejection occurs within 4 days post-transplantation and is highly dependent on innate mediators. Treatment with NECA, an adenosine receptor agonist, 2 hours prior to heterotopic heart transplantation (n=8) completely abrogated accelerated rejection compared to untreated outbred mice (incidence = 28%, n=35). CD39 and CD73 are ectoenzymes responsible for metabolism of extracellular ATP to adenosine. To determine the effect of increased extracellular ATP in the local graft environment, hearts from CD73-deficient B6 mice were transplanted to BALB/c recipients. Accelerated rejection occurred in 24% of grafts from CD73-/- mice (n=17) compared to 2% of grafts from wt inbred controls (n=42). Skin transplants from CD39-/- and CD73-/- mice to BALB/c mice (n=8/group) rejected at the same rate as wt control B6 to BALB/c skin transplants (n=8), suggesting the effect was restricted to vascularized transplants. Our data suggests that modulation of purine metabolism serves as an early trigger of the innate immune response in vascularized organ transplantation.

An optimization of protocol for mixed chimerism induction in mice model.

Studies on mixed chimerism are currently focused primarily on obtaining less toxic conditioning protocols. With these issues in mind, we have undertaken the attempt to optimize the procedure of mixed chimerism induction in mice. In order to reduce toxicity, we used decreasing doses of total body irradiation (TBI) together with combination of blocking antibodies. We also tried to eliminate immunosuppression (cyclophosphamide - CP) treatment after bone marrow transplantation. B6.SJL-PtprcaPep3b mice were injected with 20-30 x 106 bone marrow cells from Balb C mice. Mice were treated with TBI (3 - 1.5 - 0 Gy) on "-1" day of the experiment and blocking antibodies against CD40L ("0", and "4" days) and additionally anti-CD8 ("-2" day) and/or anti-NK1.1 ("-3" day). Mice in certain groups also received CP (175 mg/kg) on "2" day. Presence of mixed chimerism was assessed in peripheral blood cells by flow cytometry on the 1st, 2nd, 3rd, 4th, 6th and 8th weeks of the experiment by detecting of CD45.1 (characteristic for B6.SJL-PtprcaPep3b strain) and CD45.2 (characteristic for Balb C strain) antigens expression. We also analyzed the percentage of peripheral blood CD8 T-cells (CD3e/CD8a) and NK cells (Ly-49D/NK1.1). We found that reduction of TBI dose and elimination of CP decrease the rate of mixed chimerism formation. The highest percentage of donor cells was obtained in the group of animals treated with 3 Gy of TBI, CP and combination of anti-CD40L, anti-CD8, and anti-NK1.1 antibodies. The 3 Gy TBI was necessary to induce stable mixed chimerism, but it could be obtained without the CP use. The percentage of CD3e/CD8a and Ly-49D/NK1.1 cells was significantly lower in the groups of mice treated by corresponding antibodies. Moreover, we observed the lowest number of peripheral blood Ly-49D/NK1.1 cells in the group of animals with highest mixed chimerism. Our experiments in mice model can help in better understanding of mixed chimerism phenomenon and in selecting the method of mixed chimerism induction with lowest possible toxicity. This also might improve the protocols of stable mixed chimerism induction in humans, and in the future, the effectiveness of vascularized organ transplantation.

Mechanisms involved in antibody- and complement-mediated allograft rejection

Antibody-mediated rejection has become critical clinically because this form of rejection is usually unresponsive to conventional anti-rejection therapy, and therefore, it has been recognized as a major cause of allograft loss. Our group developed experimental animal models of vascularized organ transplantation to study pathogenesis of antibody- and complement-mediated endothelial cell injury leading to graft rejection. In this review, we discuss mechanisms of antibody-mediated graft rejection resulting from activation of complement by C1q- and MBL (mannose-binding lectin)-dependent pathways and interactions with a variety of effector cells, including macrophages and monocytes through Fcgamma receptors and complement receptors.

Transplantation immunobiology: two important themes

Organ transplantation has become the method of choice for the treatment of many diseases because of advances in both surgical and immunosuppressive techniques. These techniques have now progressed to the stage where satisfactory graft acceptance and survival can be achieved by the judicious choice of donor and recipient, and by the appropriate patient management. As a result of the improved quality of life of transplant recipients, an ever-increasing number of potential recipients are being added to the waiting lists. Despite the success of clinical transplantation, two key challenges for the field are (1) the dwindling number of suitable donors and (2)the adverse effects and the risks of long-term administration of immunosuppressive drugs. It is a well-established fact that the supply of organs for transplantation is not only inadequate but also worsening globally. Although education programs and various legal changes may increase the availability of donors, it is unlikely to solve the problem completely. The potential solutions being examined include the use of artificial organs, stem cells to engineer organs and tissues and xenotransplantation. Even though the first two options are appealing and shows potential, xenotransplantation is closest to clinical applications. The proposed use of pig organs for xenotransplantation has reintroduced the problem of hyperacute rejection, which results in the antibody-mediated destruction of a vascularized pig organ within minutes because of natural preformed antibodies. The expression of human complement regulators, either alone, or in combination, was initially used in an attempt to protect the xenograft. Several human complement regulators extended the survival of pig organs in primate transplant models from minutes or hours to days in unmodified recipients, and was further increased to 5 months with intense immunosuppression. A significant step for the field was not only the identification of the carbohydrate antigen recognized by naturally occurring human antibodies but, more importantly, the generation of homozygous pigs with a disruption in the gene that encodes the enzyme, α1,3galactosyltransferase. Using these Gal knockouts for pig-to-primate vascularized organ transplants, again using clinically inappropriate immunosuppression, showed survival times similar to those observed in the pigs expressing human complement regulators. Therefore, although inactivation of the α1,3galactosyltransferase gene prevents hyperacute rejection of vascularized organs, they still succumb to acute vascular rejection as a consequence of antibodies (natural or elicited) binding to and activating endothelial cells, or unidentified cross-species molecular incompatibilities. For transplantation, the major side effects of long-term immunosuppression are infection and malignancy. A major goal in transplantation for many years has been the induction of ‘transplant tolerance’—the long-term acceptance of the transplant with the maintenance of responses to nondonor antigens (such as viruses) without the need for chronic immunosuppression. This has involved immunological manipulations to re-educate the immune system to establish transplant tolerance and allow permanent survival of the graft. Despite continuing research in this area and success in small animal models, clinical tolerance remains elusive. Recently, interest has been renewed in the subsets of T cells, notably regulatory T cells (Tregs) and their potential role in tolerance induction. These Tregs can be identified by their expression of CD4, CD25 and the transcription factor, Foxp3. Foxp3 is currently the single best marker available for identifying Tregs, with stable expression of Foxp3, and is regarded as the ‘master switch’ for Treg function. For clinical transplantation, the ability to harness the potential of allograft-specific Treg cells would reduce the need for nonspecific immunosuppressive therapy currently required to prevent graft rejection, and may be a critical step toward inducing donor-specific tolerance and permanent graft survival. The following two Special Feature Reviews, which are contributions from the State of the Art Symposia at the recent Sydney Congress of the Transplantation Society, address progress made in these two areas: a review by Cowan and d’Apice on the current state of the field of xenotransplantation focusing on the complement and coagulation systems and their roles in vascular injury; and a review by Wang et al. on the role of histone deacetylases and their inhibitors on the generation of Foxp3+ Tregs with particular emphasis on transplantation tolerance induction. By a combination of both these approaches in the future, it is feasible that the dual problems of organ availability and long-term side effects of immunosuppression may be solved.

Comparison of different procurement regimen to reduce the ischemia reperfusion injury of murine vascular allografts

Aims: Ischemia reperfusion injury is an important non-immunological factor contributing to the development of chronic rejection after vascularized organ transplantation. The aim of this study was to compare different cell culture media in terms of vascular lesion formation as a consequence of ischemia reperfusion injury after implantation into mice. Methods: BALB/c (H2d) aortic grafts were incubated in three different cell media (ECG medium, RPMI 1640, Waymouth/Ham's F12) for various time spans (T1=5.5h, T2=6.5h, T3=8.5h) at 37°C and transplanted into syngeneic recipients. Histology and morphometric measurements were performed on day 30 after transplantation. Results: A total of 36 transplants were performed for this study with an overall survival rate of 72.0%, the most frequent complication being thrombosis of the aortic graft (n=9) and one late death of the recipient due to other courses. All recipients that had received vascular grafts incubated in ECG medium survived and showed no signs of intimal proliferation. Aortic grafts incubated in RPMI medium resulted in a reduced survival rate (67.0%) for the recipients. Vascular grafts incubated in Waymouth medium resulted in only 50% survival of the transplant recipients and analysis of the vascular morphology revealed moderate amounts of intimal proliferation in 2 grafts. Conclusions: Data from the current study suggest that vascular grafts can be safely kept in tissue culture up to 8.5 hours without the risk of significant ischemia damage. In addition we could demonstrate that there are marked differences in vascular integrity and animal survival depending on the used medium.

Platelets Influence Vascularized Organ Transplants from Start to Finish

Platelets Influence Vascularized Organ Transplants from Start to Finish

“Alternatively Activated” Dendritic Cells Preferentially Secrete IL-10, Expand Foxp3+CD4+ T Cells, and Induce Long-Term Organ Allograft Survival in Combination with CTLA4-Ig

In this study, we propagated myeloid dendritic cells (DC) from BALB/c (H2(d)) mouse bone marrow progenitors in IL-10 and TGF-beta, then stimulated the cells with LPS. These "alternatively activated" (AA) DC expressed lower TLR4 transcripts than LPS-stimulated control DC and were resistant to maturation. They expressed comparatively low levels of surface MHC class II, CD40, CD80, CD86, and programmed death-ligand 2 (B7-DC; CD273), whereas programmed death-ligand 1 (B7-H1; CD274) and inducible costimulatory ligand expression were unaffected. AADC secreted much higher levels of IL-10, but lower levels of IL-12p70 compared with activated control DC. Their poor allogeneic (C57BL/10; B10) T cell stimulatory activity and ability to induce alloantigen-specific, hyporesponsive T cell proliferation was not associated with enhanced T cell apoptosis. Increased IL-10 production was induced in the alloreactive T cell population, wherein CD4+Foxp3+ cells were expanded. The AADC-expanded allogeneic CD4+CD25+ T cells showed enhanced suppressive activity for T cell proliferative responses compared with freshly isolated T regulatory cells. In vivo migration of AADC to secondary lymphoid tissue was not impaired. A single infusion of BALB/c AADC to quiescent B10 recipients induced alloantigen-specific hyporesponsive T cell proliferation and prolonged subsequent heart graft survival. This effect was potentiated markedly by CTLA4-Ig, administered 1 day after the AADC. Transfer of CD4+ T cells from recipients of long-surviving grafts (>100 days) that were infiltrated with CD4+Foxp3+ cells, prolonged the survival of donor-strain hearts in naive recipients. These data enhance insight into the regulatory properties of AADC and demonstrate their therapeutic potential in vascularized organ transplantation.

The Impact of Corneal Allograft Rejection on the Long-Term Outcome of Corneal Transplantation

To examine the influence of corneal allograft rejection on the survival of penetrating corneal transplantation, to review the status of conventional therapies to improve graft survival, and to consider prospects for alternative approaches to reduce the impact of rejection. Perspective, including prospective, observational cohort study. An examination of the literature on human corneal graft rejection and data from the Australian Corneal Graft Registry, reviewed in the context of clinical experience. Corneal graft outcome is not improving with era. The sequelae of inflammation, whether occurring before corneal transplantation or subsequently, exert a profound influence by predisposing the graft to rejection. Of the developments that have been instrumental in reducing rejection in vascularized organ transplantation, living-related donation is not an option for corneal transplantation. However, HLA matching may be beneficial and requires reassessment. The evidence base to support the use of systemic immunosuppressive agents in corneal transplantation is thin, and topical glucocorticosteroids remain the drugs of choice to prevent or reverse rejection episodes. Experimental approaches to local allospecific immunosuppression, including the use of antibody-based reagents and gene therapy, are being developed but may be difficult to translate from the laboratory bench to the clinic. Corneal allograft rejection remains a major cause of graft failure. High-level evidence to vindicate the use of a particular approach or treatment to prevent or treat corneal graft rejection is lacking. In the absence of extensive data from randomized, controlled clinical trials, corneal graft registers and extrapolation from experimental models provide some clinically useful information.

Perforin Mediates Endothelial Cell Death and Resultant Transplant Vascular Disease in Cardiac Allografts

T cell-induced endothelial injury is an important event in the development of transplant vascular disease (TVD), the leading expression of chronic rejection of vascularized organ transplants. However, the precise contribution of perforin to vascular damage in allografts and resultant TVD has not been addressed in vivo. Minor histocompatability antigen mismatched mouse heterotopic cardiac transplants were performed from 129J donors into C57Bl/6 (wild-type (WT)) or perforin knockout (PKO) recipients. Perforin was abundant in immune infiltrates in the myocardium and vasculature of transplanted hearts in WT mice. Allograft coronary arteries in both WT and PKO mice had considerable vasculitis. There was also marked endothelial disruption, as well as TUNEL-positivity in the endothelial region, in coronary arteries of hearts transplanted into WT mice that was not evident in PKO recipients (P = 0.05). At 30 days post-transplantation, intimal thickening was assessed on elastic Van Gieson-stained ventricular sections. There was an average of 54.2 +/- 6.7% luminal narrowing of coronary arteries in allografts from WT mice as compared to 13.4 +/- 5.1% luminal narrowing in PKO counterparts (P < 0.00002). In summary, perforin plays a primary role in endothelial damage and the resultant onset and progression of TVD.