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The development of pigs transgenic for human regulators of complement activation resulted in the nearly total elimination of episodes of hyperacute rejection following discordant solid organ xenotransplantation. Following discordant heart or kidney transplantation, in subhuman primates, graft survival rates of up to several months can be observed. In contrast to these organs, the xenotransplantation of the liver is associated with the inherent problem of the immunological and metabolic compatibility of the large variety of xenoproteins generated. Based on a review of data mainly derived from experimental ex-vivo xenoliver perfusions in patients with hepatic coma, whole organ orthotopic or heterotopic liver xenotransplantation currently is not likely to become a relevant option for the treatment of patients with endstage liver failure. In contrast, clinical studies utilizing different forms of bioartificial liver assist devices are currently underway. Based on preliminary data published, this form of liver support therapy might enter the clinic in the near future.

Pig-to-primate transplantations of hDAF-transgenic kidneys achieved survival times of up to 70 days. Pilot projects for clinical kidney xenotransplantation seem to be possible in the near future. The problems of hyperacute rejection seem to be solved. Delayed xenograft rejection remains a serious problem. Further medical, ethical and practical issues have to be solved, before Phase-I-studies can be anticipated. In contrast to other organs, kidney xenotransplantation is associated with low operative risk. Xenotransplant dysfunction or rejection could be treated with conventional hemodialysis.

Transplantation of isolated pancreatic islets provides an interesting alternative to the present cure for diabetes: insulin injections and pumps. These are characterized by an insufficient glucose haemostasis and in the long run can induce kidney failure, blindness, heart failure, and amputations. Up to now more than 293 allogeneic islet transplantations have been performed in diabetics with chronical kidney failure. Despite some success, no real breakthrough has been yet achieved, though great efforts are being made to improve the various methodological steps on the way to clinical transplantation. The use of animal (xenogeneic) organs could be a solution to overcome the shortage of allogeneic donors. The current experimental and clinical research focuses on the use of pigs as organ donors, which have a number of advantages over the immunologically more compatible primates. This article reports on success and open questions concerning the efforts to isolate porcine islets for future clinical transplantation: the search for a suitable pig breed, the various isolation steps, purification and in vitro culture, transplantation models using-small and large animals, first clinical trials, and immunological reactions against the xenogeneic tissue. In addition, strategies to circumvent tissue rejection and future perspectives are discussed.

Hearts of transgenic pigs expressing a human regulator of complement activation, decay accelerating factor (hDAF), were transplanted either heterotopically into the abdomen of cynomolgus monkeys or orthotopically into baboons. None of these transgenic hearts was hyperacutely rejected. Immunosuppression with a combination of cyclosporine A, cyclophosphamide and steroids produced a maximum survival of 62 days (median 40 days) in the heterotopic model. Transgenic hearts transplanted into the orthotopic position allowed a maximum survival of 9 days (median 2.5 days). A more effective and less toxic immunosuppressive protocol for the prevention of accelerated xenograft rejection is the subject of ongoing research. The use of organs from transgenic pigs may help to solve the problem of donor shortage in clinical allotransplantation.

The recent shortage of allogeneic human organs in transplantation medicine has led to a renewed interest in evaluating the feasibility of xenotransplantation, particularly of porcine origin. Discordant xenotransplants between phylogenetically distant species such as pig and primates are challenged by a series of strong rejection reactions. Hyperacute rejection is dominantly mediated by humoral responses of the immune system (natural antibodies, complement cascade) and the activation of coagulation factors, whereas delayed xenograft rejection and T-cell mediated responses are also mediated by elements of the cellular immune system. The mechanisms of chronic xenograft rejection remain to be elucidated. The present review is focused on the mechanisms of rejection and the immunobiology of discordant, vascularized xenotransplants. We attempt to provide a brief description of the dynamics, the humoral and cellular mediators, and the pathogenesis of xenograft rejection. Finally, an overview on potentially therapeutic strategies to overcome xenograft rejection is presented.

The application of high-dose immunoglobulins during xenoperfusion of the isolated rat heart resulted in a significantly higher pressure and heart rate. The coronary flow was also elevated as compared to untreated controls. The heart function during xenoperfusion without treatment was reduced to 50% of the baseline values within 11 min. With immunoglobulin treatment the occurrence of this deficit was delayed to 30 min. A concentrate of immunoglobulins was able to reduce the early functional damage of xenoperfused hearts by interfering with complement related graft destruction.

Islet transplantation is a potential cure for diabetes mellitus. The major problem for clinical application remains the prevention of transplant rejection without major side effects. Broad application in early disease will make the usage of xenogeneic tissue necessary. Immunoisolation is an experimental strategy to prevent rejection, by separating the transplanted allogeneic or xenogeneic cells from the host immune system using a barrier device. Current methods of immunoisolation use artificial, not completely inert materials as barrier devices and induce an unwanted foreign body reaction. Using recipient own cells for encapsulation the foreign body reaction could be prevented. This study describes a new method of encapsulation of islets of Langerhans within a capsule of chondrocytes, which may serve as an immunoisolation barrier utilizing the immunoprivileged properties of the chondrocyte matrix and demonstrates the functional survival of the encapsulated islets in vivo.

The effects of COS7 cells which constitutively express the apoptosis-inducing human CD95 ligand (COSh95L cells) on in vitro alloimmune responses were investigated. In the presence of COSh95L cells a clear-cut inhibition of alloantigen-induced proliferation was observed. In these cultures fully activated blastoid T cells were absent suggesting that deletion of activated T cells is one mechanism of COSh95L-mediated inhibition of proliferation. Non-fully activated cells were not eliminated in MLR by COSh95L cells but achieved a hyporesponsive state as shown by their failure to proliferate in response to restimulation with alloantigen or mitogen. These data indicate that the effects of CD95L on T cells are heterogeneous. Further analysis of the actions of CD95L in T cells of different activation states and subsets will be crucial to estimate the therapeutical potential of this molecule as target for immunomodulation after allo- and xenotransplantation.

The ready availability of xenografts and the promising results of genetic engineering both may offer new methods in peripheral xenotransplantation. FK506 and RS61443 are able to prolong survival of nerve xenografts. The incorporation of adenoviral sequences in xenogeneic tissues is possibly a new option for decreasing the toxicity of immunosuppressive drugs and immunogenicity of grafts.

Kidney xenotransplantation is not yet a realistic clinical treatment modality. However, during the last decades more than 30 kidneys from other species have been transplanted into humans; some of the kidneys sustained some function up to 60 days. Recent progress in genetic engineering has raised the possibility to create large transgenic animals which express human complement regulatory proteins (CRP). Since early complement activation is believed to be the main triggering event for xenograft destruction, complement regulation by species-specific CRP should avoid hyperacute rejection in transspecies transplantation. The perfusion of hDAF-transgenic pig kidneys with human blood was not associated with the morphological signs of hyperacute rejection when compared to non-transgenic control organs. Specific immunohistology could demonstrate that the transgene was sufficient to regulate complement activation beyond C3 despite the endothelial deposition of xenoantibodies. In the future, these organs could be further optimized and ultimately tested in a clinical pilot protocol under appropriate immunosuppression.

Prevention of the occurrence of diabetes-specific vascular complications is the final aim of clinical islet transplantation. Pancreatic islets isolated from adult pigs may be a suitable tissue source to transplant a large number of type 1 diabetic patients. Acute cellular rejection may be finally overcome by clinically applicable protocols for tolerance induction. However, primary nonfunction of the graft, as regularly observed in the porcine islet-to-rat xenotransplantation model, may be an additional problem. In this paper, species-specific inflammatory and immunological mechanisms are discussed which prevent early porcine islet graft function in rats but not in mice.

The worldwide lack of human organ donors puts the pig as potential xenogeneic donor species into the prime of interest. Aim of the present in vitro study is the analysis of T-cell activation in the clinically attractive combination "pig-to-human". Peripheral human blood leukocytes (hPBL) and peripheral porcine blood leukocytes (pPBL) were co-cultured for 4-8 days in the xenogeneic mixed lymphocyte reaction (xMLR) and cell proliferation was measured by 3H-thymidine uptake. Both cell populations were separated into T-cells and antigen presenting cells (APC) to analyze direct and indirect antigen recognition. The results show that (a) activation of human T-cells occurs, (b) the strength of activation depends e.g. on the human responder ("high" and "low" responders), (c) the strength of activation is independent of the responder's HLA-DR status, and (d) direct T-cell activation dominates over indirect activation. Thus, T-cell activation is another immunological barrier that has to be overcome before xenotransplantation can be clinically approached.

Unlabelled: The purpose of this study was to investigate whether the "Combi-Effect" is specific for the transplanted lung tissue or not., Method: In a Guinea-pig to rat model we compared the heterotopic heart-lung transplantation (HLTx, n = 5) with a second heart transplantation (DHTx, n = 5) and a combined heart-kidney transplantation (HKTx, n = 5). Apart from the heart transplant survival time we determined the concentrations of histamine, CH-50, IgG, IgM, leucocytes and thrombocytes in the blood. At time of rejection all tissues were examined histologically and immunohistologically (ED-11, IgG, IgM, OX-39, W3-13, ED-1, NKR-P1, OX-19)., Results: We could achieve a significant prolongation of the heart transplant survival time by combined HLTx compared to HTx (25' to 12', p < 0.01). DHTx showed no effect (7' 53" to 11' 27"). But after HKTx the cardiac survival was even longer than after HTx and HLTx (62.8' to 12' and 25', p < 0.01). CH-50 showed significant lower concentrations after HLTx (180 U/l) and HKTx (178 U/l) than after HTx (260 U/l). Thrombocytes and leucocytes were lower, concentration of histamine higher than after HTx (p < 0.01). Immunohistologically C3 revealed a lower deposition in the rejected heart transplants after combined HLTx/HKTx than after isolated HTx., Conclusion: The "Combi-Effect" is stronger after HKTx than after HLTx. He is not specific for the lung tissue.

Hyperacute graft rejection triggered by the activation of the recipient's complement system represents the major obstacle to successful xenotransplantation. After the binding of preformed antibodies to vascular glycoproteins complement-induced activation and injury of endothelial cells with subsequent thrombosis leads to rapid destruction of foreign tissues. Inhibition of complement activation is therefore considered as a prerequisite for xenograft survival. Recent animal and cell culture experiments suggest that support of the physiological regulation of the complement system appears to be most promising. Besides the application of soluble complement inhibitors (e.g. soluble complement receptor 1, sCR1; C1 inhibitor) the genetic transfer of human membrane-bound complement regulatory proteins (e.g. DAF, CD59) offers new chances to protect the xenograft against the cytolytic complement attack. Results from the authors' experiments shall be included in a short overview to the issue.

The role of parathyroid transplantation for the therapy of permanent hypoparathyroidism is undisputed. Because the parathyroid hormone deficiency syndromee rarely every is a vital thread to patients affected, systemic immunosuppression for transplant recipients is not justified. A technique of microencapsulation was modified for transplantation of parathyroid tissue. Using a core substance suitable for clinical use (amitogenic alginate), we accomplished allotransplantation of functioning parathyroid tissue in the long-term animal model and, very recently, reported first clinical cases without postoperative immunosuppression. In a controlled animal model of totally parathyroidectomized rats (PTX, two groups of n = 40), we investigated the ability of microencapsulation with the amitogenic alginate to enable transplantation across the highest immunological barrier (xenotransplantation: human-rat); to ensure intact transplant function and to protect from rejection. Rat parathyroid hormone (PTHRA i.S.) and serum calcium levels served as parameters of completeness of PTX; intact human PTH (PTHRA i.S.) and serum calcium levels of recipient animals were used to assess graft function. Also, tissue integrity within explanted capsules was assessed by histology. Cultured and microencapsulated parathyroid tissue resumes and maintains function in vivo, even if transplanted across the highest immunological barrier. Functionally, PTHHU i.S. replaced (PTHRA i.S.) in PTX animals entirely and restored normocalcemia. These results suggest, that xeno-transplantation of the parathyroids can be achieved without postoperative immuno-suppression in a long term animal model. These data also imply the possibility of clinical heterotransplantation of parathyroid glands.

This paper describes the rationale behind the need for xenotransplantation and the physiological and immunological barriers associated with xenografting from pig to man. The scientific strategies developed for overcoming the immunological barriers associated with hyperacute rejection are described in detail as is the technology for producing transgenic pigs. Data on perfusion studies of transgenic pig hearts with human blood demonstrates the validity of this scientific approach.

Over the last 5 years an unexpectedly fast progress has been achieved in the field of xenotransplantation. This gives cause for substantial hope. Thus, the wish to use animal grafts for the replacement of human organs could become a reality. However, no results exist at present to suggest a biological and immunological long term acceptance of xenografts in the near future. Xenotransplantation has the goal to improve the organ shortage dramatically and to solve the existing ethical problems of organ donation.

In this study we investigated the functional and morphological properties of xenotransplanted human thyroid tissue in nude mice following systemic application of intrathyroidal lymphocyte preparations from patients with Graves' disease (GD) and non-toxic nodular goiter (NTG). Thyroid tissue samples from 17 NTG-patients were transplanted into athymic nude mice for a period of 4-5 weeks. Aliquots of lymphocyte preparations from both peripheral blood samples (PBL) and thyroid tissue (ITL) of 13 patients with GD and 12 patients with NTG were analyzed by flow-cytometry (CD3, CD4, CD8, CD56) and injected (i.v.) into transplanted nude mice. Animals injected with saline solution served as a control. After 48 h transplants were harvested and histological (H&E) as well as immunohistological evaluation was performed (MHCII, IgG, IgM). Control animals and mice treated with both PBL and ITL from NTG patients showed regular thyroid tissue without lymphocytic infiltrates or local expression of human immunoglobulins. Application of PBL and ITL of GD patients caused scant to moderate lymphocytic infiltrates with detection of human immunoglobulin production. Injection of GD-ITL was accompanied by a significantly higher proportion of intrathyroidal CD3+ lymphocytes and MHCII expression of adjacent thyroid epithelium as compared to injection of GD-PBL preparations. Our results demonstrate that GD-lymphocytes of both peripheral but especially intrathyroidal origin migrate specifically to human thyroid transplants in the nude mouse model, survive for at least two days, secrete immunoglobulins and induce MHCII expression.