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Cardiac xenotransplantation has seen remarkable success in recent years and is emerging as the most promising alternative to human cardiac allotransplantation. Despite these achievements, acute vascular rejection still presents a challenge for long-term xenograft acceptance and new insights into innate and adaptive immune responses as well as detailed characterizations of signaling pathways are necessary. In allotransplantation, endothelial cells and their sugar-rich surface—the endothelial glycocalyx—are known to influence organ rejection. In xenotransplantation, however, only in vitro data exist on the role of the endothelial glycocalyx so far. Thus, in the current study, we analyzed the changes of the endothelial glycocalyx components hyaluronan, heparan sulfate and syndecan-1 after pig-to-baboon cardiac xenotransplantations in the perioperative (n = 4) and postoperative (n = 5) periods. These analyses provide first insights into changes of the endothelial glycocalyx after pig-to-baboon cardiac xenotransplantation and show that damage to the endothelial glycocalyx seems to be comparable or even less pronounced than in similar human settings when current strategies of cardiac xenotransplantation are applied. At the same time, data from the experiments where current strategies, like non-ischemic preservation, growth inhibition or porcine cytomegalovirus (a porcine roseolovirus (PCMV/PRV)) elimination could not be applied indicate that damage of the endothelial glycocalyx also plays an important role in cardiac xenotransplantation. [ABSTRACT FROM AUTHOR]

Summary: The future clinical application of animal‐to‐human transplantation (xenotransplantation) is of importance to society as a whole. Favourable preclinical data relevant to cell, tissue and solid organ xenotransplants have been obtained from many animal models utilizing genetic engineering and protocols of pathogen‐free husbandry. Findings have reached a tipping point, and xenotransplantation of solid organs is approaching clinical evaluation, the process of which now requires close deliberation. Such discussions include considering when there is sufficient evidence from preclinical animal studies to start first‐in‐human xenotransplantation trials. The present article is based on evidence and opinions formulated by members of the European Society for Organ Transplantation who are involved in the Transplantation Learning Journey project. The article includes a brief overview of preclinical concepts and biology of solid organ xenotransplantation, discusses the selection of candidates for first‐in‐human studies and considers requirements for study design and conduct. In addition, the paper emphasizes the need for a regulatory framework for xenotransplantation of solid organs and the essential requirement for input from public and patient stakeholders. [ABSTRACT FROM AUTHOR]

Xenotransplantation using pig organs has achieved survival times up to 195 days in pig orthotopic heart transplantation into baboons. Here we demonstrate that in addition to an improved immunosuppressive regimen, non-ischaemic preservation with continuous perfusion and control of post-transplantation growth of the transplant, prevention of transmission of the porcine cytomegalovirus (PCMV) plays an important role in achieving long survival times. For the first time we demonstrate that PCMV transmission in orthotopic pig heart xenotransplantation was associated with a reduced survival time of the transplant and increased levels of IL-6 and TNFα were found in the transplanted baboon. Furthermore, high levels of tPA-PAI-1 complexes were found, suggesting a complete loss of the pro-fibrinolytic properties of the endothelial cells. These data show that PCMV has an important impact on transplant survival and call for elimination of PCMV from donor pigs. [ABSTRACT FROM AUTHOR]

Abstract: Background: In pig‐to‐human xenotransplantation, early cellular rejection reactions are mediated by natural killer cells (NK cells). Human NK cells are inhibited by HLA‐E via CD94/NKG2A receptors. To protect porcine grafts against human NK cell responses, transgenic GTKO pigs expressing hCD46 and HLA‐E have been generated. The aim of this study was to test the effect of this genetic modification on xenogeneic, and in particular human NK cell response, using an ex vivo perfusion model of pig hearts with human blood. Methods: Cardiopleged and explanted genetically modified (gm) pig hearts (GTKO/hCD46/HLA‐E/hβ2‐microglobulin) and wild‐type (wt) controls (n = 6 each) were reperfused and tested in an 8 hours ex vivo perfusion system using freshly drawn human blood. Cardiac function was evaluated during a 165‐minute period in working heart mode. Myocardial damage, antibody deposition, complement activation, and coagulation parameters were evaluated histologically at the end of perfusion. The number of NK cells in the perfusate was determined by flow cytometry at baseline and at 8 hours; tissue infiltration by NK cells was quantified by immunofluorescence microscopy using NKp46 staining of frozen sections. Results: Deposition of IgG (1.2 ± 1 × 107 vs 8.8 ± 2.9 × 106; P < .01), IgM (4.4 ± 3.7 × 106 vs 1.7 ± 1.2 × 106; P < .01), and the complement activation product C4b/c (3.5 ± 1.3 × 106 vs 2.3 × 106 ± 9.4 × 105; P > .01) was lower in gm than wt hearts. NK cell percentages of leukocytes in the perfusate decreased from 0.94 ± 0.77% to 0.21 ± 0.25% (P = .04) during xenoperfusion of wt hearts. In contrast, the ratio of NK cells did not decrease significantly in the gm hearts. In this group, NK cell myocardial infiltration after 480 minutes of perfusion was lower than in wt organs (2.5 ± 3.7 × 104/mm3 vs 1.3 ± 1.4 × 105/mm3; P = .0001). The function of gm hearts was better preserved compared to wt organs, as demonstrated by higher cardiac index during the first 2 hours of ex vivo perfusion. Conclusion: GTKO, hCD46, and HLA‐E expression in porcine hearts reduced complement deposition, complement dependent injury, and myocardial NK cell infiltration during perfusion with human blood. This tested combination of genetic modifications may minimize damage from acute human‐anti‐pig rejection reactions and improve myocardial function after xenotransplantation. [ABSTRACT FROM AUTHOR]

Abstract: For clinical xenotransplantation, transplants must be free of porcine cytomegalovirus (PCMV). Piglets become infected primarily in the perinatal period by the mother sow. While individual donor animals can be protected from infection by isolation husbandry, success is not guaranteed and this strategy poses the risk of undetected infections and raises animal welfare questions. Here, we present the establishment of a completely PCMV‐negative pig herd for breeding donor animals for xenotransplantation. Eleven pregnant DanAvl Basic hybrid sows were purchased from a designated pathogen‐free (DPF), PCMV‐positive colony and transferred to a new pig facility at the Centre for Innovative Medical Models (CiMM) 4 weeks prior to farrowing. At the age of 24 hours, piglets were early‐weaned and transferred to a commercially available Rescue Deck system dedicated to motherless rearing of piglets. Sows were removed from the facility. The PCMV status of F1‐generation animals was determined at regular intervals over a period of 14 months by a sensitive real‐time PCR‐based detection method testing blood, nasal swabs and cultured peripheral blood mononuclear cells (PBMCs). F1 sows were used as recipients of genetically modified embryos to generate a xenotransplant donor herd. Offspring were tested for PCMV accordingly. All offspring have remained PCMV negative over the whole observation period of 14 months. A completely PCMV‐negative pig herd for xenotransplantation has thus been successfully established. [ABSTRACT FROM AUTHOR]

The porcine cytomegalovirus (PCMV) is a herpesvirus that may pose a risk for xenotransplantation using pig cells, tissues, or organs. Here, three orthotopic pig heart transplantations into baboons were studied. To detect PCMV, a real-time PCR and a Western blot assay based on four PCMV protein sequences, including two tegument proteins, were used. The transmission of PCMV from the donor pig to the recipient baboon was found in two cases, despite PCMV not being detected in the blood of the donor pigs by real-time PCR. Although it was not in the blood, PCMV was detected in different organs of the donor pigs, and in sibling animals. Immunohistochemistry using an antiserum that is specific for PCMV detected virus protein-expressing cells in all of the organs of the recipient baboon, most likely representing disseminated pig cells. Therefore, for the first time, the distribution of PCMV in organs of the donor pigs and the recipient baboons was described. In addition, baboon cytomegalovirus (BaCMV) was found activated in the recipient, and a screening for hepatitis E virus (HEV) and porcine lymphotropic herpesviruses (PLHV) was performed. For the first time, a cross-reactivity between antibodies directed against PCMV and BaCMV was found. [ABSTRACT FROM AUTHOR]

Abstract: Background: Regulatory T cells (Treg) play an important role in maintenance of homeostasis in vivo. Treg application to alleviate allo‐organ rejection is being studied extensively. However, natural Treg (nTreg) expansion in vitro is laborious and expensive. Antigen‐specific Treg are more effective and require lower cell numbers than use of nTreg for immune control. The baboon, as a non‐human primate experimental animal model, is widely used in xenotransplantation research. An effective method to generate baboon xeno‐specific Treg would benefit research on immune tolerance in xenotransplantation using this model system. Method: Baboon tolerogenic dendritic cells (tolDC) were generated in 3 days from monocytes isolated from baboon peripheral blood mononuclear cells in medium supplemented with anti‐inflammatory cytokines. After loading with porcine‐specific (PS) in vitro‐transcribed RNA (ivtRNA), tolDC were used to induce CD4+ T cells to become porcine‐specific Treg (PSTreg) in cocultures supplemented with IL‐2 and rapamycin for 10 days. Anti‐inflammatory and inflammatory cytokine expression was evaluated at the mRNA and protein levels in both baboon tolDC and PSTreg. Functional assays, suppression of activation markers on porcine‐specific effector T cells (PSTeff) and inhibition of PSTeff proliferation, were used to test PSTreg specificity. Results: TolDC generated with this method exhibited a tolerogenic phenotype, expressed CCR7 and produced high levels of IL‐10 and TGF‐β1, whereas IL‐12p40 and IFN‐γ were not expressed. PSTreg were successfully generated in cocultures of CD4+ T cells and PS ivtRNA‐loaded tolDC. They exhibited a CD3+ CD4+ CD25+ CD127low/− CD45RAlow Foxp3+ phenotype and were characterized by high expression of IL‐10 and TGF‐β1 mRNA and protein. They showed upregulated expression of EBI3 and GARP mRNA. PSTreg exhibited highly suppressive effects toward PSTeff, secreting high amounts of IL‐10 and TGF‐β1 cytokine upon interaction with PSTeff and suppressing IFN‐γ expression on PSTeff. Conclusion: In this study, a fast 3‐day method to generate baboon‐derived tolDC is provided that allows subsequent induction of PSTreg displaying high porcine‐antigen specificity and expression of IL‐10 and TGF‐β1. Porcine‐specific baboon Treg can be used in porcine solid organ or cell xenotransplantation studies through adoptive cell transfer into host baboons. [ABSTRACT FROM AUTHOR]

Background: The aim of this study was to establish a new experimental model to directly analyse the coronary microcirculation in cardiac xenografts. Methods: Intravital fluorescence microscopy (IVM) of the subepicardial microcirculation in heterotopically transplanted hamster-to-rat cardiac xenografts was performed at 30 and 90 min of reperfusion. We quantitatively assessed the microcirculatory perfusion characteristics as well as the interactions of leukocytes and platelets with the endothelium of postcapillary coronary venules in non-sensitised as well as sensitised recipients. Results: In this first experimental IVM study of cardiac xenografts, we successfully visualised the subepicardial microcirculation, i.e. feeding arterioles, nutritive capillaries and draining postcapillary venules, during reperfusion. Leukocyte-endothelial and platelet-endothelial cell interactions could be quantified. In the non-sensitised group, the myocardial microcirculation remained stable during the observation period of 90 min, whereas in the sensitised group, xenografts were rejected immediately. Conclusions: We established a model for the assessment of the microcirculatory dysfunction and inflammation during ischaemia/reperfusion injury in hamster-to-rat cardiac xenografts. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Significant progress in understanding and overcoming cardiac xenograft rejection using a clinically relevant large animal pig-to-baboon model has accelerated in recent years. This advancement is based on improved immune suppression, which attained more effective regulation of B lymphocytes and possibly newer donor genetics. These improvements have enhanced heterotopic cardiac xenograft survival from a few weeks to over 2 years, achieved intrathoracic heterotopic cardiac xenograft survival of 50 days and orthotopic survival of 57 days. This encouraging progress has rekindled interest in xenotransplantation research and refocused efforts on preclinical orthotopic cardiac xenotransplantation. [ABSTRACT FROM AUTHOR]

The authors reflect on the guidelines offered by the Xenotransplantation International Society For heart and Lung (ISHLT) Advisory Board. They inform that initial clinic xenogeneic heart transplantation has provided a solution for heart failure patients suffering from severe right ventricular disfunction and coagulation disorders. They mention that xenotransplantation in non-human primates related to their health is time consuming and costly.

Bauer A, Renz V, Baschnegger H, Abicht J-M, Beiras-Fernandez A, Brenner P, Thein E, Schmoeckel M, Reichart B, Christ F. Microcirculatory alterations after orthotopic pig-to-baboon heart transplantation. Xenotransplantation 2011; 18: 232-238. © 2011 John Wiley & Sons A/S. Abstract: Background: Whilst macrohemodynamic function of porcine xenografts transplanted into baboons has been assessed perioperatively, the ability of the xenograft to maintain systemic microcirculatory perfusion has not been investigated after pig-to-baboon xenotransplantation so far. Methods: We investigated the sublingual microcirculation of six baboons undergoing orthotopic transplantation of hCD46-transgenic pig hearts using orthogonal polarization spectral imaging. Microvascular measurements were performed after induction of anesthesia, in the early phase of cardiopulmonary bypass (CPB), during reperfusion of the porcine heart and 1 h after the xenograft had resumed its life-supporting function. Microvascular blood flow was analyzed semiquantitatively and the number of visualized cell-to-cell interactions was counted. Results: The proportion of continuously perfused microvessels was 97 (96 to 97) % at baseline and 95 (94 to 97) % in the early phase of CPB. It decreased significantly (P < 0.05) during CPB to 89 (84 to 91), and alterations were still present (P < 0.05) when CPB was terminated and the xenograft had taken over systemic perfusion 83 (81 to 85) %. The microcirculatory changes correlated with the lactate levels ( y = 18.1-0.18 x; r2 = 0.55; P < 0.001), but no correlation with macrohemodynamic parameters was found. Conclusion: Microvascular blood flow is altered after orthotopic pig-to-baboon heart transplantation, despite systemic hemodynamic parameters being well maintained by the porcine xenograft. These changes are moderate but persist after termination of CPB. Further studies need to elucidate whether these changes are transient or add to the mortality associated with cardiac xenotransplantation. [ABSTRACT FROM AUTHOR]

Background: Cytotoxic anti-Galα(1,3)Gal antibodies play a key role in the rejection of pig organs transplanted into primates. Regimens reducing anti-Galα(1,3)Gal antibodies were associated with severe side effects unable to prevent antibody rebound until soluble synthetic oligosaccharides with terminal Galα(1,3)Gal inhibiting antigen binding became available. We displayed kinetics of anti-pig and anti-Galα(1,3)Gal IgM and IgG antibody levels using GAS914, a Galα(1,3)Gal trisaccharide conjugated to poly-l-lysine, and investigated corresponding changes of parameters of heart function. Methods: Using a working heart model, hDAF pig hearts were perfused with human blood containing GAS914 (group 1). As controls hDAF pig hearts (group 2) and landrace pig hearts (group 3) were perfused with human blood only. Levels of anti-Galα(1,3)Gal (IgM, IgG) and anti-pig antibodies were assessed to prove the effectiveness of GAS914. As parameters of heart function, cardiac output (CO), stroke work index (SWI), coronary blood flow (CBF) and coronary resistance were measured. Creatine phosphokinases, lactate dehydrogenase and aspartate aminotransferase were evaluated as markers of myocardial damage. Histological and immunohistochemical investigations were performed at the end of perfusion. Results: In group 1 an immediate and extensive reduction in both IgM and IgG anti-Galα(1,3)Gal was found. Anti-pig antibodies were eliminated accordingly. Antibody binding to GAS914 was complete before the start of organ perfusion. Corresponding to rapid antibody elimination in group 1 GAS914 not only was able to significantly prolong the beating time of the heart in hDAF pigs, but also to clearly improve functional parameters. When switching to the working heart mode hDAF pig hearts perfused with human blood containing GAS914 (group 1) revealed a CO starting at a significantly higher level than hDAF (group 2) and non-transgenic pig hearts (group 3) perfused with human blood only. Similarly, in group 1 SWI was significantly increased at the beginning of perfusion compared to that of group 2 and group 3. The increase in CBF during perfusion and the corresponding fall of coronary resistance occurred without significant differences between the groups revealing the independence of hDAF and GAS914. Conclusions: Due to an immediate and profound reduction in Galα(1,3)Gal-specific antibodies, soluble Galα(1,3)Gal conjugates not only prolong survival, but also improve the hemodynamic performance of the heart in DAF pigs. [ABSTRACT FROM AUTHOR]

Background: Only limited data are available on the physiological functional compatibility of cardiac xenografts after orthotopic pig to baboon transplantation (oXHTx). Thus we investigated hemodynamic parameters including cardiac output (CO) before and after oXHTx. Methods: Orthotopic xenogeneic heart transplantation from nine hDAF transgeneic piglets to baboons was performed. We used femoral arterial thermodilution for the invasive assessment of CO and stroke volume. Results: Baseline CO of the baboons after induction of anesthesia was 1.36 (1.0–1.9) l/min. 30 to 60 min after termination of the cardiopulmonary bypass, CO of the cardiac xenograft was significantly increased to 1.72 (1.3–2.1) l/min (P < 0.01). The stroke volumes of the baboon heart before transplantation and the cardiac xenograft was comparable [14.9 (11–26) vs. 11.8 (10–23) ml]. Thus the higher CO was achieved by an increase in heart rate after oXHTx [75.0 (69–110) vs. 140.0 (77–180)/min; P < 0.01]. Despite the increased CO, oxygen delivery was reduced [256 (251–354) vs. 227 (172–477) ml/min; P < 0.01] due to the inevitable hemodilution during the cardiopulmonary bypass and the blood loss caused by the surgical procedures. Conclusion: Our results demonstrate that in the early phase after orthotopic transplantation of hDAF pig hearts to baboons, cardiac function of the donor heart is maintained and exceeds baseline CO. However, in the early intraoperative phase this was only possible by using inotropic substances and vasopressors due to the inevitable blood loss and dilution by the priming of the bypass circuit. [ABSTRACT FROM AUTHOR]

A review is presented of the article "Meta-analysis of the independent and cumulative effects of multiple genetic modifications on pig lung xenograft performance during ex vivo perfusion with human blood" by D.G. Harris and colleagues.

Background Cell injury, particularly as consequence of ischemia/reperfusion or acute rejection after (xeno)transplantation, leads to release of intracellular components like RNA. Extracellular RNA, (i) causes edema as a result of increasing permeability of blood vessels [1], (ii) is described as a procoagulation factor by activation of the contact phase system leading to thrombus formation and vessel occlusion [2], and (iii) promotes leukocyte recruitment to the vascular system by mobilising proinflammatory cytokines [3], and thus, triggers immune response. In rodent models of stroke and thrombosis, treatment with counteracting RNase A, a ribonuclease that specifically degrades single-stranded RNA, was shown to result in less edema formation and vessel occlusion [4]. In search of additive drugs to protect (xeno)grafts from dysfunction, we investigated if treatment with RNase A in a rat model of heterotopic heart transplantation results in less edema and less coronary occlusion, and, therefore, in improved graft survival. Methods Brown Norway rat cardiac allografts were transplanted into the abdomen of Lewis rats after perfusion with Bretschneider cardioplegic solution. Microvascular technique for aorto-aortic anastomosis and pulmonary artery to inferior vena cava anastomosis (Langendorff) was used. Recipients were intravenously treated directly before transplantation and every other day with RNase A (50 μg/kg) or vehicle (saline, n = 6 in each group). In addition, in six cases the donor graft was perfused with Bretschneider solution containing RNase A (8 μg in 10 ml), and afterwards transplanted into RNase A treated recipients. The primary end point of the study was graft survival, which was determined by daily examination of the graft function by palpation and in case of inconclusiveness by echocardiography. A tolerance study was performed treating animals daily with 50 μg/kg of RNase A, with 1,000 μg/kg of RNase A, or vehicle, respectively, for 28 days (n = 3 in each group). All explanted hearts were collected for histological evaluation. Results Mean graft survival was 6.5 ± 0.55 day (range 6-7 day) in vehicle treated animals vs. 10.2 ± 1.0 day (range 9-11 day) in RNase A treated animals (P = 0.002). Directly after explantation donor grafts had a mean weight of 0.94 ± 0.03 g (range 0.89-0.98 g). After been rejected, donor graft weight increased to 2.06 ± 0.52 g (range 1.59-2.77 g) in vehicle treated recipients, in contrast to 1.28 ± 0.27 g (1.05-1.61 g) in RNase A treated recipients (P = 0.017). No supplemental benefit was observed by additive using of RNase A in Bretschneider solution (graft survival 9.50 ± 1.52 day, graft weight 1.38 ± 0.15 g) compared to single treatment of the recipient (graft survival P = 0.394, graft weight P = 0.589). Tolerance studies showed neither macroscopic nor histological differences compared to saline treated animals. Outlook: Further histological analyses will help to clarify if RNase A will keep the promise to reduce edema formation even in the field of transplantation, as suggested by the significant less graft weight in RNase A treated recipients. Moreover, we will investigate, if RNase A prevents leukocyte recruitment in the context of rejection reaction. Therefore, grafts of RNase A treated and untreated rats will be explanted on day 4 after transplantation. Conclusions RNase A significantly improved graft survival. On the basis of these sweeping results, however, we suppose that RNase A could be an important adjuvant drug not only in allotransplantation but even in xenotransplantation. References [1] Fischer S, Gerriets T, Wessels C, et al. Extracellular RNA mediates endothelial-cell permeability via vascular endothelial growth factor. Blood 2007; 110(7): 2457-2465. [2] Kannemeier C, Shibamiya A, Nakazawa F, et al. Extracellular RNA constitutes a natural procoagulant cofactor in blood coagulation. Proc Natl Acad Sci U S A 2007; 104(15): 6388-6393. [3] Fischer S, Grantzow T, Pagel JI, et al. Extracellular RNA promotes leukocyte recruitment in the vascular system by mobilising proinflammatory cytokines. Thromb Haemost. 2012; 108(4): 730-741. [4] Walberer M, Tschernatsch M, Fischer S, et al. RNase therapy assessed by magnetic resonance imaging reduces cerebral edema and infarction size in acute stroke. Curr Neurovasc Res. 2009; 6(1): 12-19. [ABSTRACT FROM AUTHOR]

It is more than 1 year since our Consortium commenced. The following abstracts provide an account of what has been achieved so far and document our understanding of xenotransplantation and some of our future strategies: Since (microencapsulated) islets are the first in the clinic, our basic research focuses on immediate (innate) rejection reactions and means of counteracting them. Once transplanted and working, porcine and primate islets may respond differently, for example to glucose challenges., We will gain preclinical experience regarding the safety of genetically modified islets as quickly as possible., We are currently in dialogue with German authorities (Paul-Ehrlich-Institution as representative of the European Medicines Agency) to decide how best to formulate a successful clinical application within European standards., Are there any ethical objections from within German society? A recent 3-day meeting, held at the Munich Catholic Academy explored the issues, including opinions from the Jewish and Islamic communities. In brief, the outcome revealed no major objections as long as xenogeneic procedures are safe and effective. All talks will be published in German, with abstracts in English. There will also be an English review of the meeting possibly published in the journal Xenotransplantation., Successful preclinical organ (heart, kidney) xenotransplantation is still difficult to achieve. Heterotopic thoracic (working) heart transplantation has been established with consistent results. It seems that hyperacute and delayed humoral rejection reactions are manageable; but thrombotic microangiopathy currently presents the main obstacle. Porcine hearts that express human thrombomodulin will be used next, together with complement antibodies and/or co-stimulation blockade. Human thrombomodulin has provided promising results in the laboratory (cell based)., Porcine heart valves will be tested using single and multiple antigen knock out., In conclusion, the German Xenotransplantation Consortium supported by the German Research Foundation (DFG) is on track for a successful (first?) period. There seems to be broad support from a majority of German cardiovascular surgeons, and this is spreading to those in other parts of Europe [1]. Reference [1] Reichart B., Guethoff S., Mayr T. et al. Discordant cardiac xenotransplantation - broadening the horizons. Eur J Cardiothorac Surg 2013; Epub ahead of print. [ABSTRACT FROM AUTHOR]

Discordant xenotransplantation research started in Hanover/Mariensee and in Munich in the late nineties - the latter receiving a substantial grant from the Bavarian Research Foundation. In 2003 both groups were invited by the German Research Foundation (DFG) to make a joint application to further develop xenotransplantation technology. This was peer-reviewed, approved and continued with considerable success from 2004 to 2012. The Paul-Ehrlich and Robert-Koch Institutes, the official bodies responsible for microbiological safety in Germany, were subsequently included. Our unique interdisciplinary consortium combines basic scientists, animal biotechnologists and clinicians demonstrating an excellent co-operation over the last 8 years. The tool box available to us includes pigs transgenic for human thrombomodulin, hemoxygenase-1, A20, LEA 29Y, HLA-E and TRAIL; key enabling technologies include gene targeting, large transgene vectors, transposon mediated transgenesis, zinc finger and TALE nuclease mediated gene targeting. In July 2012 our new Collaborative Research Centre (again supported by the DFG) commenced, continuing in the first instance to 2016 and ultimately until 2024. The strong Diabetes group at the University of Dresden, which currently runs the only successful allogeneic pancreatic islet transplantation in Germany, has now been incorporated. Our main goal is to translate preclinical studies into benefits for patients as quickly as possible. The consortium focuses on islet, cardiac and renal xenotransplantation; new biological heart valves for paediatric cases and young adults will be developed. Our endeavours will be supported by two ethicists to ensure best practice and promote public dialogue. [ABSTRACT FROM AUTHOR]