Trotz Inkrafttretens des ersten gesamtdeutschen Transplantationsgesetzes im Dezember 1997 steigt die Anzahl terminal niereninsuffizienter Patienten auf der Warteliste, ebenso wie die Wartezeit auf eine Nierentransplantation (NTX). Verschiedene Programme existieren, um der Organknappheit zu begegnen. Im Eurotransplant Senior Programm (ESP) werden seit 1999 Nieren älterer verstorbener Spender (65+) an ältere Empfänger (65+) unabhängig von der Gewebeübereinstimmung auf lokaler Ebene übertragen. Unsere Studien konnten zeigen, dass die Transplantat- und Empfängerüberlebensraten sich nicht von denen nach Transplantation jüngerer Spendernieren in alte Empfänger unterscheiden. Transplantatverlust im ESP tritt meist als Tod mit funktionierendem Transplantat auf. Dies ist die Folge vermehrt vorkommender Komorbiditäten in dieser Empfängergruppe und unterstreicht die Notwendigkeit einer intensiven prä-operativen Evaluation sowie engmaschigen Nachsorge. Zudem konnten wir zeigen, dass die Grundannahme einer verminderten Immunkompetenz älterer Empfänger nicht zutrifft. Eine Änderung der Allokationsregeln von Eurotransplant wird deshalb diskutiert, die auch im ESP die Gewebetypisierung berücksichtigt. Der Anteil der Lebendnierenspende (LNTX) in Deutschland hat sich in den vergangenen Jahren auf ca. 19% verdreifacht. An unserer Klinik macht sie ca. 30% der durchgeführten NTX aus. Zudem wird an unserer Klinik deutschlandweit die laparoskopische Spendernierenentnahme am häufigsten durchgeführt. Folge einer hier durchgeführten prospektiven Studie zur präoperativen Bildgebung der renalen Gefäßarchitektur des Spenders ist der Verzicht auf invasive Methoden. In weiteren Untersuchungen konnten wir zeigen, dass die Akzeptanz von Lebendspendern mit erweiterten Spenderkriterien gute Möglichkeiten bieten NTX-Programme zu intensivieren. In einer Lebensqualitätsstudie untersuchten wir erstmalig für den deutschen Sprachraum die postoperativen Verläufe von Lebendnierenspendern und konnten neben einer grundsätzlichen Zufriedenheit auch besondere Bedürfnisse aufdecken. Verschiedene Studien zur Laparoskopie in der LNTX, Handhabung und Optimierung dieses neuen Verfahrens sowie dessen Anwendbarkeit bei anatomischen Besonderheiten konnten zeigen, dass diese Methode ein sicheres Verfahren darstellt. Anhand unserer Erfahrungen in der LNTX haben wir zudem Empfehlungen zur Evaluation und Nachbetreuung von Lebendnierenspendern erarbeitet. Insgesamt sind das ESP und die LNTX, hier besonders die laparoskopische Spendernierenentnahme, effektive Konzepte zur Erweiterung der Nierentransplantationsprogramme. / Despite the implementation of a transplantation law in 1997 the number of patients with end-stage renal disease in Germany is rising, as is the waiting time for a kidney transplantation (KTX). Different programs exist to counterbalance organ shortage. The Eurotransplant Senior Programm (ESP), initiated in 1999, allocates kidneys of deceased donors aged 65 and older (65+) irrespective of tissue matching locally to recipients 65+. Our studies revealed that graft- and patient survival in this program are comparable to that after transplantation of young donor-kidneys in old recipients. Graft loss is mostly due to recipient death, which is a consequence of the co-morbidities of the recipient. Meticulous evaluation and intensified follow-up in this recipient-group are most important. Furthermore, despite the assumption of a reduced immunologic response of old recipients we could show that they are very well immunocompetent. Therefore, a change in allocation rules at Eurotransplant is discussed, possibly involving HLA-matches in the ESP. Living donor kidney transplantation (LDKTX) has increased three-fold over the last years and features about 19% of all hat KTX in Germany. In our department LDKTX features about 30% of all KTX and our department also performs the most laparoscopic explantations of living kidney donors in Germany. With regard to the preoperative donor evaluation we could show in a prospective study that non-invasive methods for renal vessel imaging are better than the standard invasive approach. Further research proved that applying extended donor criteria renders good results for graft and recipient and thus is a viable option to increase LDKTX. In the first German study on donors Quality of Life we mostly found satisfied donors and also could detect specific donor demands. For the laparoscopic approach we performed different studies on the technique, its improvement and peculiarities, proving that laparoscopic organ retrieval is a safe method in LDKTX. ESP and LDKTX, especially the laparoscopic organ retrieval, are effective concepts to increase the number of kidney transplantations.

Il est prouvé que la conservation des greffons rénaux marginaux en machine de perfusion (MP) est bénéfique. Cependant, cette méthode nécessite des améliorations afin de minimiser les lésions d’ischémie-reperfusion (I/R), par l’ajout d’oxygène et/ou d’un transporteur d’oxygène. Nous avons cherché à évaluer les effets de l’oxygénation et de l’ajout d’une hémoglobine de ver marin (HbAm, M101) durant la perfusion rénale hypothermique avant transplantation. Nos critères de jugement étaient basés sur la reprise de fonction du greffon et sur les lésions tardives de dysfonction rénale. Nous avons utilisé un modèle porcin : les reins ont été exposés à 1h d’ischémie chaude, puis perfusés dans une MP WAVES® pendant 23h à 4°C avant autotransplantation. Quatre groupes ont étudié : W (MP-21% O2), W-O2 (MP-100% O2), W-M101 (MP-21% O2 + 2g/L HbAm), W-O2+M101 (100% O2 + 2g/L HbAm), (n=6 per groupe). Les reins du groupe W-M101 ont montré un débit de perfusion plus élevé et une résistance rénale plus faible comparé aux autres groupes. Pendant la première semaine post-transplantation, les groupes W-O2 et W-M101 ont montré une créatininémie significativement plus faible et un meilleur taux de filtration glomérulaire (GFR). Les niveaux circulants de KIM-1 et IL-18 étaient plus faibles dans le groupe W-M101, tandis que les niveaux de NGAL et d’ASAT étaient plus faibles dans les groupes d’oxygénation active. Trois mois post-transplantation, la fraction excrétée de sodium et le ratio protéinurie/créatininurie étaient plus élevé dans le groupe W. La créatininémie était plus faible dans le groupe W-M101. La fibrose interstitielle a évalué à 3 mois post-transplantation étaient plus faible dans les groupes W-M101 et W-O2+M101. Nous avons révélé histologiquement que l’infiltration de mastocytes était significativement élevée dans le groupe W comparé aux autres groupes. Nous avons montré que la combinaison de 21% O2 + hémoglobine améliorent la reprise de fonction du greffon rénale. / Introduction: It is proved that preservation of marginal kidney graft in machine perfusion (MP) is beneficial. However, this method requires improvement to minimize the ischemia-reperfusion injuries (IRI), as addition of oxygen and/or an oxygen carrier. We aimed to evaluate the effects of oxygenation (100% or 21%) and the addition of marine worm hemoglobin (HbAm, M101) during hypothermic renal perfusion before transplantation. Our endpoints were based on graft function recovery and late renal dysfunction. Method and materials: We use a porcine model where kidneys were submitted to 1h warm ischemia, followed by WAVES® MP preservation for 23h before auto-transplantation. Four groups were studied: W (MP-21% O2), W-O2 (MP-100% O2), W-M101 (MP-21% O2 + 2g/L HbAm), W-O2+M101 (100% O2 + 2g/L HbAm), (n=6 per group). Results: Kidneys preserved in W-M101 group showed a higher perfusion flow and lower renal resistance, compared to other groups. During the first week post-transplantation, W-O2 or W-M101 groups showed lower blood creatinine and better glomerular filtration rate. Blood levels of KIM-1 and IL-18 were lower in W-M101 group, while blood levels of AST and NGAL were lower in groups with 100% O2. Three months after transplantation, the fractional excretion of sodium and the proteinuria/ creatininuria ratio were higher in W group. Blood creatinine was lower in W-M101 group. Interstitial fibrosis evaluated at 3 months was lower in groups W-M101 and W-O2+M101. We showed that the combination 21% O2 + hemoglobin improves the kidney graft outcome.Conclusion: We showed that the combination of 21% O2 + hemoglobin improved the kidney graft outcome.

Estudos com células tronco mesenquimais (CTm) têm despertado grande interesse devido a seu promissor potencial terapêutico e representam uma alternativa para o tratamento de diversas patologias em diferentes órgãos, inclusive em transplante renal. A rejeição crônica é um dos maiores desafios no transplante tardio e se caracteriza por perda progressiva da função renal causado pela intensa fibrogênese no aloenxerto. Os tratamentos convencionais com imunossupressores, apesar de reduzirem significativamente as crises de rejeição aguda, não interferem na sobrevida do enxerto a longo prazo. A compreensão dos processos fisiopatológicos da doença depende de seu estudo em modelos experimentais, que são de grande importância pois também propiciam uma melhor compreensão dos possíveis tratamentos. O presente estudo teve como objetivo analisar a terapia com células-tronco mesenquimais derivadas de tecido adiposo (CTmTA) no modelo experimental de transplante renal em ratos, para estudar seu efeito na rejeição crônica e avaliar seu potencial efeito imunomodulador. O modelo foi estabelecido com ratos das linhagens isogênicas Fisher (doador) e Lewis (receptor) e os animais transplantados foram divididos em três grupos: ISO (transplante isogênico de Lewis para Lewis, n=6), ALO (transplante alogênico de Fisher para Lewis, n=6) e ALO+CTmTA (transplante alogênico, tratado com CTmTA, n=6). As CTmTA foram caracterizadas por aderência ao plástico, diferenciação nas linhagens adipogênica, condrogênicas e osteogênicas e por citometria de fluxo. Foram inoculadas 1 x 106 células na região subcapsular renal no dia da realização da nefrectomia unilateral direita (10 dias pós-transplante). Após 6 meses foram realizadas análises dos parâmetros clínicos e laboratoriais, além de análise histológica, imunohistoquímica e PCR em tempo real. As CTmTA foram eficientes em prevenir significativamente a elevação da ureia e da creatinina séricas, manter clearence de creatinina em níveis normais, e prevenir a elevação da fração de excreção de Na+ e K+. Além disso, impediram o desenvolvimento de proteinúria e da hipertensão arterial. A análise histológica mostrou uma redução significativa do infiltrado inflamatório de macrófagos e linfócitos T, além de uma diminuição da fibrose intersticial no grupo ALO+CTmTA. O tratamento com CTmTA reduziu significativamente a expressão relativa dos fatores e citocinas pró-inflamatórios tais como INF-y, TNF-alfa, IL1beta e IL-6, além de aumento importante na expressão de IL-4 e IL-10, conhecidas por seu potencial antiinflamatório. Em conclusão, o tratamento com ADMSC em um modelo experimental de transplante renal pode trazer uma nova abordagem terapêutica para controle da rejeição crônica do enxerto. A aparente modulação da resposta imune observada neste trabalho, pode estar associada a uma possível polarização de macrófagos e células T. Outros estudos pré-clínicos e clínicos são necessários para confirmar nossos resultados / Studies involving mesenchymal stem cells (MSCs) have aroused great interest due to their promising therapeutic potential representing an alternative for the treatment of several pathologies in different organs, including renal transplantation. Chronic rejection is one of the major challenges in late transplantation and is characterized by progressive loss of renal function caused by intense fibrogenesis in the allograft. Conventional immunosuppressive treatments, while significantly reducing acute rejection crises, do not interfere with long-term graft survival. Animal model of kidney transplantation can provide a better understanding of the pathophysiological processes and bring a new path to treat chronic rejection. The aim of this project was to analyze the therapy with mesenchymal stem cells derived from adipose tissue (ADMSCs) in the experimental model of kidney transplantation in rats, focus on chronic rejection and evaluate its potential immunomodulatory effect. The model was established with rats of isogenic strains Fisher (donor) and Lewis (recipient), and the transplanted animals were divided into three groups: ISO (isogenic transplantation from Lewis to Lewis, n = 6), ALO (allogenic transplant from Fisher to Lewis, n = 6) and ALO + ADMSCs (allogenic transplantation, treated with ADMSCs, n = 6). ADMSCs were characterized by adhesion to plastic, differentiation in adipogenic, condrogenic and osteogenic lines and by flow cytometry. One million of cells were inoculated under the renal capsule on the day of the right unilateral nephrectomy (10 days after transplantation). After 6 months, clinical and laboratory parameters were analyzed, as well as histological analysis, immunohistochemistry and real-time PCR. ADMSCs were effective in preventing elevation of serum urea and creatinine, elevation of the Na + and K + excretion fraction as well as maintained creatinine clearence at normal levels. Furthermore, the treatment also prevented the development of proteinuria and preserved blood pressure. Histological analysis showed a significant reduction of macrophages and T cells infiltrate, associated to a decreased of interstitial fibrosis in the ALO + ADMSCs group. In the presence of ADMSCs, there was a significant decrease in the relative expression of INF-y, TNF-alpha, IL1beta and IL-6 factors and pro-inflammatory cytokines, as well as a significant increase in the relative expression of anti-inflammatory cytokines as IL-4 and IL-10. In conclusion, treatment with ADMSC in a transplantation model could open a new approach to control chronic rejection. This apparent modulation of the immune response may be associated with a possible polarization of macrophages and T cells. Further pre-clinical and clinical studies are needed to confirm our findings

A doença cardiovascular (DCV) é uma condição clínica comum entre pacientes (pcts) portadores de doença renal crônica (DRC) e é causa de eventos fatais observados peri transplante renal (TX). A melhor estratégia de avaliação cardiovascular em candidatos a transplante (CTR) ainda é controversa.Ignora-se se todos os pacientes devem ser submetidos a testes não-invasivos/invasivos ou se estes devem ser reservados aqueles com determinadas características clínicas, como população geral. O objetivo deste estudo foi comparar a estratificação de risco baseada em método nãoinvasivo de detecção de doença coronária com dois métodos de estratificação clínica de risco cardiovascular preconizados pela American Society of Transplantation (AST) e European Renal Association (ERA). A AST subdivide os pcts em : alto risco (idade maior ou igual a 50 anos e/ou diabete e/ou DCV clínica) e baixo risco (os demais). A ERA subdivide em: alto risco (DCV clínica), risco intermediário (diabéticos e/ou idade maior ou igual a 50 anos) e baixo risco (os demais). Nós estudamos 386 pcts com DRC em diálise enviados ao nosso serviço para avaliação cardiovascular antes da inclusão na lista de espera de TX. Foram estratificados quanto ao risco de eventos de acordo com os dois algoritmos acima e alterações na cintilografia de perfusão miocárdica (SPECT-MIBI) com dipiridamol e acompanhados até a morte, TX ou ocorrência de eventos. A estratificação clínica (RR:1,8 [IC95% 1,3 2,6- P<0,0001] e o SPECT-MIBI (RR:1,5 [IC95% 1,2-1,9-P=0,002] identificaram os pcts de maior risco de eventos cardiovasculares . Apenas os pcts ASTalto risco (RR1,4 [IC95%1,1-1,8-P=0,002] e ERA médio risco com SPECTMIBI alterado (RR:1,7 [IC95% 1,2-2,3-P=0,003] tiveram maior incidência de eventos. Os pcts de baixo risco pelos dois algorítmos de estratificação clínica (P=0,50) e do sistema ERA alto risco (RR:1,1 [IC95% 0,8-1,5-P=0,41], não se beneficiaram dos resultados do estudo não-invasivo. Concluímos que os estudos não-invasivos não devem ser utilizados em todos os CTR mas devem ser reservados aos pcts previamente identificados pela estratificação clínica de risco. Esses resultados permitem uma abordagem mais racional da avaliação pré- TX com melhor uso dos recursos econômicos escassos. / Cardiovascular (CV) disease is a common condition in chronic kidney disease (CKD) patients and is the leading cause of fatal events during and after renal transplantation. The best strategy for CV evaluation and coronary risk stratification in renal transplant candidates remains controversial. Moreover, there is no consensus regarding the best strategy for detection of coronary artery disease (CAD). We still do not know if all patients should be evaluated by noninvasive testing or if this approach should be restricted to individuals with clinical evidence of CAD, as in the general population. The objective of this study was to compare CV risk stratification based on nonivasive testing for CAD with two clinical stratification methods as advanced by The American Society of Transplantation (AST) and by The European Renal Association (ERA), respectively. The AST divides patients in high risk (age50 years and/or diabetes and/or CV disease) and low risk (all others).The ERA divides : high risk (CV disease), intermediate risk (age 50 years and/or diabetes), and low risk (as above). We studied 386 CKD patients treated by hemodyalisis, to CV evaluation before being admitted to the renal transplant waiting list. All patients were stratified for the risk of future major cardiovascular events (MACE) using the clinical algorithms and also by myocardial scintigraphy (SPECT-MIBI) with dipyridamol and followedup until death, transplant or MACE. Clinical algorithms (RR:1,8 [IC95% 1,3 2,6-P<0,0001] and SPECT-MIBI(RR:1,5 [IC95% 1,2-1,9-P=0,002] identified patients at increased risk of events. The combined use of clinical stratification followed by SPECT showed that the only patients that would benefit from SPECT risk stratification were those belonging the AST-high risk (RR1,4 [IC95%1,1-1,8-P=0,002] and ERA-intermediate risk groups (RR:1,7 [IC95% 1,2-2,3-P=0,003]. In all other groups :ERA-high-risk (RR:1,1[IC95% 0,8-1,5- P=0,41] and ERA and AST-low-risk (P=0,50) SPECT did not add to the probability of events defined by clinical stratification alone. We conclude that SPECT should not be applied to all renal transplant candidates but should be restricted to those considered at a category of risk as defined by clinical algorithms. These results delineate a more rational approach to risk stratification in renal transplant candidates with a better utilization of economical resources.

Contexte : La réponse allo-immune induite par la transplantation à partir d'un donneur génétiquement différent est un obstacle majeur au succès de la greffe. Notre objectif est de caractériser les différents phénotypes de rejet d'allogreffe rénale et d'identifier la façon dont chacun est associé aux anticorps anti-HLA. Nous avons également évalué l’impact de certaines propriétés de ces anticorps, comme leur intensité ou leur capacité à fixer le complément, sur l'échec des allogreffes rénales. Pour finir, nous avons étudié l’impact pronostic des formes indolentes de rejets ainsi que l’apport des nouvelles technologies d’analyses transcriptomique des biopsies de patients transplantés. Méthodes : Nous avons utilisé une approche en population, basée sur l’étude de larges cohortes de receveurs de greffes rénales. L’étude concomitante des données immunologiques et histologiques, nous a permis de corréler les caractéristiques des anticorps anti-HLA circulants aux phénotypes lésionnels. Résultats : Nous avons identifié et caractérisé 4 types distincts de rejet : les rejets vasculaires médiés par les lymphocytes T (9%) et par les anticorps (21%), non reconnus par les classifications internationales, et les rejets cellulaires (46%) et humoraux sans vascularite (24%). Le risque de perte de greffons est le plus important dans les cas de rejet vasculaire médié par anticorps. Les anticorps dirigés contre le donneur (DSA) fixant le complément induisent un phénotype histologique plus sévère, dominé par des lésions inflammatoires et plus de dépôts de la fraction C4d du complément dans les greffons. En leur présence, le risque de perte de greffons est augmenté de 3,7 fois (IC95 1,9-7,2). Les formes indolentes de rejet médié par les anticorps sont également associées à un risque accru de perte du greffon. L’utilisation d’approches moléculaires permet d’améliorer la stratification du risque au sein du groupe des patients présentant des rejets humoraux. Conclusion : Ce travail répond à un besoin clinique pressant dans le domaine de la transplantation, celui de déterminer l’impact clinique des anticorps anti-HLA et d’améliorer la stratification du risque immunologique en se basant sur leurs propriétés et l’utilisation de nouvelles technologies pour mieux caractériser l’activité et le stade des rejets humoraux. / Background : The alloimmune response induced by transplantation from a donor who differs genetically from the kidney recipient has always been the major obstacle to graft success. The present work aimed to improve characterization of kidney-allograft rejection phenotypes and identify how each one is associated with anti-HLA antibodies. We also sought to determine whether characteristics of these antibodies i.e., their levels or complementbinding ability, might play a role in kidney allograft failure. Finally, we evaluated the clinical relevance of indolent forms of ABMR and the clinical relevance of new genes expression technologies to stratify the kidney recipients at risk for failure. Methods : We used a population-based approach in precisely phenotyped cohorts of kidney recipients. The design of our study, which is based on the concomitant evaluation of immunologic and histologic data, permits a precise connection of circulating anti-HLA antibodies with a phenotype of graft injury. Findings : We identified four distinct patterns of kidney allograft rejection: T cell-mediated vascular rejection (9%), antibody-mediated vascular rejection (21%), not included in international classifications, T cell- (46%) and antibody-mediated rejection without vasculitis (24%). Risk of graft loss was 9.07 times (95CI 3.6-19.7) higher in antibody-mediated vascular rejection than in T-cell mediated rejections (p<0.0001). Patients with post-transplant complement-binding DSA had more severe graft injury phenotype with higher inflammation and increased deposition of complement fraction C4d. They have the poorest graft survival with 3.7 fold increased risk of graft loss (95CI 1.9-7.2). Subclinical ABMR is a truncated for of rejection associated with risk of kidney allograft failure. Gene expression assessment in kidney allografts with early ABMR improves classification of individuals at risk for kidney allograft loss. Conclusion : This work addresses the unmet need of the deleterious impact of anti-HLA antibodies and the improvement of risk stratification in kidney transplantation. Recognition of distinct phenotypes could lead to the development of new treatment strategies. Gene expression assessment appears useful to evaluate disease activity, disease state and prediction of failure.

<p>Anemia is common in patients with chronic kidney disease (CDK), contributes to reduced Quality of Life (QoL) and is associated with cardiovascular disease, morbidity and mortality. Epoetin raises hemoglobin (Hb) and increases QoL and physical exercise capacity. Because of concerns about safety and economics, current anemia treatment with epoetin aims to achieve subnormal Hb (110-120 g/l). Normalization of Hb may be of additional benefit regarding QoL and cardiovascular effects. The present study examines the effects of Hb normalization with epoetin on safety variables, QoL, graft function after kidney transplantation, dialysis adequacy, hemorheology, hemodynamics and cardiac autonomic function in CKD patients. </p><p>In a randomized, multicenter study comprising 416 pre-dialysis and dialysis patients no difference was observed between patients treated to a normal or a subnormal Hb level on mortality, thrombovascular events, serious adverse events, vascular access thrombosis and residual renal function. QoL was enhanced in a subgroup of hemodialysis patients. Pretransplant epoetin treatment directed toward normal Hb levels did not result in worse graft function during 6 postoperative months. Dialysis adequacy was reduced in a subgroup of hemodialysis patients after normalization of Hb. The blood flow properties of pre-dialysis patients were altered. The hemorheological investigation demonstrated that Hb normalization caused a parallel increase in hematocrit and blood viscosity without other hemorheological changes. While the total peripheral resistance index increased, the cardiac index (CI) decreased. In a separate study cardiac autonomic function, measured by heart rate variability, was decreased in pre-dialysis patients. It was improved, but not fully normalized, by Hb normalization. </p><p>On the basis of this study, Hb normalization with epoetin appears to be safe and increases QoL in hemodialysis patients though may result in lower dialysis adequacy and increased blood pressure. A reduction in CI and improved cardiac autonomic function indicate a positive effect on cardiovascular function.</p>

Anemia is common in patients with chronic kidney disease (CDK), contributes to reduced Quality of Life (QoL) and is associated with cardiovascular disease, morbidity and mortality. Epoetin raises hemoglobin (Hb) and increases QoL and physical exercise capacity. Because of concerns about safety and economics, current anemia treatment with epoetin aims to achieve subnormal Hb (110-120 g/l). Normalization of Hb may be of additional benefit regarding QoL and cardiovascular effects. The present study examines the effects of Hb normalization with epoetin on safety variables, QoL, graft function after kidney transplantation, dialysis adequacy, hemorheology, hemodynamics and cardiac autonomic function in CKD patients. In a randomized, multicenter study comprising 416 pre-dialysis and dialysis patients no difference was observed between patients treated to a normal or a subnormal Hb level on mortality, thrombovascular events, serious adverse events, vascular access thrombosis and residual renal function. QoL was enhanced in a subgroup of hemodialysis patients. Pretransplant epoetin treatment directed toward normal Hb levels did not result in worse graft function during 6 postoperative months. Dialysis adequacy was reduced in a subgroup of hemodialysis patients after normalization of Hb. The blood flow properties of pre-dialysis patients were altered. The hemorheological investigation demonstrated that Hb normalization caused a parallel increase in hematocrit and blood viscosity without other hemorheological changes. While the total peripheral resistance index increased, the cardiac index (CI) decreased. In a separate study cardiac autonomic function, measured by heart rate variability, was decreased in pre-dialysis patients. It was improved, but not fully normalized, by Hb normalization. On the basis of this study, Hb normalization with epoetin appears to be safe and increases QoL in hemodialysis patients though may result in lower dialysis adequacy and increased blood pressure. A reduction in CI and improved cardiac autonomic function indicate a positive effect on cardiovascular function.

Die Nierentransplantation ermöglicht Patienten die Wiederherstellung der Nierenfunktion. Aufgrund der begrenzten Verfügbarkeit an Organen nimmt dabei die Zahl der Transplantationen von einem lebenden Spender stetig zu. Zudem ermöglichen die präzisen und genauen Vorbereitungen und Abläufe bei Lebendnierenspenden eine bessere 5-Jahres-Überlebensrate als bei Kadaverspenden. Die genetische Verschiedenheit zwischen Spender und Empfänger bedingt jedoch eine lebenslange immunsuppressive Therapie, um Abstoßungsreaktionen und damit das Scheitern einer Organtransplantation zu verhindern. An den Universitätskliniken Leipzig und Halle/Saale besteht diese Therapie aus einer Dreifachkombination von Tacrolimus, Mycophenolat-Mofetil und Prednisolon, wobei mögliche Nebenwirkungen wie opportunistische Infektionen, kardiovaskuläre und metabolische Erkrankungen sowie Tumore in Kauf genommen werden. Zudem besteht für den immunsupprimmierten Organismus die ständige Gefahr einer Abstoßungsreaktion. Diese Aspekte führen bei den Empfängern zu einer massiven Einschränkung der Gesundheit und Lebensqualität. Inwieweit die ausgeprägte Immunsuppression notwendig ist, bleibt unklar und muss individuell festgelegt werden. Bisher existiert kein geeignetes Verfahren für ein Immunmonitoring, weshalb in vielen Fällen eine umfangreiche und überdosierte Immunsuppression in Kauf genommen wird. Im Rahmen dieser Arbeit wurde ein geeignetes Testverfahren, der Elispot-Assay, für die Expression der beiden proinflammatorischen Zytokine IFN-γ und IL-17 erstellt. Dafür wurden die PBMC der Spender und Empfänger aus Vollblut separiert, um sie anschließend sowohl separat als auch in einer Lymphozytenmischreaktion zu untersuchen. Die Darstellung von IL-17 konnte nur aufgrund einer zusätzlichen Stimulation mit OKT3 gelingen, während der IFN-γ-Elispot sowohl im Leerwert als auch unter Stimulation mit IL-2 zu ausreichenden Spotanzahlen führte. Die Spotanzahlen der Spender-PBMC wurden mit Hilfe von γ-Strahlung signifikant reduziert (IFN-γ: p=0,047 | IFN-γ + IL-2: p=0,007 | IL-17: p = 0,001), um in den Lymphozytenmischreaktionen die alleinige Zytokinausschüttung der Empfänger-PBMC messen zu können. Die Spender- PBMC fungierten dabei nur als Antigene. Insgesamt konnten zwischen 2009 und 2012 zwölf von siebzehn Patientenpaaren in die Studie eingeschlossen werden. Die Spotanzahlen der Paare wurden dabei sowohl im IFN-γ- als auch im IL-17-Elispot-Assay zu vier unterschiedlichen Zeitpunkten gemessen (vor Transplantation | 21±3 d postoperativ | 28±3 d postoperativ | 75±15 d postoperativ). In den meisten Fällen zeigte sich vor Transplantation eine erhöhte Spotanzahl im Vergleich zu den drei postoperativen Werten. Zudem stiegen die Spotanzahlen sowohl für IFN-γ als auch für IL-17 nach niedrigen Messergebnissen kurz nach der Transplantation im postoperativen Verlauf wieder an und erreichten in einigen Fällen die Spotanzahl der präoperativen Ausgangswerte. Ein signifikanter Unterschied konnte aufgrund der geringen Fallzahl nicht erreicht werden. Die kurzfristige Reduktion der Spotanzahlen postoperativ ist dabei aller Wahrscheinlichkeit nach auf die hohen Dosen an immunsuppressiven Medikamenten zurückzuführen. Insgesamt zeigten die Verläufe der IFN-γ- und der IL-17- Elispot-Assays ähnliche Verläufe. Daraus lässt sich schlussfolgern, dass der IL-17-Elispot- Assay in Bezug auf mögliche Abstoßungsreaktionen eine ähnliche Aussagekraft besitzen könnte wie der bereits vielfach untersuchte IFN-γ-Elispot-Assay. Weiterhin wurden die Messergebnisse mit der Serumkreatininmolarität verglichen. Diese zeigte präoperativ höhere Molaritäten als postoperativ, wobei die postoperativen Molaritäten im Verlauf, im Gegensatz zu den Elispot-Messungen, abnahmen, was das Einsetzen der Nierenfunktion widerspiegelt. Unter den zwölf Patientenpaaren gab es keine einzige nachgewiesene akute Abstoßungsreaktion, der Verlauf der Serumkreatininmolaritäten war bei allen zwölf Empfängern vergleichbar. Demzufolge konnten die Werte der Elispot-Assays nicht herangezogen werden, um an ihnen eine Abstoßungsreaktion der transplantierten Nieren erkennen zu können. Das präoperative Abschätzen einer möglichen Abstoßungsreaktion anhand der Elispot-Assays konnte aufgrund fehlender Abstoßungsreaktionen ebenfalls nicht untersucht werden. Zusätzlich wurde bei den Patienten eine HLA-Typisierung vorgenommen, wobei der Bereich von optimalen bis maximal ungünstigen Konstellationen reichten (HLA-Mismatch: 0-0-0 bis 2-2-2). Auch hier konnten die Ergebnisse nicht mit möglichen Abstoßungsreaktionen verglichen werden. In der vorliegenden Arbeit wurden zahlreiche Varianten untersucht, die das Abschätzen einer Immunreaktion nach Nierentransplantation (Immunmonitoring) ermöglichen könnten. Aufgrund fehlender Abstoßungsreaktionen bei den Empfängern konnte das Testverfahren nicht an den klinischen Verläufen validiert werden. Mit dem in dieser Arbeit entwickelten Messverfahren kann jedoch eine neue und größer angelegte Studie erfolgen, die in Zukunft ein Immunmonitoring bei Patienten nach Nierentransplantation ermöglicht. / Introduction Since the first kidney transplantation in the 1950ies, kidney transplantation is still being challenged by graft dysfunction and complete graft failure. Permanent immunsuppressive treatment is mandatory to avoid an unfavourable outcome. The treatment with Prednisolone, Tacrolimus and Mycophenolat-Mofetil may cause toxic side effects resulting in Diabetes mellitus, hypertension, infections and cancer. In the present study we tried to demonstrate that the amount of spots in the Enzyme linked immunospot assay (Elispot-Assay) of IFN-γ and IL-17 correlates with the probability of graft dysfuction and complete graft failure. We also compared the results to clinical parameters. Methods Between the years 2009 and 2012, twelve pairs of related living kidney transplantations were included in this study. From each pair blood samples were taken at four time points (before transplantation, and at 21±3, 28±3 and 75±15 days after kidney transplantation, respectively). After establishing the technique of IFN-γ- and IL-17-Elispot-Assays, we separated the periphale blood mononuclear cells (PBMC) and performed follow up examinations at the four time points mentioned above. The PBMC of each donor and each recipient were examined separatly, and in addition together in a lymphocyte mixed reaction. We stimulated the PBMC of the IFN-γ-Elispot with Interleukin-2 (IL-2) and the PBMC of the IL-17-Elispot with OKT3 to get significant characteristics. PBMC of the donors were irradiated with 30 Gy before mixing them with the PBMC of the recipients. We also took the HLA-matches and serum creatinine molarity to compare important clinical parameters with the results of the Elispot-Assays. Results Sufficient spots were measured using the unstimulated and stimulated IFN-γ-Elispot and the stimulated IL-17-Elispot. Radiation was significant at all three tests (IFN-γ: p=0,047 | IFN-γ + IL-2: p=0,007 | IL-17: p = 0,001). All twelve recipients showed a high number of spots before transplantation in both types of Elispot-Assays and most of them an increasing number of spots after a minimal turning point three weeks after transplantation. Due to the small number of cases, no significant results could be obtained at follow up. Non recipient developed a graft rejection as proven by biopsy or graft failure. The molarity of serum creatinine was permanently reduced whereas it was high before transplantation. Because of the abscence of any rejection episodes, HLA matches could not be compared. Discussion Due to the absence of rejection episodes or graft failure, no prediction for rejection by the IFN-γ- and IL-17-Elispot was possible. The low number of cases of living related kidney transplantation demonstrated the challange of the investigation of living related kidney transplantation. Although we could prove a significant effect of the irradiation of PBMC, there was no significant result in the follow up investigations. A higher number of cases are needed in future investigations. The established method of the IFN-γ- and IL-17-Elispot can be used in a future study with an extended number of cases and a longer follow up of time.

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