Martin Wissing

Critical role of interleukin 4 in the induction of neonatal transplantation tolerance

Neonatal injection of semiallogeneic cells is known to promote differentiation of donor-specific CD4+ T cells into TH2-like cells in the peripheral lymphoid organs. We reasoned that the propensity of neonatal T cells to synthesize high levels of IL-4 might be involved in this polarization of the alloreactive response and thereby in the development of neonatal transplantation tolerance. First, analysis of cytokine gene expression in lymph nodes after neonatal injection of 107 (A/J×BALB/c)Fl cells in BALB/c mice indicated that IL-4 but not IL-2 is rapidly produced by CD4+ cells after allogeneic challenge in vivo. To determine whether the early production of IL-4 was involved in the establishment of allotolerance, BALB/c mice neo-natally injected with (A/JxBALB/c)Fl spleen cells received on days 1 and 3 after birth 1 mg of anti-IL-4 mAb (11B11) or the same amount of control mAb. When grafted with A/J skin at 4 weeks, 88% of mice treated with control mAb retained their graft for more than 50 days, whereas rejection occurred within 30 days in 93% of mice treated with anti-IL-4 mAb. Analysis of T cell functions after in vitro restimulation with A/J spleen cells indicated that early IL-4 neutralization did not prevent donor-specific CTL unresponsiveness but allowed the emergence of alloreactive T cells secreting increased levels of IL-2 and IFN–y. We conclude that early production of IL-4 is critical for the establishment of neonatal transplantation tolerance in this strain combination, which has disparities across the entire H-2 region.

Fatal primary infection due to human herpesvirus 6 variant A in a renal transplant recipient.

We describe a fatal primary human herpesvirus 6 (HHV-6) variant A infection in a kidney transplanted adult woman. On day 20 post transplantation (TX), after rejection therapy, the patient presented an acute hemophagocytic syndrome with hepatitis and central nervous system involvement. HHV-6 IgG and IgM antibodies seroconversion was demonstrated. HHV-6 variant A was the sole pathogen detected by nested PCR and/or culture in blood, bone marrow aspiration, liver biopsy, cerebrospinal fluid and bronchoalveolar lavage. The graft was HHV-6 seropositive and the patient was not transfused before day 28 post TX, suggesting that the virus was transmitted by the graft. Despite immunoglobulins, ganciclovir and foscarnet therapy, the HHV-6 infection progressed and led to severe aplasia. The patient developed Aspergillus fumigatus pneumonia and died from fulminant candidemia. This case demonstrated for the first time that HHV-6 variant A primary infection can cause life-threatening disseminated infection in immunosuppressed patients.

Male recipients of kidneys from female donors are at increased risk of graft loss from both rejection and technical failure

The aim of the present retrospective study was to uncover the factor(s) responsible for the poor outcome of cadaver kidney grafts from female donors in male recipients.The 741 transplantations performed at our center from August 1983 to September 1997 were distributed into four groups according to recipient and donor gender: female donor to female recipient (F to F: n=117), male donor to female recipient (M to F: n=172), female donor to male recipient (F to M: n=170), and male donor to male recipient (M to M: n=282). All the patients received immunosuppressive therapy based on corticosteroids and cyclosporine, associated or not with either azathioprine or prophylactic anti‐lymphocyte globulin.Overall graft survival was lower in the F to M group than in the three other groups (p=0.009). Failures due to rejection were more frequent during the 1st post‐transplant trimester in female than in male donor grafts, irrespective of recipient gender (p=0.025). All failures due to technical problems occurred during the first 3 months post‐transplantation: they were more frequent in the F to M group than in the three other groups (p=0.040); this could be related to the older age of the donors in the former group. After the first post‐transplant year, failures due to causes other than rejection remained low in the F to F group but increased steadily in the three other groups (p=0.007). Specific survival rates were not correlated with the time‐evolution of mean serum creatinine values, daily doses and trough levels of cyclosporine in the four groups of grafts. In conclusion, the poor outcome of F to M grafts results from combined immunologic and technical factors exerting their effects early in the course of transplantation.

Modulation of the release of cytokines and reduction of the shock syndrome induced by anti-CD3 monoclonal antibody in mice by interleukin-10.

Since IL-10 was recently shown to inhibit several T cell functions in vitro, we investigated the effects of IL-10 on the cytokine release syndrome induced in mice by the 145–2C11 anti-CD3 mAb. As OKT3 in man, this mAb induces a massive polyclonal T cell activation before to induce immunosuppression. First, we found that administration of 1000 U of recombinant mouse IL-10 (mIL-10) 30 min before injection of 10 μg of the 145–2C11 antimouse CD3 mAb markedly reduced the systemic release of IFN-γ and TNF. In contrast, IL-10 pretreatment did not significantly modify the release of IL-6. To determine the effect of IL-10 pretreatment on the endogenous secretion of IL-10 induced by the 145–2C11 mAb, mice were injected with human IL-10 (hIL-10) which does not cross-react in the ELISA for mIL-10 determination. While hIL-10 was as efficient as mIL-10 in reducing TNF and IFN-γ release, it did not modify peak serum levels of IL-10. The modulation of cytokine production by mIL-10 was associated with a significant reduction of the toxicity of the 145–2C11 mAb, as assessed by the attenuation of hypothermia and by the reduced lethality in D-galac-tosamine-sensitized mice. We conclude that IL-10 differentially regulates the in vivo production of cytokines and decreases the systemic toxicity induced by the 145–2C11 mAb. These observations suggest potential therapeutic applications of IL-10 in organ transplantation, especially in association with anti-CD3 mAb.

Efficacy and cardiovascular safety of daclizumab, mycophenolate mofetil, tacrolimus, and early steroid withdrawal in renal transplant recipients: a multicenter, prospective, pilot trial

Abstract:  This single‐arm, open‐label, pilot study was designed to assess the efficacy and cardiovascular safety profile of daclizumab, a humanized monoclonal interleukin (IL)‐2Rα antibody, in combination with mycophenolate mofetil (MMF), tacrolimus, and early corticosteroid withdrawal in renal transplant recipients. Seventy‐nine renal allograft recipients were treated with daclizumab (1 mg/kg; five doses starting on the day before transplant and then every two weeks), MMF (1 g b.i.d.), tacrolimus (0.2 mg/kg/d), and low‐dose prednisolone, which was withdrawn at day 150 after transplant. The rate of acute rejection was determined at 12 months. Lipid profile, oral glucose tolerance, and adverse events were monitored. Of the 76 patients eligible for analysis, eight (10.5%) developed biopsy‐proven acute rejection (BPAR). Ten (13.2%) experienced clinical and/or BPAR. Corticosteroids were withdrawn completely in 91% of patients at 12 months. Graft and patient survival were 97.5% and 98.7% respectively. Mean total cholesterol and triglycerides were significantly lower at 12 months post‐transplant than at baseline (201 ± 47.5 vs. 190.8 ± 43.6 mg/dL, p = 0.005 and 196.2 ± 133.2 vs. 144.5 ± 76.8 mg/dL, p < 0.001, respectively). Mean hemoglobin A1c levels did not differ between baseline (5.54%) and 12 months (5.48%). New‐onset post‐transplant diabetes mellitus occurred in 6.6% of the non‐diabetic transplanted patients. The proportion of patients with abnormal oral glucose tolerance test (OGTT) was 47% at 3 months and 39% at 12 months (p = NS). Daclizumab induction in combination with MMF, tacrolimus, and low‐dose (followed by withdrawal) prednisolone appears to be effective and safe in patients receiving renal allografts. The regimen appears to be associated with a favorable cardiovascular profile.

Ticlopidine and clopidogrel, sometimes combined with aspirin, only minimally increase the surgical risk in renal transplantation: a case–control study

BACKGROUND Patients undergoing kidney transplantation are sometimes being treated with antiplatelet agents such as ticlopidine or clopidogrel. Some teams refuse to wait-list these patients for fear of bleeding during transplant surgery. METHODS We retrospectively reviewed the records of 702 adult patients with a kidney transplant alone between 2000 and 2010. Nineteen (2.7%) patients were taking clopidogrel or ticlopidine when called in for transplantation. Furthermore, 10 of these 19 patients were also taking low-dose aspirin (ASA). We compared the risk of bleeding peri- and postoperatively, and the occurrence of cardiovascular complications within 30 days after renal transplantation between 19 cases and 39 controls randomly selected within the cohort. RESULTS Platelets were administered to 7 cases (37%) versus 0 controls (P<0.001). A single case (5.3%) presented with significant bleeding during surgery following an implantation biopsy, and required 4 red bood cell (RBC) units. During the first day, 3 of the 19 cases (16%) and 1 of the 39 controls required RBC (P=0.1). No reoperation was performed for bleeding. After the transplant, clopidogrel or ticlopidine was resumed in only two patients. The platelet count and haemoglobin were similar between cases and controls at Day 30. No cardiovascular event occurred in cases or controls during the first month post-transplantation. At 5 years, graft and patient survival was similar in cases and controls. CONCLUSIONS Clopidogrel and ticlopidine, sometimes in combination with ASA, are associated with a low risk of bleeding during renal transplantation and does not seem to be a contraindication for renal transplant surgery.

Belgian experience of DCD kidney transplantation

O-098 – Table 1. LTx in patients with incidental PPHT Indication labMELD mPAP at induction (mmHg) hospital stay (days) Medical treatment post-LTx Outcome Follow-up (months) m/54yr Post-ethyl cirrhosis 32 44 38 Spontaneous resolution Alive and well 15 f/37yr Post-ethyl cirrhosis 26 26 40.6 42 Epoprostenol IV during 13 months Alive and well 13 f/62yr HBV 31 46.7 92 Sildenafil PO during 9 months Alive and well 12 m/61yr Post-ethyl cirrhosis and HCC 12 36.7 6 Extra-corporeal membrane oxygenation †6d post-LTx – f/67yr Sarcoidosis 36 34.3 – Aborted LTx †1d post-LTx – f/54yr Post-ethyl cirrhosis 24 35.3 22 Spontaneous resolution Alive and well 16 f/53yr Postethyl + HBV cirrhosis 19 37.3 19 Spontaneous resolution Alive and well 55 f/53yr PBC 17 51.7 58 Sildenafil PO Alive and well 6 m/42yr HCV 17 53.3 – Listed for combined heart/lung/LTx Alive and well 120 Eleven (9.9%) patients did not even achieve 65% of the predicted target heart rate, and notably all of them were on β-blockers. Thirty (73.1%) of 41 patients who achieved the target heart rate had MELD score ≤15 compared with 11 (26.9%) patients with MELD score > 15 (p < 0.05). Conclusions: Chronotropic incompetence on DSE is frequent in patients with ESLD. In absence of any cardiac symptoms or/and ECG findings during DSE, a lower cut-off for target heart rate may be acceptable when patients are on βblockers or/and MELD score >25 to avoid unnecessary further investigations. Large prospective studies are needed to support these findings. O-098 INCIDENTAL PORTOPULMONARY HYPERTENSION DISCOVERED AT THE START OF LIVER TRANSPLANTATION, “TO GO AHEAD OR TO LET GO...” Filip Gryspeerdt, Marion Dupont, Wim Laleman, Raymond Aerts, Dieter Mesotten, Geert Meyfroidt, Marleen Verhaegen, Arne Neyrinck, Frederik Nevens, Jacques Pirenne, Diethard Monbaliu. Leuven liver Transplant Team, University Hospitals Leuven, Leuven, Belgium Background: Portopulmonary hypertension (PPHT) is the association of pulmonary hypertension (mean pulmonary artery pressure [mPAP] >25 mmHg) and portal hypertension with or without chronic liver disease. Moderate PPHT (mPAP >35 mmHg) is associated with higher morbidity/mortality and severe PPHT (mPAP> 45mmHg) is generally considered a contra-indication for Liver Transplantation (LTx). Moderate to severe PPHT may develop during the waiting time of LTx period. A retrospective analysis was done to review the shortterm outcome of LTx in patients with incidental PPHT (e.g. diagnosed at the start of LTx and unkown at time of listing). Methods: All medical records of patients with incidental PPHT were reviewed. Lab-MELD at time of LTx, mPAP immediately pre-LTx, post-LTx hospital stay, type/length of post-LTx medical treatment for PPHT and patient survival were analyzed (see Table 1). Results: Between 2000-2011, 9/653 patients were diagnosed with moderate to severe PPHT at time of LTx induction. LTx was pursued in 7 patients. Of those, 6 had uneventful post-LTx recovery with spontaneous or medically assisted (vasodilators) resolution of PPHT; and 1 succumbed to complications of extra-corporeal membrane oxygenation. LTx was started but aborted in 1 due to hemodynamic unstability. LTx was not started in 1 who later received combined heart/lung/LTx. Conclusion: The incidental discovery of a previously unknown moderate to severe PPHT at the start of LTX is a possibility that LTx teams should be aware of. PPHT has usually been seen as a contra-indication for LTx. However, favorable outcome in 6/7 recipients suggests that LTx should not necessarily be aborted in case of incidental PPHT. 28 Oral Sessions Oral Session 13: Liver / intestine miscellaneous O-099 LIVER INMUNOPROTECTIVE EFFECT ON THE KIDNEY ALLOGRAFT IN SIMULTANEOUS LIVER AND KIDNEY TRANSPLANTATION Nuria N. Esforzado 1, Ana Yurena A.Y. Sánchez 1, José Vicente J.V. Torregosa 1, Nuria N. Serra 1, Rafael R. Pascualin 1, Jaume J. Martorell 2 , Federico F. Oppenheimer 1 , Josep Maria J.M. Campistol 1 . 1Renal Transplant Unit, 2Inmunology Unit, Hospital Clinic, Barcelona, Spain Background: Simultaneous liver-kidney transplantation (SLK) has less incidence of renal graft rejection and inmunological graft lost against the receptors of an isolated renal transplantation (RT). In addition, a low rejection incidence and a good renal graft evolution have ben reported in cross-match (CM) positive (+) SLK patients. The low prevalence of immunological complications in high-risk immune (“HRI”) SLK patients, suggests a liver’s inmunoprotective effect on the kidney graft. Material and methods: We present our experience in “HRI” SLK patients, defined as CM by cytotoxicity (CDC) post DTT + and/or “HRI” + pre transplantion (Tx). From May 1993 until December 2010, 58 SLK Tx were made (27 retransplanted patients), and eight patients had CM + pre Tx and other four patients had negative CM but positive “HRI”. Results: Of 12 “HRI” patients, 3 (25%) patients had graft dysfunction related to humoral acute rejection (HAR) during the first month after SLK Tx. Only one of these patients (33%) received Apheresis and Rituximab treatment with a good response. In the other two patients, HAR was resolved without specific treatment. None of 12 patients after 45±40 months follow-up, loss graft related to inmunological etiology. Six of 8 CM + pre Tx patients became negative post Tx. Conclusion: High-risk inmune SLK patients have a low prevalence of immunological complications which suggests an inmunoprotector role of the liver on the kidney allograft in these patients. O-100 EVOLUTION OF KIDNEY FUNCTION AFTER LIVER TRANSPLANTATION FOR ADULT POLYCYSTIC LIVER DISEASE AND INDICATIONS FOR COMBINED LIVER AND KIDNEY TRANSPLANTATION Tom Darius 1, Alexander Patris 1, Ziad Hassoun 1, Diethard Monbaliu 2, Tania Roskams 2, Olga Ciccarelli 1, Yves Pirson 1, Yves Vanrenterghem 2 , Frederik Nevens 2, Jacques Pirenne 2, Jan Lerut 1 . 1Abdominal Transplant Unit, University Hospitals Saint Luc, Brussels, Belgium; 2Liver Transplant programme, University Hospitals Gasthuisberg, Leuven, Belgium Background: Adult polycystic liver disease (PLD) is frequently associated with autosomal dominant polycystic kidney disease (ADPKD). Established indication for combined liver and kidney transplantation (CLKTx) is end stage renal failure. If renal insufficiency is less advanced, indications for combined kidney transplantation (KTx) are controversial. We reviewed our experience with isolated liver transplantation (LTx) and CLKTx in patients with PLD. Methods: Between 1995-2008, 56 patients originating from 2 collaborating centers underwent LTx for PLD. 7 patients with isolated PLD received LTx alone. Of 49 patients with combined PLD and ADPKD, 31 underwent isolated LTx and 18 CLKTx. Among the 18 CLKTx recipients, 11 were dialysisdependent pre-transplant whereas 7 had a creatinine clearance (CrCl) between 15 and 38 mL/min. Results: Median follow up is 34 months (range, 26-167). 1 and 5-year patient and liver graft survival were 96% and 94%, and 96% and 90%, respectively. The 1 and 5-year kidney graft survival (death censored) is 100%. Of the 31 patients who underwent isolated LTx for combined PLD and ADPKD, 29% (n=9) developed terminal renal failure post-LTx. Their mean pre-LTx CrCl was 76 mL/min (range, 48-110). The mean pre-LTx CrCl in the 71% patients who display stable kidney function post-LTx was 78 mL/min (range, 47-153). Pre-LTx CrCl was not a significant factor for the development of renal failure after isolated LTx for combined PLD and ADPKD (p=0,835). Conclusion: This series demonstrates that short& long-term survival after LTx and CLKTx for PLD is excellent. In patients with clearly-proven & evolving renal impairment pre-transplant, CLKTx is the preferred option, anticipating the need for later KTx. In patients with preserved/mildly disturbed renal function, nephron sparing strategies are essential since evolution towards renal failure is seen in 29%, without clear prognosis factors. O-101 LIVER TRANSPLANTATION (LTx) FOR TRANSTHYRETIN SYSTEMIC AMYLOIDOSIS DISORDERS: ANALYSIS FROM THE FAMILIAL AMYLOIDOTIC POLYNEUROPATHY WORLD TRANSPLANT REGISTER (FAPWTR) Bo-Göran Ericzon. Transplantation Surgery, Karolinska Institutet, Stockholm, Sweden Background: Transthyretin (TTR) systemic amyloidosis disorders are treatable with Ltx. The FAPWTR was established in 1993 to assemble data on such patients. Methods/Results: By December 2009, 1798 patients/1953 liver transplantations were reported to the FAPWTR from 72 centers in 19 different countries. Most transplantations were done in Portugal (n=866), France (n=216), Sweden (n=130) and Brazil (n=91). More than 45 different TTR-variants have been reported, the commonest being Val30Met (85%) followed by Ser77Tyr, Thr60Ala and Tyr114Cys. Gastrointestinal, cardiovascular and extraneurological manifestations appear more often in non-Val30Met than in Val30Met patients. 15% of the non-Val30Met patients underwent liver and heart transplantation compared to 0.1% of the Val30Met patients. Different countries show varying age at onset in Val30Met patients, with Brazil having the youngest patients and Sweden the oldest (32 years and 45 years, respectively). After Ltx, 80-90% of the ValMet30 patients reported stable or improved clinical symptoms compared to 60-65% in non-Val30Met patients. The overall 5-, 10and 15-year patient survival is 79%, 70% and 64%, respectively. Most common cause of death is cardiac. Val30Met patients with a disease duration >7 years disclose inferior 5-year survival than patients with a duration ≤7 years (58.2% and 84.7%, respectively p<0.001). Results improve when analyzing patients transplanted in the last 5 years, but the 5-year survival still remains significantly better in patients with less than 7 years disease duration (72.1% and 88.7%, respectively p<0.05). Conclusion: LTx is lifesaving in patients with TTR amyloidosis. Val30Met and non-Val30Met TTR mutations differ clinically. Cardiac related post transplant death i

Susceptibility and resistance to autoimmunity following neonatal injection of semi-allogeneic spleen cells in rats

A model of neonatal allotolerance was developed in rats. Brown-Norway (BN) neonates injected with semi-allogeneic (BN x Lewis) F1 hybrid spleen cells express a long-lasting chimerism and exhibit polyclonal B cell activation demonstrated by hyperimmunoglobulinemia affecting mainly IgE and IgG1, anti-laminin and anti-DNA autoantibodies as well as glomerulonephritis and anti-hapten antibodies. These abnormalities are autoregulated although the chimerism persists. In contrast, Lewis (LEW) neonates injected with semi-allogeneic (BN x LEW) F1 hybrid spleen cells exhibit a very short-lasting chimerism and transient activation of B cells, as reflected by increased allo-class II antigen expression, but do not develop an autoimmune disease. The autoimmune syndrome observed in BN rats is similar to that reported in mice during host-versus-graft reaction. Similarities between the drug-induced models of autoimmunity and allogeneic reactions in BN rats are also striking. The susceptibility of BN rats and the resistance of LEW rats to these autoimmune diseases might respectively reflect the involvement of TH2-like or of TH1-like subsets.

Modulation of murine host-versus-graft disease by anti-CD3 monoclonal antibody

BALB/c mice made tolerant to A/J alloantigens by neonatal injection of (A/J × BALB/c)FI spleen cells develop a host‐versus‐graft (HVG) disease due to the activation of donor B cells by a subset of host alloreactive helper T cells. We have investigated the effects of a single neonatal injection of the 145‐2C11 anti‐mouse CD3 monoclonal antibody (MoAb) on the establishment of allotolerancc and on the development of the immunopathological features of HVG disease. First, this treatment did not modify the specific anti‐donor cytotoxic T lymphocyte (CTL) unresponsiveness or the persistence of circulating immunoglobulins bearing donor allotype. Second, the hyper IgE, the hyper IgG 1 and the increased expression of Ia antigens on B cells found in untreated HVG mice were not observed after injection of the 145‐2C11 MoAb. Likewise, treated mice displayed lower levels of anti‐DNA IgG antibodies and less glomerular immune deposits as compared with untreated HVG mice. We conclude that the administration of the anti‐CD3 MoAb did not interfere with the induction of allotolerance but exerts a pronounced inhibitory effect on the associated immunopathological syndrome.