A.P. Navarro

Evaluation of the ischemic protection efficacy of a laparoscopic renal cooling device using renal transplantation viability assessment criteria in a porcine model.

PURPOSE We assessed the efficacy of a prototype laparoscopic topical cooling device. The aim of regional renal hypothermia in laparoscopic surgery is to limit ischemic injury and extend safe operative time. A reliable model for assessing renal ischemic injury exists in the field of nonheart beating donor renal transplantation. Hypothermic machine perfusion allows calculation of the pressure flow index and measurement of glutathione S-transferase in the perfusate. These parameters allow accurate assessment of the extent of renal damage. MATERIALS AND METHODS The device incorporates a 2-layer cooling bag and coolant circuit. The system achieves hypothermia by circulating coolant across the surface of the kidney. Using 10 pigs individual kidneys were subjected to periods of renal ischemia with or without device in situ cooling. Each kidney was then machine perfused and assessed using nonheart beating donor viability criteria. RESULTS The best performance of the device achieved a renal parenchymal temperature of 15C in 11.2 minutes (mean +/- SD 21.4 +/- 8.42). In the warm ischemia groups significant deterioration of pressure flow index compared to controls occurred by 60 minutes (p = 0.0001). In cooled kidneys at 60 minutes the mean pressure flow index was not significantly different from that in controls. Greater mean glutathione S-transferase measurements were associated with the warm ischemia groups. CONCLUSIONS Our study reinforces the efficacy of topical renal cooling in the laparoscopic setting. We report the use of assessment techniques capable of accurate quantitative measurement of renal injury in an animal model. Our cooling device is currently undergoing further development to enhance its efficiency.

Donation after Cardiac Death Kidneys with Low Severity Pre‐Arrest Acute Renal Failure

The widening gap between supply and demand for renal transplantation has prompted many centers to use donors after cardiac death. Some of these donors exhibit signs of acute renal failure (ARF) prior to cardiac arrest. Concern has been expressed about poor quality of graft function from such donors. In response to this perception, we reviewed 49 single renal transplant recipients from category III donors after cardiac death between 1998 and 2005, at out center. All kidneys but one had hypothermic machine perfusion and viability testing prior to transplantation. According to the RIFLE criteria, nine recipients had kidneys from donors with “low severity pre‐arrest ARF”. The remainder of the recipients were used as control group. There was no statistical significant difference in delayed graft function and rejection rates between these two groups. Recipients GFR at 12 months was 44.4 ± 17.1 and 45.2 ± 14.7 (mL/min/1.73m2) from donors with ARF and without ARF, respectively (p = 0.96). In conclusion, low severity ARF in kidneys from controlled after cardiac death donors can be a reversible condition after transplantation. Short‐term results are comparable to the kidneys from same category donors without renal failure, providing that some form of viability assessment is implemented prior to transplantation.

Laparoscopic versus open live donor nephrectomy: meta-analysis of randomized controlled trials

1 A Novel siRNA-Containing Solution Protecting Donor Organs in Heart Transplantation. Xiufen Zheng,1 Dameng Lian,1 Mu Li,1 Xusheng Zhang,1 Mahdieh Khoshniat,3 Hongtao Sun,1 Weihua Liu,1 Jifu Jiang,1 Dong Chen,1 Costin Vladau,1 Motohiko Suzuki,1 James C. Lacefi eld,3 Bertha Carcia,1,2 Anthony M. Jevnikar,1,2,3 Wei-Ping Min.1,2,3 1Department of Surgery, Pathology, and Microbiology & Immunology, University of Western Ontario, London, ON, Canada; 2Multi-Organ Transplant Program, London Health Sciences Centre, London, ON, Canada; 3Robarts Research Institute, London, ON, Canada. Background: Ischemia-reperfusion (I/R) injury occurs in organ transplantation. We hypothesize that siRNA can effectively suppress infl ammatory, apoptosis and complement genes which contribute to I/R injury. This study was designed to develop a novel siRNA-containing preservation solution, which offers an alternative for protection of donor organs. Methods: Multiple siRNAs that specifi cally target TNFα, Fas, and C3 genes were prepared using our newly-developed bio-synthesis method. Heart grafts from BALB/c mice were preserved in UW solution (control) or siRNA-containing UW solution (siRNA solution) at 4°C for 48 hrs, and subsequently transplanted into syngeneic BALB/c mice. Cardiac functions were quantitatively assessed by High Frequency Ultrasound. The I/R injury was assessed by immunohistochemistry. Results: After 48 hrs preservation in the siRNA solution, the expressions of TNFα, Fas, and C3 genes in the grafts were inhibited at mRNA and protein levels detected by qPCR and immunoblot. Using siRNA solution preserved organ as donor, the graft survival was signifi cantly prolonged in heart transplantation. While siRNA solution-treated heart grafts retained strong heartbeat up to the end point of observation (>70 days), the control grafts lost function within 7 days. In addition, an improved cardiac function was observed in the graft preserved in siRNA solution. Grafts treated by siRNA solution displayed normal function, whereas grafts preserved in control solution showed dysfunction as detected by ultrasound scanning. The protection of graft by siRNA solution is associated with prevention of I/R injury. siRNA solution-treated organs exhibited almost normal histological structures, less neutrophil and lymphocyte infi ltration as compared with control solution-treated organs. Furthermore, siRNA solution prevented apoptosis and necrosis of cardiac tissues. Conclusions: This is the fi rst demonstration of a novel siRNA solution that can prevent cardiac I/R injury, protect cardiac function, and prolong graft survival in heart transplantation, implying a signifi cant and potential application in clinic. Abstract# 2 The Long-Term Evaluation of HLA-Mismatched Combined Kidney and Bone Marrow Transplant Recipients after Complete Withdrawal of Immunosuppression. Tatsuo Kawai,1 David H. Sachs,2 Thomas Spitzer,3 Nina Tolkoff-Rubin,1 Susan Saidman,4 Winfred Williams,1 Bimalangshu Dey,3 Nelson Goes,1 Dicken Ko,1 Waichi Wong,1 Robert B. Colvin,4 Megan Sykes,2 A. Benedict Cosimi.1 1Transplantation Unit, Massachusetts General Hospital, Boston, MA; 2Trasnplant Biology Research Center, MGH, Charlestown, MA; 3Bone Marrow Transplant Unit, MGH, Boston, MA; 4Pathology, MGH, Boston, MA; 5this Study was Performed in Partnership with Immune Tolerance Network. BACKGROUND: Induction of allograft tolerance has been the ultimate goal in organ transplantation. METHODS: Five subjects with ESRD received combined kidney and bone marrow transplantation (CKBMT) from HLA mismatched donors. The preparative regimen consisted of cyclophosphamide, thymic irradiation, anti-CD2 mAb and a 9-14 months course of a calcineurin inhibitor. Two subjects also received two doses of antiCD20 mAb prior to transplant. RESULTS: All recipients developed transient mixed chimerism without GVHD.Complete withdrawal of immunosuppressive medication was achieved in 4/5 recipients with current immunosuppression-free survival exceeding 3.2, 2 The Long-Term Evaluation of HLA-Mismatched Combined Kidney and Bone Marrow Transplant Recipients after Complete Withdrawal of Immunosuppression. Tatsuo Kawai,1 David H. Sachs,2 Thomas Spitzer,3 Nina Tolkoff-Rubin,1 Susan Saidman,4 Winfred Williams,1 Bimalangshu Dey,3 Nelson Goes,1 Dicken Ko,1 Waichi Wong,1 Robert B. Colvin,4 Megan Sykes,2 A. Benedict Cosimi.1 1Transplantation Unit, Massachusetts General Hospital, Boston, MA; 2Trasnplant Biology Research Center, MGH, Charlestown, MA; 3Bone Marrow Transplant Unit, MGH, Boston, MA; 4Pathology, MGH, Boston, MA; 5this Study was Performed in Partnership with Immune Tolerance Network. BACKGROUND: Induction of allograft tolerance has been the ultimate goal in organ transplantation. METHODS: Five subjects with ESRD received combined kidney and bone marrow transplantation (CKBMT) from HLA mismatched donors. The preparative regimen consisted of cyclophosphamide, thymic irradiation, anti-CD2 mAb and a 9-14 months course of a calcineurin inhibitor. Two subjects also received two doses of antiCD20 mAb prior to transplant. RESULTS: All recipients developed transient mixed chimerism without GVHD.Complete withdrawal of immunosuppressive medication was achieved in 4/5 recipients with current immunosuppression-free survival exceeding 3.2, 2.5, 1.1 and 0.2 years. A recipient rejected his kidney allograft on day 10. (1) The most recent creatinine/GFR of these 4 subjects were 1.2/61, 1.6/75, 1.6/96 (by renogram), 1.8/71 ml/min, respectively. (2)Although Subjects 2 and 5 initially developed anti-donor antibodies (ADA), both recipients lost ADA by 10th and 3rd month, respectively. Subject 4 developed post-transplant de novo anti-class II ADA one month after discontinuation of his immunosuppression, but without any functional derangements. (3) Subjects 1, 2 and 4 showed donor specifi c hyporesponsiveness in CML and MLR after nine months. (4) Allograft biopsies taken at 3 years from Subjects 1 and 2 revealed normal glomerulus, no tubulitis and no vasculopathy by light and electron microscopic examination with no C4d staining. Biopsy taken at 1.5 years from Subject 4 showed positive C4d deposition with no changes diagnostic of rejection. A most recent biopsy taken from Subject 5 on day 243 showed no rejection. (5) In Subject 1, extensive warts due to human polyoma virus existed before transplantation, but it disappeared after discontinuation of immunosuppression. All four patients show no hypertension, no DM, no abnormal lipid profi les. CONCLUSIONS: Renal allograft function of CKBMT recipients has been stable up to 3 years after complete withdrawal of their immunosuppression. Abstract# 3 A Novel Human Anti-CD40 Monoclonal Antibody, 4D11, for Kidney Transplantation in Cynomolgus Monkeys, Effect of Induction and Maintenance Therapy. Takeshi Aoyagi,1 Tomomi Suzuki,1 Atsushi Sugitani,2 Atsushi Imai,1 Motohiro Uno,1 Ryoichi Goto,1 Kenichiro Yamashita,1 Toru Miura,3 Nobuaki Takahashi,3 Tomoo Itoh,4 Hiroyuki Furukawa,1 Satoru Todo.1 11st Dept of Surgery, Hokkaido Univ., Sapporo, Hokkaido, Japan; 21st Dept of Surgery, Kyusyu Univ., Hakata, Fukuoka, Japan; 3Kirin Brewery Co., Ltd, Takasaki, Gunma, Japan; 4Dept of Surgical Pathology, Hokkaido Univ. Hospital, Sapporo, Hokkaido, Japan. Background/Aim: We have previously demonstrated that a novel human anti-CD40 monoclonal antibody (mAb), 4D11, markedly prolongs renal allograft survival in cynomolgus monkeys. In this study, we evaluated the effect of induction and maintenance therapy using 4D11. Methods: Twenty-four monkey renal transplants were divided into three groups: no treatment (Control), induction (Group A) and maintenance (Group B) treatments. Groups A and B were further subdivided according to the dose of 4D11 (n=3 each). The 4D11 was given intravenously on days 0, 4, 7, 11 and 14 for the induction therapy. In the group B, a half of the initial dose was given weekly thereafter until day 180. Graft survival, graft histology, anti-4D11 and anti-donor antibodies were examined. Results: Treatment Group 4D11 (mg/kg) Graft survival (days) Mean (days) Cause of death No treatment Control 5, 6, 7 6.0 AR, AR, AR Induction A-I 5 >96, 79, 75 83.3 Alive, AR, AR A-II 10 >200, 107, 92 133.0 Alive, CAN, AR A-III 20 169, 105, 90 121.3 AR, AR, AR Maintenance B-I 1 80, 79, 10 56.3 AR, AR, AR B-II 5 >202, 98, 44 114.3 Alive, AR, AR B-III 10 374, 253, 27 218.3 AR, AR, UTI B-IV 20 216, 169, 153 179.3 CAN, AR, CAN AR: Acute rejection, CAN: Chronic allograft nephropathy, UTI: Urinary tract infection No adverse events were observed. Graft survival was markedly prolonged by the induction treatment, which was further extended by the maintenance treatment. While graft biopsies revealed grade II or III rejection in the control, majority of those from groups A and B at 1 month post-transplantation showed only a borderline change. Anti 4D11-Ab was detected only in group A-III after drug cessation. In addition, animals receiving induction treatment and those given maintenance treatment with high doses of 4D11 did not develop anti-donor Ab unless the treatment was discontinued. Conclusion: 4D11 exhibited potent immunosuppressive effect on renal allograft survival both by induction and maintenance treatments. Besides, anti-drug and anti-donor antibody productions were effectively suppressed by 4D11. Abstract# 4 Pre-Transplant (Tx) Donor-Specifi c B Cells in Highly Sensitized Patients (HS), Detected by Intracellular Cytokine Flow Cytometry (CFC) Predict Antibody Mediated Rejection (AMR). Mieko Toyoda,1 Andy Pao,1 Ashley Vo,1 Marina Lukovsky,1 Raju Radha,1 Tetsu Sado,1 Lara Baden,1 Anna Petrosyan,1 Stanley C. Jordan.1 1Transplant Immunology Laboratory and Comprehensive Transplant Center, Cedars-Sinai Medical Center, Los Angeles, CA. Background: Intravenous immunoglobulin treatment (IVIG-Rx) reduces panel reactive antibodies and crossmatch (CMX) positivity, facilitating CMX(-) kidney Tx in HS. However, high rates of allograft rejec