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Background: Lecithin cholesterol acyl transferase (LCAT) deficiency is a rare autosomal recessive disorder of lipoprotein metabolism that results in end-stage renal disease (ESRD) necessitating transplantation. As LCAT is produced in the liver, combined kidney and liver transplantation was proposed to cure the clinical syndrome of LCAT deficiency., Methods: A 29-year-old male with ESRD secondary to LCAT deficiency underwent a sequential kidney-liver transplantation from the same living donor (LD). One year following the kidney transplant, auxiliary partial orthotopic liver transplant (APOLT) of a left lateral segment from the same donor was performed., Results: At 5 years follow-up, there have been no major complications, readmissions, or rejection episodes. Serum lipid abnormalities recurred within the first year, but liver and kidney allograft function remains intact., Conclusion: Few cases of sequential transplantation from the same LD have been performed in adults. This is the first APOLT and multi-organ transplant performed for LCAT deficiency. Sequential organ transplant from the same LD for ESRD secondary to a metabolic disorder of the liver is feasible in adults and should be further investigated., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Timing of bilateral nephrectomy (BN) is controversial in patients with refractory symptoms of autosomal dominant polycystic kidney disease (APKD) in need of a renal transplant., Methods: Adults who underwent live donor renal transplant (LRT) + simultaneous BN (SBN) from August 2003 to 2013 at a single transplant center (n = 66) were retrospectively compared to a matched group of APKD patients who underwent LRT alone (n = 52). All patients received general health and polycystic kidney symptom surveys., Results: Simultaneous BN increased operative duration, estimated blood loss, transfusions, intravenous fluid, and hospital length of stay. Most common indications for BN were pain, loss of abdominal domain, and early satiety. There were more intraoperative complications for LRT + SBN (6 vs 0, P = 0.03; 2 vascular, 2 splenic, and 1 liver injury; 1 reexploration to adjust graft positioning). There were no differences in Clavien-Dindo grade I or II (39% vs 25%, P = 0.12) or grade III or IV (7.5% vs 5.7%, P = 1.0) complications during the hospital course. There were no surgery-related mortalities. There were no differences in readmission rates (68% vs 48%, P = 0.19) or readmissions requiring procedures (25% vs. 20%, P = 0.51) over 12 months. One hundred percent of LRT + SBN allografts functioned at longer than 1 year for those available for follow-up. Survey response rate was 40% for LRT-alone and 56% for LRT + SBN. One hundred percent of LRT + SBN survey responders were satisfied with their choice of having BN done simultaneously., Conclusions: Excellent outcomes for graft survival, satisfaction, and morbidity suggest that the combined operative approach be preferred for patients with symptomatic APKD to avoid multiple procedures, dialysis, and costs of staged operations.

The objective of this review is to summarize the collective knowledge regarding the risks and complications in vascularized composite tissue allotransplantation (VCA), focusing on upper extremity and facial transplantation. The field of VCA has entered its second decade with an increasing experience in both the impressive good outcomes, as well as defining challenges, risks, and experienced poor results. The limited and selective publishing of negative outcomes in this relatively new field makes it difficult to conclusively evaluate outcomes of graft and patient survival and morbidities. Therefore, published data, conference proceedings, and communications were summarized in an attempt to provide a current outline of complications. These data on the medical complications of VCA should allow for precautions to avoid poor outcomes, data to better provide informed consent to potential recipients, and result in improvements in graft and patient outcomes as VCA finds a place as a therapeutic option for selected patients., Competing Interests: Conflict of InterestArthur Nam, Rolf Barth, Jhade Woodall, Mehmet Uluer, Philip Brazio, and Stephen Bartlet declare that they have no conflict of interest., (© Springer International Publishing AG 2016.)

The importance of further elucidating the properties surrounding microchimerism in various experi- mental models and clinical transplantation are limited by current techniques and the sensitivity of available platforms. Development of reliable methods and use routine use of microchimerism detection in clinical practice could guide clinical decision making regarding rejection, stable function, and tolerance.

We describe the incidence of early graft failure (EGF, defined as loss of function from any cause within 3 days after transplant) in a large cohort of GalTKO pig organs transplanted into baboons in three centers, and the effect of additional expression of a human complement pathway-regulatory protein, CD46 or CD55 (GalTKO.hCPRP). Baboon recipients of life-supporting GalTKO kidney (n = 7) or heterotopic heart (n = 14) grafts received either no immunosuppression (n = 4), or one of several partial or full immunosuppressive regimens (n = 17). Fourteen additional baboons received a GalTKO.hCPRP kidney (n = 5) or heart (n = 9) and similar treatment regimens. Immunologic, pathologic, and coagulation parameters were measured at frequent intervals. EGF of GalTKO organs occurred in 9/21 baboons (43%). hCPRP expression reduced the GalTKO EGF incidence to 7% (1/14; P < 0.01 vs. GalTKO alone). At 30 mins, complement deposits were more intense in organs in which EGF developed (P < 0.005). The intensity of peri-transplant platelet activation (as β-thromboglobulin release) correlated with EGF, as did the cumulative coagulation score (P < 0.01). We conclude that (i) the transgenic expression of a hCPRP on the vascular endothelium of a GalTKO pig reduces the incidence of EGF and reduces complement deposition, (ii) complement deposition and platelet activation correlate with early GalTKO organ failure, and (iii) the expression of a hCPRP reduces EGF but does not prevent systemic coagulation activation. Additional strategies will be required to control coagulation activation., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Introduction: We have aggressively used continuous veno-venous hemofiltration (CVVH) on high model for end-stage liver disease (MELD) score liver transplant patients with acute kidney injury and hypothesized that the addition of intraoperative CVVH therapy would improve overall outcomes., Methods: We performed a retrospective review of all adult, single organ, liver transplant recipients requiring preoperative renal replacement therapy between January 1, 2011 and June 1, 2013. Intraoperative and perioperative records and laboratory values were collected and used to create a database of these patients. Patients were grouped according to whether or not they underwent CVVH at the time of liver transplantation., Results: Twenty-one patients with new-onset renal failure requiring preoperative renal replacement therapy received a liver transplant alone. Fourteen received intraoperative CVVH and 7 patients did not. The average MELD score was similar between groups (34 for intraoperative CVVH vs 35; P = .8). Preoperative sodium and potassium were higher for the group receiving intraoperative CVVH, but still fell within normal ranges. Preoperative lactate levels were higher in the group that received intraoperative CVVH (4.7 vs 2.0 mmol/L; P = .01). Intraoperative CVVH did not decrease intraoperative transfusion requirements or intensive care unit (ICU) and hospital lengths of stay. Differences in reoperative rates did not reach statistical significance. All patients were weaned off renal replacement therapy. One-year patient survival rate was 86% for intraoperative CVVH versus 71% without., Conclusion: The judicious use of intraoperative CVVH therapy may permit patients with increasing severity of illness to achieve outcomes comparable with less ill patients., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: There is a paucity of data regarding long-term renal graft survival in hepatitis C virus positive (HCV+) patients. We analyzed our institution's experience with HCV+ renal transplantation and factors contributing to subsequent renal graft failure., Methods: We analyzed 1,679 adult, deceased donor, single-organ renal transplants occurring between 2000 and 2012. Recipient and donor demographics, HCV serostatus, and graft outcome and function were evaluated., Results: Of 1,679 patients, 195 HCV+ recipients (R+) received renal transplants from HCV+ donors (D+), in contrast to 1,418 HCV negative (HCV-) recipients (R-) who received grafts from HCV- donors (D-), and 66 R+ patients who received D- kidneys. Death-censored graft survival in the R+/D+ population was better than graft survival for R+/D- patients, despite R+/D+ patients having higher rates of hypertension and African Americans. Waitlist times for patients accepting HCV+ grafts was 318 days (for R+/D+ patients) versus 613 days (R-/D-) or 570 days (R+/D-). On multivariate analysis, waitlist times were independently predictive of graft failure., Conclusion: R+/D+ patients spent less time on the transplant waitlist, which contributed to improved death censored graft survival when compared with R+/D- patients.

Since the first facial transplantation in 2005, 28 have been done worldwide with encouraging immunological, functional, psychological, and aesthetic outcomes. Unlike solid organ transplantation, which is potentially life-saving, facial transplantation is life-changing. This difference has generated ethical concerns about the exposure of otherwise young and healthy individuals to the sequelae of lifelong, high-dose, multidrug immunosuppression. Nevertheless, advances in immunomodulatory and immunosuppressive protocols, microsurgical techniques, and computer-aided surgical planning have enabled broader clinical application of this procedure to patients. Although episodes of acute skin rejection continue to pose a serious threat to face transplant recipients, all cases have been controlled with conventional immunosuppressive regimens, and no cases of chronic rejection have been reported., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Human diffuse large B-cell lymphomas (DLBCLs) often aberrantly express oncogenes that generally contain complex secondary structures in their 5' untranslated region (UTR). Oncogenes with complex 5'UTRs require enhanced eIF4A RNA helicase activity for translation. PDCD4 inhibits eIF4A, and PDCD4 knockout mice have a high penetrance for B-cell lymphomas. Here, we show that on B-cell receptor (BCR)-mediated p70s6K activation, PDCD4 is degraded, and eIF4A activity is greatly enhanced. We identified a subset of genes involved in BCR signaling, including CARD11, BCL10, and MALT1, that have complex 5'UTRs and encode proteins with short half-lives. Expression of these known oncogenic proteins is enhanced on BCR activation and is attenuated by the eIF4A inhibitor Silvestrol. Antigen-experienced immunoglobulin (Ig)G(+) splenic B cells, from which most DLBCLs are derived, have higher levels of eIF4A cap-binding activity and protein translation than IgM(+) B cells. Our results suggest that eIF4A-mediated enhancement of oncogene translation may be a critical component for lymphoma progression, and specific targeting of eIF4A may be an attractive therapeutic approach in the management of human B-cell lymphomas.

The mechanisms of skin rejection in vascularized composite allotransplantation (VCA) remain incompletely understood. The formation of tertiary lymphoid organs (TLO) in hand transplantation has been recently described. We assess this phenomenon in experimental and clinical VCA rejection. Skin biopsies of human (n = 187), nonhuman primate (n = 11), and rat (n = 15) VCAs were analyzed for presence of TLO. A comprehensive immunohistochemical assessment (characterization of the cell infiltrate, expression of adhesion molecules) including staining for peripheral node addressin (PNAd) was performed and correlated with rejection and time post-transplantation. TLO were identified in human, nonhuman primate, and rat skin samples. Expression of PNAd was increased in the endothelium of vessels upon rejection in human skin (P = 0.003) and correlated with B- and T-lymphocyte numbers and LFA-1 expression. PNAd expression was observed at all time-points after transplantation and increased significantly after year 5. In nonhuman primate skin, PNAd expression was found during inflammatory conditions early and late after transplantation. In rat skin, PNAd expression was strongly associated with acute rejection and time post-transplantation. Lymphoid neogenesis and TLO formation can be uniformly found in experimental and human VCA. PNAd expression in vascular endothelium correlates with skin rejection and T- and B-cell infiltration., (© 2014 Steunstichting ESOT.)

Predicting graft outcome after renal transplantation based on donor histological features has remained elusive and is subject to institutional variability. We have shown in a retrospective study that the Maryland Aggregate Pathology Index score reliably predicts graft outcome. We sought to validate the scoring system in our center and a second transplant center. We analyzed 140 deceased donor kidneys pre-implantation biopsies from center 1 and 65 from center 2. The patients had a mean follow-up of 695 ± 424 and 656 ± 305 d respectively. Although MAPI scores were similar, there were significant differences in donor and recipient parameters between both centers. Despite this, MAPI was predictive of graft outcome for both centers by Cox univariate, multivariate and time dependent ROC analysis. For center 1 and 2, three yr graft survival within each MAPI group was statistically equivalent. The three-yr graft survival at center 1 for low, intermediate, and high MAPI groups were 84.3%, 56.5%, and 50.0%, respectively, p ≤ 0.0001, and at center 2 were 83.3%, 33.3%, and 33.3%, p = 0.006. MAPI, which is based on a pre-implantation biopsy, demonstrated similar predictive and outcome results from both centers. As expanded criteria donors (ECD) criteria have redefined marginal kidneys, MAPI has the potential to further define ECD kidneys, increase utilization, and ultimately improve outcomes., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Although transplantation of genetically modified porcine livers into baboons has yielded recipient survival for up to 7 days, survival is limited by profound thrombocytopenia, which becomes manifest almost immediately after revascularization, and by subsequent coagulopathy. Porcine von Willebrand's factor (VWF), a glycoprotein that adheres to activated platelets to initiate thrombus formation, has been shown to constitutively activate human platelets via their glycoprotein Ib (GPIb) receptors. Here, we report our pig-to-primate liver xenoperfusion model and evaluate whether targeting the GPIb-VWF axis prevents platelet sequestration., Methods: Twelve baboons underwent cross-circulation with the following extracorporeal livers: one allogeneic control with a baboon liver, 4 xenogeneic controls with a GalTKO.hCD46 pig liver, 3 GalTKO.hCD46 pig livers in recipients treated with αGPIb antibody during perfusion, and 4 GalTKO.hCD46 pig livers pre-treated with D-arginine vasopressin (DDAVP) in recipients treated with αGPIb antibody during perfusion., Results: All perfused livers appeared grossly and macroscopically normal and produced bile. Xenograft liver perfusion experiments treated with αGPIb antibody may show less platelet sequestration during the initial 2 h of perfusion. Portal venous resistance remained constant in all perfusion experiments. Platelet activation studies demonstrated platelet activation in all xenoperfusions, but not in the allogeneic perfusion., Conclusion: These observations suggest that primate platelet sequestration by porcine liver and the associated thrombocytopenia are multifactorial and perhaps partially mediated by a constitutive interaction between porcine VWF and the primate GPIb receptor. Control of platelet sequestration and consumptive coagulopathy in liver xenotransplantation will likely require a multifaceted approach in our clinically relevant perfusion model., (© 2014 John Wiley & Sons A/S.)

Background: Perioperative renal dysfunction in liver transplant recipients complicates maintenance immunosuppressive therapy, particularly in patients with hepatitis C. Calcineurin inhibitors exacerbate renal dysfunction and mammalian target-of-rapamycin inhibitors are generally avoided because of perceived perioperative risks. The authors' experience with seven liver transplant patients who received belatacept and mycophenolic acid maintenance immunosuppression is reported., Methods: A retrospective review of adult liver transplant recipients with hepatitis C receiving belatacept was conducted under Institutional Review Board approval. All patients were Epstein-Barr virus IgG seropositive. The primary endpoint was patient and graft survival, with secondary endpoints including the incidence of acute rejection, degree of renal function recovery, and occurrence of major side effects., Results: Between December 19, 2011 and January 25, 2013, seven liver transplant recipients with hepatitis C received belatacept immunosuppression in the perioperative period. The primary indication for belatacept was perioperative renal dysfunction. Belatacept was initiated between 2 and 90 days posttransplant and the duration of belatacept therapy ranged from 19 to 89 days. Patients were transitioned onto calcineurin inhibitor therapy when they reached chronic kidney disease stage 2 or better. Six-month patient and graft survival was 86%. There was one episode of graft rejection on belatacept therapy in a patient who had also had early rejection before initiation of belatacept., Conclusions: The results in this initial group of patients suggest that belatacept with mycophenolic acid may be a safe maintenance immunosuppression regimen in hepatitis C-positive liver transplant recipients with renal dysfunction, and that this regimen can serve as an effective bridge to calcineurin inhibitor therapy.

Procurement of a facial vascularized composite allograft (VCA) should allow concurrent procurement of all solid organs and ensure their integrity. Because full facial procurement is time-intensive, "simultaneous-start" procurement could entail VCA ischemia over 12 h. We procured a total face osteomyocutaneous VCA from a brain-dead donor. Bedside tracheostomy and facial mask impression were performed preoperative day 1. Solid organ recovery included heart, lungs, liver, kidneys, and pancreas. Facial dissection time was 12 h over 15 h to diminish ischemia while awaiting recipient preparation. Solid organ recovery began at 13.5 h, during midfacial osteotomies, and concluded immediately after facial explantation. Facial thoracic and abdominal teams worked concurrently. Estimated blood loss was 1300 mL, requiring five units of pRBC and two units FFP. Urine output, MAP, pH and PaO2 remained normal. All organs had good postoperative function. We propose an algorithm that allows "face first, concurrent completion" recovery of a complex facial VCA by planning multiple pathways to expedient recovery of vital organs in the event of clinical instability. Beginning the recipient operation earlier may reduce waiting time due to extensive recipient scarring causing difficult dissection., (© Copyright 2013 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Background: T regulatory cells (Tregs) have been associated with prolonged allograft survival and tolerance across a wide variety of species and organ types. We used our nonhuman primate model of facial vascularized composite allotransplantation (VCA) to study the association of Tregs with graft outcomes., Methods: We quantified Tregs in peripheral blood and allograft biopsies from nonhuman primates after heterotopic partial facial segment allotransplantation from major histocompatibility complex class I-mismatched donors using flow cytometry and immunohistochemistry. Immunosuppression consisted of tacrolimus and mycophenolate mofetil without induction or depletional therapies. Circulating and graft skin Treg values were compared with graft outcomes and with histologic grade from concurrent biopsies., Results: Treg proportion in peripheral blood ranged from 0.156% to 9.00% with a mean of 3.34%±0.22%. FoxP3 staining was observed in 3 of 29 graft biopsies. Median circulating Treg value did not predict time to Banff grade II rejection (hazard ratio, 0.9; confidence interval, 0.4-2.2) or graft loss (hazard ratio, 0.5; confidence interval, 0.01-5.3). Animals that experienced rejection did not have significantly different peripheral blood or graft skin Treg values from those that did not. Biopsy specimens with grade I or II rejection were more likely to contain Tregs (25% vs. 0%; P=0.044) despite no difference in concurrent circulating Tregs (3.56% vs. 3.36%; P=0.704)., Conclusions: These findings in a clinically relevant model suggest that Tregs may have limited prognostic value with standard immunosuppressive protocols used in VCA. Further studies are necessary to determine the specific role of Tregs in VCA and any role of Treg monitoring in clinical practice.

Background: Although the LigaSure device is widely used, its use in liver transplantation, where compounding factors of portal hypertension, coagulopathy, and thrombocytopenia exist, is poorly described., Methods: From October 1, 2011, to December 31, 2011, 6 patients underwent liver transplantation with recipient hepatectomy utilizing the LigaSure device. Outcomes using the device were compared with 6 contemporaneous patients in whom the device was not used., Results: Patient demographics, preoperative laboratory values, and Model for End-Stage Liver Disease scores were not different. Recipient hepatectomy was performed, on average, 43 minutes faster using the LigaSure device (P = .02). Although total operative time and intraoperative blood product usage were lower when the LigaSure was used, these differences did not attain statistical significance. Duration of stay and recipient readmission rates were similar., Conclusions: LigaSure vessel sealing is an efficient method for recipient hepatectomy in liver transplantation. Vessel sealing of caval, portal, and other structures can be safely performed in the setting of end-stage liver disease., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Background: Chronic rejection of vascularized composite allografts (VCA) is an emerging phenomenon that may decrease long-term allograft survival and impair allograft function. Although intimal hyperplasia has been reported in human hand transplants, chronic changes in VCAs remain poorly described., Methods: We developed a nonhuman primate model of face transplantation to evaluate the effect of various immunosuppressive regimens on allograft survival that we have previously reported. Nineteen grafts were successfully transplanted and serially biopsied to assess for rejection. Five VCA grafts with long-term survival (>200 days) were weaned off immunosuppression. We performed additional histologic and immunohistochemical studies on previously collected samples., Results: All five grafts developed features consistent with chronic rejection, including neointimal proliferation, transplant vasculopathy, vessel wall fibrosis, progressive luminal occlusion, and tertiary lymphoid follicles. Review of 186 serial allograft skin and subcutaneous tissue biopsies revealed that tertiary follicles and vascular changes developed in the absence of acute skin rejection. No relationship was found between alloantibody production and these changes., Conclusions: Recognition of these characteristics of VCA chronic rejection is important for diagnosis in clinical hand and face transplantation. Studies directed towards minimizing VCA chronic rejection responses may be required to improve long-term outcomes.

Objective: Minimally invasive techniques have expanded the donor pool for living kidney donation. We changed our approach to single-port donor nephrectomy in 2009 and have compared outcomes with traditional multiple-port laparoscopic donor nephrectomy., Background: The development of minimally invasive surgical techniques to procure kidneys from living donors has allowed expansion of living donor renal transplantation to account for one third of all renal transplants. Recent technical advancement allows for the entire surgical procedure to be done through a single incision contained within the umbilicus., Methods: We compared outcomes from 135 single-port donor nephrectomies with an immediately preceding cohort of 100 multiple-port laparoscopic donor nephrectomies. Survey data were collected from both groups to compare outcomes. Additional comparisons were made to total center experience with 1300 laparoscopic donor nephrectomies., Results: A total of 135 patients completed successful single-port donor nephrectomy without major complication or open conversion. Another 16 patients required additional port placement because of excessive intra-abdominal fat or limited abdominal domain. Compared with multiple-port donor nephrectomy, single-port patients had similar operative times to cross clamp (2.8 vs 2.6 hours; P = 0.11) that normalized after a learning curve of approximately 50 cases. Recipient creatinine levels were similar at 1 week and 1 month posttransplant. Although 36-Item Short Form Health Surveys demonstrated no significant differences, additional survey data revealed that single-port patients were more satisfied with cosmetic outcomes (P < 0.01) and the overall donation process (P = 0.01). Single-port approach had similar outcomes compared with all previous laparoscopic donor nephrectomies., Conclusions: Single-port donor nephrectomy can be integrated as a standardized approach for renal donation without additional donor risk, and with benefits of improved patient satisfaction with cosmetic and overall outcomes. Although the primary benefit is cosmetic, (a single incision predominantly contained within the umbilicus) outcomes justify application for kidney donors in experienced centers and may motivate additional living kidney donation.

Background: The central face high-energy avulsive injury has been frequently encountered and predictably managed at the R Adams Cowley Shock Trauma Center. However, despite significant surgical advances and multiple surgical procedures, the ultimate outcome continues to reveal an inanimate, insensate, and suboptimal aesthetic result., Methods: To effectively address this challenging deformity, a comprehensive multidisciplinary approach was devised. The strategy involved the foundation of a basic science laboratory, the cultivation of a supportive institutional clinical environment, the innovative application of technologies, cadaveric simulations, a real-time clinical rehearsal, and an informed and willing recipient who had the characteristic deformity., Results: After institutional review board and organ procurement organization approval, a total face, double jaw, and tongue transplantation was performed on a 37-year-old man with a central face high-energy avulsive ballistic injury., Conclusions: This facial transplant represents the most comprehensive transplant performed to date. Through a systematic approach and clinical adherence to fundamental principles of aesthetic surgery, craniofacial surgery, and microsurgery and the innovative application of technologies, restoration of human appearance and function for individuals with a devastating composite disfigurement is now a reality., Clinical Question/level of Evidence: Therapeutic, V.

Background: Transplantation of a facial vascularized composite allograft is a highly complex procedure that requires meticulous planning and affords little room for error. Although cadaveric dissections are an essential preparatory exercise, they cannot simulate the true clinical experience of facial vascularized composite allograft recovery., Methods: After obtaining institutional review board approval to perform a facial vascularized composite allograft research procurement, a 66-year-old, brain-dead donor was identified. The family graciously consented to donation of a total face, double jaw, and tongue allograft and multiple solid organs., Results: A craniofacial computed tomographic angiogram was obtained preoperatively to define the vascular anatomy and facilitate virtual computerized surgical planning. The allograft was procured in 10 hours, with an additional 2 hours required for an open tracheostomy and silicone facial impression. The donor was coagulopathic throughout the recovery, resulting in an estimated blood loss of 1500 ml. Fluorescence angiography confirmed adequate perfusion of the entire allograft based on lingual and facial arterial and external jugular and thyrolinguofacial venous pedicles. The solid organ transplant team initiated abdominal organ isolation while the facial allograft procurement was in progress. After completion of allograft recovery, the kidneys and liver were recovered without complication., Conclusions: Before conducting a clinical face transplant, adequate preparation is critical to maximize vascularized composite allotransplantation outcomes and preserve solid organ allograft function. As more centers begin to perform facial transplantation, research procurement of a facial vascularized composite allograft offers a unique educational opportunity for the surgical and anesthesia teams, the organ procurement organization, and the institution.

Background: Clinical vascularized composite allografts (VCA), although performed with good success, have been characterized by rejection episodes and postoperative graft edema. We investigated lymphatic donor-recipient reconstitution and lymphatic regeneration in a nonhuman primate facial VCA model., Methods: Heterotopic partial face (n = 9) VCAs were performed in cynomolgus macaques. Grafts were monitored for rejection episodes and response to immunosuppressive therapies as previously described. Donor and recipient lymphatic channels were evaluated using a near-infrared handheld dual-channel light-emitting diode camera system capable of detecting fluorescence from indocyanine green injections. Graft lymphatic channels were serially evaluated from postoperative day 0 to 364., Results: Preoperative imaging demonstrated superficial lymphatic anatomy similar to human anatomy. Initial resolution of facial allograft swelling coincided with superficial donor-recipient lymphatic channel reconstitution. Reconstitution occurred despite early acute rejection episodes in 2 animals. However, lymphatic channels demonstrated persistent functional and anatomic pathology, and graft edema never fully resolved. No differences in lymphatic channels were noted between grafts that developed transplant vasculopathy (n = 3) and those that did not (n = 6). Dynamic changes in patterns of lymphatic drainage were noted in 4 animals following withdrawal of immunosuppression., Conclusions: Donor-recipient lymphatic channel regeneration following VCA did not result in resolution of edema. Technical causes of graft edema may be overcome with alternative surgical techniques, allowing for direct investigation of the immunologic relationship between VCA graft edema and rejection responses. Mechanisms and timing of dynamic donor-recipient lymphatic channel relationships can be evaluated using fluorescent imaging systems to better define the immunologic role of lymphatic channels in VCA engraftment and rejection responses, which may have direct clinical implications.

Background: Controversy exists regarding the benefit of ureteral stents in kidney transplantation. We aimed to examine the association of stents with risk of ureteral complications, particularly in relationship with donor type., Methods: Kidney transplants from 2005 to 2009 were evaluated (n=1224). Patients with previous or simultaneous nonkidney transplants, death, or lost to follow-up within 90 days were excluded, unless already developed a ureteral complication. Only cases with a single extravesical ureteroneocystostomy were included. The cohort (n=961) was divided into stent (32.2%) and no-stent (67.7%) groups. Poisson regression was used to examine the association of stent with ureteral complications (leak or stricture) and urinary tract infections (UTI)., Results: Ureteral complication rate was 1.9% in stent versus 5.8% in no-stent group (P=0.007). UTI rate was 14.2% with stent versus 7.9% without stent (P=0.003). Stent use was independently associated with reduction in ureteral complications (incidence rate ratios [IRR], 0.40; P=0.04; 95% confidence interval [CI], 0.17-0.96) and an increase in UTI risk (IRR, 1.79; P=0.006; 95% CI, 1.18-2.74). Stent protective effect was primarily related to reduction in stricture risk (IRR, 0.23; P<0.05; 95% CI, 0.05-0.99). Stents were associated with a decrease in ureteral complications in deceased donor recipients (IRR, 0.34; P=0.03; 95% CI, 0.13-0.88), but not living donors (IRR, 1.24; P=0.84; 95% CI, 0.15-10.2)., Conclusions: Ureteral stents are associated with a significant reduction in ureteral complications but increases UTI risk. Routine stenting in deceased donor transplants is recommended as its protective effect was observed in this group. The value of stents in living donor transplants warrants further investigation.

Background: Vascularized composite tissue allotransplantation has demonstrated clinical success with standard immunosuppression in hand and upper extremity transplantation. The authors developed a fibular vascularized composite tissue allotransplantation model in nonhuman primates to investigate healing and rejection patterns of bone and associated tissues., Methods: Five fibular vascularized composite tissue allotransplantations were performed between mismatched cynomolgus macaques (Macaca fascicularis). Vascularized fibular segments with associated muscle and skin were transplanted to recipient forearm radius defects. Recipients were treated with either tacrolimus monotherapy or tacrolimus plus co-stimulatory blockade with a novel anti-CD28 antibody. Animals were followed for 6 months with serial radiographs, blood sample collection, and biopsies. At the study endpoint, angiographic, biomechanical, histologic, and immunologic assays were performed., Results: All animals survived to the experimental endpoint of 180 days. Rapid or immediate skin loss was evident secondary to vascular compromise (n = 3) or rejection (n = 1) in four animals. Despite loss of nonbony segments and the development of transplant arteriopathy consistent with chronic rejection in two animals, serial radiologic imaging and histology demonstrated bone healing and donor-recipient bony union by 10 weeks in all animals. Histology confirmed the presence of viable cortical and marrow elements. Biomechanical analysis supported donor-recipient bony union. Short-tandem repeated genotypic analysis revealed that donor marrow had been completely replaced by recipient marrow., Conclusions: In contrast to successes in extremity vascularized composite tissue allotransplantation, the authors' nonhuman primate fibular vascularized composite tissue allotransplantation model showed early skin loss, replacement of donor bone marrow, and chronic rejection. Donor-recipient bone union did occur and supports the potential for reconstruction of bony continuity defects using isolated vascularized bone allotransplants.

Vascularized composite allograft (VCA) transplantation (also referred to as composite tissue allotransplantation) has demonstrated clinical success in cases of hand, arm and face transplantation despite prior belief that skin provides an insurmountable barrier to allograft rejection. These overall good outcomes are facilitated by substantial immunosuppressive requirements in otherwise healthy patients, yet still demonstrate frequent rejection episodes. We developed a nonhuman primate model of facial segment allotransplantation to elucidate the unique pathophysiology and immunosuppressive requirements of VCA with addition of concomitant vascularized bone marrow (VBM). Heterotopically transplanted facial segment VCA with VBM treated only with tacrolimus and mycophenolate mofetil (MMF) demonstrated prolonged rejection-free survival, compared to VCA without VBM that demonstrated early rejection episodes and graft loss. While VCA with VBM demonstrated sporadic macrochimerism, acute and chronic rejection and graft loss occurred after discontinuation of immunosuppression. These data support an immunomodulatory role of VBM in VCA that reduces immunosuppressive requirements while providing improved outcomes., (2011 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Objectives: Cytomegalovirus (CMV) infection is responsible for significant morbidity and mortality among solid organ transplant recipients. Prophylaxis using valganciclovir (VGCV) in orthotopic liver transplant (OLT) recipients is not approved by the Food and Drug Administration and its use is controversial. This study aimed to evaluate the effectiveness of VGCV in CMV prophylaxis in OLT recipients., Methods: We carried out a retrospective, single-centre study including all OLT procedures performed during 2005-2008. Patients with early death (at ≤ 30 days), without CMV serology or prophylaxis, or with follow-up of <1 year were excluded., Results: The overall incidence of CMV disease was 6% (n= 9). The ganciclovir (GCV) and VGCV groups had similar incidences of CMV disease (4.6% vs. 7.0%; P= 0.4) and similar distributions of disease presentation (CMV syndrome vs. tissue-invasive CMV; P= 0.4). Incidences of CMV infection, as well as disease presentation, were similar between the high-risk (CMV D+/R-) and non-high-risk groups (P= 0.16). Although acute cellular rejection occurred more frequently in patients who developed CMV disease (P= 0.005), overall survival in these patients did not differ from that in patients who did not develop CMV infection (P= 0.5)., Conclusions: Valganciclovir is an effective antiviral for the prevention of CMV disease in liver transplant recipients. Our data support its use in high-risk OLT patients.

Liver transplantation is performed based on ABO blood type compatibility without dependence on crossmatch results. Combined liver-kidney transplantation (CLKT) is similarly performed without dependence of crossmatch results as the liver is thought to confer protection to the renal allograft against alloantibody. We report a case of CLKT in a sensitized patient with antibody-mediated rejection (AMR) of the renal allograft. AMR was confirmed with C4d staining and serial monitoring of donor-specific antibody (DSA). Despite intensive therapy directed against AMR and the presence of the liver allograft, the patient demonstrated increasing titers of alloantibody, never demonstrated adequate renal function, and ultimately expired after two months. This result demonstrates the potential for AMR of the renal allograft in sensitized recipients of CLKT., (© 2009 John Wiley & Sons A/S.)

C4d+ antibody-mediated rejection following pancreas transplantation has not been well characterized. Therefore, we assessed the outcomes of 27 pancreas transplantation patients (28 biopsies), with both C4d staining and donor-specific antibodies (DSA) determined, from a cohort of 257 patients. The median follow-up was 50 (interquartile range [IQR] 8-118) months. Patients were categorized into 3 groups: group 1, patients with minimal or no C4d staining and no DSA (n = 13); group 2, patients with either DSA present but no C4d, diffuse C4d+ and no DSA or focal C4d+ and DSA (n = 6); group 3, patients with diffuse C4d+ staining and DSA (n = 9). Active septal inflammation, acinar inflammation and acinar cell injury/necrosis were significantly more abundant in group 3 than in group 2 (respective p-values: 0.009; 0.033; 0.025) and in group 1 (respective p-values: 0.034; 0.009; 0.002). The overall uncensored pancreas graft survival rate for groups 1, 2 and 3 were 53.3%, 66.7% and 34.6%, respectively (p = 0.044). In conclusion, recipients of pancreas transplants with no C4d or DSA had excellent long-term graft survival in comparison with patients with both C4d+ and DSA present. Hence, C4d should be used as an additional marker in combination with DSA in the evaluation of pancreas transplant biopsies.

Background: Composite tissue allotransplantation may have different immunosuppressive requirements and manifest different complications compared with solid organ transplantation. We developed a non-human primate facial composite tissue allotransplantation model to investigate strategies to achieve prolonged graft survival and immunologic responses unique to these allografts., Methods: Composite facial subunits consisting of skin, muscle, and bone were heterotopically transplanted to mixed lymphocyte reaction-mismatched Cynomolgus macaques. Tacrolimus monotherapy was administered via continuous intravenous infusion for 28 days then tapered to daily intramuscular doses., Results: Five of the six animals treated with tacrolimus monotherapy demonstrated rejection-free graft survival up to 177 days (mean, 113 days). All animals with prolonged graft survival developed posttransplant lymphoproliferative disorders (PTLD). Three animals converted to rapamycin after 28 days of rejection of their allografts, but did not develop PTLD. Genotypic analysis of PTLD tumors demonstrated donor origin in three of the five analyzed by short-tandem repeats. Sustained alloantibodies were detected in rejecting grafts and absent in nonrejecting grafts., Conclusions: Tacrolimus monotherapy provided prolonged rejection-free survival of composite facial allografts in a non-human primate model but was associated with the development of a high frequency of donor-derived PTLD tumors. The transplantation of a large volume of vascularized bone marrow in composite tissue allografts may be a risk factor for PTLD development.

Background: Renal transplantation (RTx) in the geriatric population (age >65 years) accounts for 14% of all RTx performed nationally in 2007., Methods: We reviewed 3297 RTx recipients from our database over a 15-year period to evaluate recipient and donor age, date of transplant and graft loss, cause of graft loss, and cold ischemic time in the geriatric population., Results: Since 1991, we have performed 468 living donor RTx (LDRTx) and deceased donor RTx (DDRTx) in patients more than 65 years: 280 (65-69 years), 128 (70-74 years), and 60 (>75 years). Geriatric recipients of DDRTx demonstrated 83.0%, 74.1%, and 64.1% uncensored graft survival at 1, 3, and 5 years, respectively. Interestingly, these rates were similar compared with DDRTx in adults (18-64 years, P=0.49). Geriatric recipients of LDRTx demonstrated 1-year, 3-year, and 5-year graft survival rates of 94.3%, 88.8%, and 72.3%, respectively. Although better than geriatric DDRTx recipients, these results were not equal to the success of adult LDRTx recipients, potentially because of poorer graft survival in LDRTx recipients more than 75 years (P=0.004). Death-censored graft survivals were similar between adult and geriatric recipients of LDRTx (P=0.28). Graft loss secondary to death was twice as great in geriatric versus adult recipients (P<0.01)., Conclusions: DDRTx geriatric recipients of each group showed similar uncensored graft survivals to adult DDRTx recipients. LDRTx had better outcomes than DDRTx in geriatric recipients. Death-censored outcomes were similar between adult and geriatric LDRTx recipients. These data support the equivalent outcomes of RTx in appropriately selected geriatric recipients.

Background: The role of the innate immune system in the development of thrombotic microangiopathy (TM) after alpha1,3-galactosyltransferase gene-knockout (GTKO) pig organ transplantation in primates is uncertain., Methods: Twelve organs (nine hearts, three kidneys) from GTKO pigs were transplanted into baboons that received no immunosuppressive therapy, partial regimens, or a full regimen based on costimulation blockade. After graft failure, histologic and immunohistologic examinations were carried out., Results: Graft survival of less than 1 day was prolonged to 2 to 12 days with partial regimens (acute humoral xenograft rejection) and to 5 and 8 weeks with the full regimen (TM). Clinical or laboratory features of consumptive coagulopathy occurred in 7 of 12 baboons. Immunohistochemistry demonstrated IgM, IgG, and complement deposition in most cases. Histopathology demonstrated neutrophil and macrophage infiltrates, intravascular fibrin deposition, and platelet aggregation (TM). Grafts showed expression of primate tissue factor (TF), with increased mRNA levels, and TF was also expressed on baboon macrophages/monocytes infiltrating the graft., Conclusions: Our data suggest that (1) irrespective of the presence or absence of the adaptive immune response, early or late xenograft rejection is associated with activation of the innate immune system; and (2) porcine endothelial cell activation and primate TF expression by recipient innate immune cells may both contribute to the development of TM.

Background: Widespread application of composite tissue allotransplantation has been impeded by risks of rejection and conventional immunosuppression. The authors have developed a nonhuman primate composite tissue allotransplantation model that demonstrated reliable and long-term success necessary to progress to preclinical studies., Methods: Composite facial subunits (e.g., skin, muscle, and bone) were transplanted between mismatched cynomolgus monkeys. Vascular supply was based on the common carotid artery and external and internal jugular veins. Facial allografts were heterotopically transplanted to the recipient's lower abdomen with end-to-side anastomoses of the common carotid artery to the common femoral artery and of both the internal and external jugular veins to the common femoral vein. Animals received tacrolimus monotherapy. Grafts were inspected daily, submitted to biopsy regularly, and studied with magnetic resonance imaging., Results: Thirteen transplants were performed. Two grafts based on the common carotid artery and only the internal jugular vein failed within 3 to 5 days because of venous thrombosis not related to rejection. Subsequent transplants included anastomoses of both the internal and external jugular veins to the common femoral vein without thromboses. Immunosuppression consisted of tacrolimus monotherapy and was tolerated without complications. Long-term success was achieved with rejection-free graft survival (60 to 177 days)., Conclusions: The authors have developed the first successful model of facial composite tissue allotransplantation in a nonhuman primate. Technical requirements include preservation of both internal and external jugular venous outflow. Tacrolimus monotherapy permitted prolonged rejection-free graft survival without early complications. This model provides a platform for further investigation of composite tissue allotransplantation tolerance and requirements for indefinite survival.

Background: Pancreas transplant alone (PTA) is a controversial procedure. Without clearly demonstrated patient survival, recipients report improved quality of life. Nephrotoxic immunosuppression (IS) may exacerbate diabetic renal injury post-PTA., Methods: A single institution retrospective review of patients receiving PTA over a 14-year period was completed. Patient and donor demographics, surgical outcomes, rejection, and patient or graft survival were analyzed. Pre- and Postoperative estimated glomerular filtration rates (eGFR) were calculated based on the modification of diet and renal disease. Multivariate analysis was performed., Results: One hundred twenty-three patients undergoing 131 PTAs had an average age of 40.0 years. Seven patients were retransplanted and one received a third pancreas. Mean graft survival was 3.26 years (0-11.3 years) with 21 patients (17%) lost to follow-up. One- and 5-year patient survivals were 96.6% and 91.5%, respectively (mean, 7.15 year). Seventeen patients had an eGFR less than 50 mL/min/1.73 m preoperatively, whereas 64 patients did so post-PTA and 24 had an eGFR less than 30 mL/min. Mean eGFR pretransplantation was 88.9 vs. 55.6 posttransplantation (P<0.0001) with mean follow-up of 3.68 years. All but 16 (12%) patients showed a decrease in eGFR. Mean decrement was 32.1 mg/min/1.73 m. Thirteen developed end-stage renal disease chronic kidney disease (CKD 5) requiring kidney transplantation (KT) at a mean of 4.36 years. Eighty-three patients had an episode of rejection. In post-PTA RF, graft survival was 3.2 vs. 2.4 years (P=0.13). In those requiring KT, graft survival was 7.9 vs. 2.9 years (P<0.0001). Cold ischemia times, donor age, and preoperative eGFR for those with and without RF-requiring KT were not significant. Body mass index was statistically significant. Leukocyte-depleting agents was evaluated, but was not significant. All patients received calcineurin inhibitor IS., Conclusions: Patients who undergo PTA may be at increased risk for RF. After comparing patient and donor demographics, IS, and human leukocyte antigen mismatch, it seems that PTA is an independent risk factor for the development of renal failure. Patients with more successful pancreatic grafts demonstrated lower eGFR. Patients should be made aware of the risks of long-term IS. Only the most appropriate patients should be chosen for PTA.

Despite the common use of diagnostic pretransplant deceased donor kidney biopsy, there is no consensus on the prognostic significance of the pathologic findings. In order to assist clinicians with interpretation we analyzed 371 pretransplant biopsies and correlated the findings with graft failure. Glomerular pathology was assessed with percent glomerulosclerosis (GS), glomerular size and periglomerular fibrosis (PGF); vascular pathology with arterial wall-to-lumen ratio (WLR) and arteriolar hyalinosis and interstitial pathology with measurement of cumulative fibrosis and presence of scar. Using two-thirds of the study population as a model-development cohort, we found that biopsy features independently associated with an increased risk of graft failure were GS > or =15%, interlobular arterial WLR > or =0.5 and the presence of PGF, arteriolar hyalinosis or scar. The Maryland Aggregate Pathology Index (MAPI), was developed from these parameters and validated on the remaining one-third of the population. Five-year actuarial graft survival was 90% for kidneys with MAPI scores between 0 and 7, 63% for scores from 8 to 11 and 53% for scores from 12 to 15 (p < 0.001). We conclude MAPI may help transplant physicians estimate graft survival from the preimplantation biopsy findings, in clinical situations similar to this study population (cold ischemia over 24 h, GS < 25%).

Objective: To determine whether eliminating routine nasogastric (NG) decompression after simultaneous pancreas-kidney (SPK) transplantation would reduce hospital length of stay without any increase in complications., Background: University of Wisconsin performs all pancreas transplantations with enteric drainage of exocrine secretions. Traditionally, NG tube decompression has been used for 5 postoperative days. This strategy was supported by the fact that most patients with diabetes have a history of gastroparesis. However, to date, no study has evaluated whether NG decompression is necessary post pancreas transplantation., Methods: We have conducted a retrospective review of 182 primary SPK transplant recipients from 2002 to 2005. Before August 2004 we used NG decompression for 5 days postoperatively and resumed diet 24 hours after tube removal. After this period, diets were initiated with return of bowel function. We eliminated routine NG decompression in 2004. One hundred and thirty-two patients had NG decompression and 50 patients did not. Induction therapy changed during the study timeframe from basiliximab in the NG group to alemtuzumab in the no NG group. Maintenance therapy was similar between the 2 groups consisting of prednisone, mycophenolate mofetil, and tacrolimus., Results: Patients managed without NG tubes had significantly shorter length of stay (9.1 +/- 3.93 days) compared with patients managed with NG tube decompression (13.8 +/- 8.99 days) (P < 0.0001). Only 6 patients initially managed without NG tubes required NG placement during their hospital stay, including 2 patients returned to the operating room for nongastrointestinal complications. No differences existed between groups in complications, graft or patient survival., Conclusions: Historically, NG tube decompression has been recommended postoperatively in SPK patients. These data refute this traditional clinical practice. SPK patients managed without NG decompression have shorter hospital length of stay, equivalent graft survival, and no increased morbidity.

Campath-1H (alemtuzumab) induction was used for renal transplantation in combination with sirolimus as immunosuppression. We previously reported a high (28%) rate of early rejection with this regimen, and now report 3-year outcomes. Twenty-nine patients were recipients of either deceased donor or non-HLA (Human Leukocyte Antigen) identical living donor primary renal allografts. Clinical parameters including infection, malignancy, kidney function, and kidney histology were followed prospectively for 3 years. Three-year cumulative graft and patient survival were 96% and 100%, respectively. Twenty patients were maintained on steroid-free immunosuppressive regimens, and 15 patients were maintained on monotherapy for immunosuppression (12 on sirolimus). No serious infectious complications were observed and two patients developed basal cell skin cancer. The 3-year results of our initial pilot study demonstrate good graft (96%) and patient (100%) outcomes. Campath-1H induction has yielded a high proportion of patients maintained on immunosuppressive monotherapy (57%) without serious infectious- and no malignancy-related complications. The reported regimen yielded novel insights into both Campath-1H and sirolimus therapy in renal transplantation. Because of the higher incidence of early rejection, we recommend a modified strategy of immunosuppression including a brief course of a calcineurin inhibitor.

Models of pig-to-baboon xenografting were examined to identify the mechanisms and pathologic characteristics of acute humoral xenograft rejection (AHXR). Thymus and kidney (composite thymokidney) from human decay accelerating factor-transgenic swine were transplanted into baboons (n = 16) that were treated with an immunosuppressive regimen that included extracorporeal immunoadsorption of anti-alphaGal antibody and inhibition of complement activation. Morphologic and immunohistochemical studies were performed on protocol biopsies and graftectomy samples. All renal xenografts avoided hyperacute rejection. However, graft rejection coincided with the increase of anti-alphaGal antibody in the recipient's circulation. The 16 xenografts studied were divided into two groups dependent on the rapid return (group 1) or gradual return (group 2) of anti-alphaGal antibody after immunoadsorption. In group 1 (n = 6), grafts were rejected to day 27 with development of typical AHXR, characterized by marked interstitial hemorrhage and thrombotic microangiopathy in the renal vasculature. In group 2 (n = 10), grafts also developed thrombotic microangiopathy affecting mainly the glomeruli by day 30 but also showed minimal evidence of interstitial injury and hemorrhage. In the injured glomeruli, IgM and C4d deposition, subsequent endothelial cell death and activation with upregulation of von Willebrand factor and tissue factor, and a decrease of CD39 expression developed with the formation of fibrin-platelet multiple microthrombi. In this model, the kidney xenografts, from human decay accelerating factor-transgenic swine, in baboons undergo AHXR. In slowly evolving AHXR, graft loss is associated with the development of thrombotic microangiopathic glomerulopathy. Also, anti-alphaGal IgM deposition and subsequent complement activation play an important role in the mechanism of glomerular endothelial injury and activation and the formation of multiple microthrombi.

Tissue-invasive disease due to porcine cytomegalovirus (PCMV) has been demonstrated after pig-to-baboon solid-organ xenotransplantation. Porcine lymphotropic herpesvirus (PLHV)-1 is associated with B cell proliferation and posttransplant lymphoproliferative disorder after allogeneic bone marrow transplantation in swine but has not been observed in pig-to-primate xenotransplantation. Activation of PCMV and PLHV-1 was investigated in 22 pig-to-baboon xenotransplants by use of quantitative polymerase chain reaction. PCMV was found in all xenografts; increased viral replication occurred in 68% of xenografts during immunosuppression. PLHV-1 was found in 12 xenografts (55%); no increases in viral replication occurred during immunosuppression. Control immunosuppressed swine coinfected with PCMV and PLHV-1 had activation of PCMV but not PLHV-1. PCMV, but not PLHV-1, is activated in solid-organ xenotransplantation.

As the major site of self-nonself discrimination in the immune system, the thymus, if successfully transplanted, could potentially carry with it the induction of central tolerance to any other organ or tissue from the same donor. We have recently developed a technique for transplantation of an intact, vascularized thymic lobe (VTL) in miniature swine. In the present study, we have examined the ability of such VTL allografts to support thymopoiesis and induce transplantation tolerance across fully MHC-mismatched barriers. Six miniature swine recipients received fully MHC-mismatched VTL grafts with a 12-day course of tacrolimus. Three of these recipients were thymectomized before transplantation and accepted their VTL allografts long-term, with evidence of normal thymopoiesis. In contrast, three euthymic recipients rejected their VTL allografts. Donor renal allografts, matched to the donor VTL grafts, were transplanted without immunosuppression into two of the three thymectomized recipients, and one of the three euthymic recipients. These renal allografts were accepted by thymectomized recipients, but rejected by the euthymic recipient in an accelerated fashion. This study thus demonstrates that successful transplantation of a vascularized thymus across a fully MHC-mismatched barrier induces tolerance in this preclinical, large-animal model. This procedure should enable studies on the role of the thymus in transplantation immunology as well as offer a potential strategy for tolerance induction in clinical transplantation.

Background: This study determines whether composite thymokidney (TK) grafts, created by implantation of autologous thymic tissue beneath the donor's renal capsule before transplantation, could induce allogeneic transplantation tolerance across two-haplotype fully major histocompatibility complex (MHC)- mismatched barriers in juvenile MGH-miniature swine., Methods: TK grafts were prepared by implanting autologous thymic tissue under the renal capsule of donor animals 2 to 3 months before transplantation. Four recipients were treated with a T-cell-depleting immunotoxin and received fully MHC-mismatched TK grafts plus a 12-day course of cyclosporine A (CsA). Control animals were treated with CsA alone or both CsA and immunotoxin, but with a normal kidney or a kidney implanted with autologous lymph node rather than thymus. Renal graft function was assessed by plasma creatinine levels and histologic analyses. Immunologic status was monitored by cell-mediated lympholysis assays., Results: All four recipients of fully MHC-mismatched TK transplants treated with immunotoxin and a 12-day course of CsA accepted their composite renal allografts long-term. All control recipients receiving a TK and CsA alone, a normal kidney or a composite kidney containing lymph node tissue acutely rejected their grafts., Conclusions: To our knowledge, this is the first demonstration that functional vascularized thymic grafts can induce transplantation tolerance across fully MHC-mismatched barriers in a large animal model.

Background: The potential of xenotransplantation for clinical application will require overcoming barriers of humoral and cellular rejection, through strategies using immune suppression or tolerance induction. This laboratory has previously reported the induction of tolerance in the discordant xenogeneic model of pig-to-rodent thymic transplantation. We also have described a miniature swine model of fully mismatched allogeneic composite vascularized thymokidney transplantation that induced transplantation tolerance. We tested a combination of these approaches in a clinically relevant pig-to-primate model of xenotransplantation., Methods: Composite thymokidney grafts were prepared 40 to 80 days before transplantation by the autologous implantation of thymic tissue under the renal capsule of human decay-accelerating factor transgenic swine. Baboons received xenotransplants of both human decay-accelerating factor composite thymokidneys and omental implants of thymic tissue. Recipients were treated with an immunosuppressive-conditioning regimen including thymectomy or thymic irradiation, extracorporeal immunoadsorption of anti-alphaGal antibodies and T-cell depletion. Recipients were followed for indicators of xenograft rejection, T-cell depletion and reconstitution, anti-alphaGal antibody levels, and mixed lymphocyte responses. Immunologic responses were studied in those animals that survived for more than 3 weeks., Results: Thymokidney xenografts survived for up to 30 days, with evidence of viable thymic epithelium and Hassall's corpuscles under the renal capsule and in the omental implants, and with evidence of few host lymphocytes. Three animals demonstrated donor-specific unresponsiveness, while maintaining normal alloresponses, in mixed-lymphocyte-response assays performed after immunosuppression had been stopped. Rejected grafts demonstrated humoral damage without evidence of cellular infiltrates. After graftectomy, one animal maintained donor-specific cellular unresponsiveness and stable anti-alphaGal antibody levels for more than 2 months., Conclusions: We concluded that composite thymokidney and thymic-tissue xenotransplantation from swine to baboons can induce donor-specific cellular unresponsiveness and stable anti-alphaGal antibody levels, suggesting avoidance of sensitization after xenotransplantation. The presence of viable donor-swine thymic epithelium could have a role in the development of donor-specific T-cell tolerance. Further strategies to address humoral rejection could prolong graft survival and result in long-term tolerance to xenografts.

Background: Whereas clinical pancreatic transplantation has been highly successful in correcting the hyperglycemia of insulin-dependent diabetes mellitus (type 1), the results of islet transplantation have been disappointing. This discrepancy may be because of, at least in part, nonspecific loss of islets during the time required for revascularization. To test this hypothesis, we have designed composite kidney grafts containing vascularized autologous islets that can be used to compare the engraftment potential of vascularized versus nonvascularized islet tissue., Methods: (1) Islet-cell isolation: miniature swine underwent either partial pancreatectomy to isolate autologous islets or total pancreatectomy to isolate minor antigen-mismatched islets. Islets were purified from excised pancreatic tissue by enzymatic digestion and discontinuous density gradient purification. Isolated islets were cultured for 3 days before transplant. (2) Creation of vascularized islet kidneys (IK): autologous islets alone (n=6), minor-mismatched islets alone (n=3), and minor-mismatched islets plus simultaneous autologous thymic tissue (n=3) were transplanted beneath the renal capsule of juvenile miniature swine. Minor antigen-mismatched islets were also transplanted into both the vascularized thymic graft of a thymokidney (to produce a thymo-islet kidney [TIK]) and the contralateral native kidney (n=3) and both the host thymus and beneath the renal capsule (n=2). All recipients receiving minor-mismatched islets were treated with a 12-day intravenous (IV) course of either cyclosporine A (CsA) at 10 mg/kg per day or FK506 at 0.15 mg/kg per day. (3) Assessment of Function: to evaluate the function of the transplanted islets, three animals bearing TIK and IK underwent total pancreatectomy 3 months following islet transplantation., Results: (1) Islet-cell yields: an average of 254,960+/-51,879 (4,452+/-932 islet equivalents [IEQ]/gram of pancreas) and 374,410+/-9,548 (4,183+/-721 IEQ/gram of pancreas) viable islets were obtained by partial pancreatectomy and complete pancreatectomy, respectively. (2) Creation of IK: autologous islets engrafted indefinitely, whereas recipients of minor-mismatched islets alone rejected the islets within 2 months. However, when minor-mismatched islets were implanted into both the thymokidney and the contralateral kidney of animals bearing a thymokidney, the islets engrafted indefinitely in both sites (>3 months). Simultaneous implantation of islets into the host thymus and under the renal capsule also led to permanent engraftment of minor-mismatched islets. (3) Function of vascularized islets: three animals with both a TIK and an IK in place for 3 months underwent total pancreatectomy. All three animals maintained normoglycemia thereafter. In two of these animals, the IKs were removed 2 months after the pancreatectomy, and in both cases normoglycemia was maintained thereafter by the TIK., Conclusions: The implantation of islets beneath the autologous renal capsule permitted the establishment of a vascular supply and thereby supported normal islet-cell growth and function. The presence of thymic tissue beneath the autologous renal capsule facilitated the engraftment of minor-mismatched islets, and such grafts achieved results similar to autologous islet transplants. Therefore, the ability to create vascularized islet grafts may provide a strategy for successful islet transplantation across allogeneic and potentially across xenogeneic barriers.

We have previously reported the preparation of vascularized islet-kidneys (IKs) by transplantation of islets under the autologous kidney capsule. Here, we compare the efficacy of transplanting vascularized versus nonvascularized islets into diabetic allogeneic swine recipients. In the vascularized islet transplantation (5,000 islet equivalents [IE]/kg), recipients received minor-mismatched (n = 4) or fully-mismatched (n = 2) IKs after pancreatectomy, with a 12-day course of cyclosporine A (CyA) or FK506, respectively. For the nonvascularized islet transplantation (7,000 IE/kg), three recipients received minor-mismatched islets alone and two recipients received minor-mismatched donor islets placed in a donor kidney on the day of transplantation. All recipients of nonvascularized islets were treated with a 12-day course of CyA. With vascularized islet transplantation, pancreatectomized recipients were markedly hyperglycemic pretransplant (fasting blood glucose >300 mg/dl). After composite IK transplantation, all recipients developed and maintained normoglycemia (<120 mg/dl) and stable renal function indefinitely (>3 months), and insulin therapy was not required. Major histocompatibility complex-mismatched recipients demonstrated in vitro donor-specific unresponsiveness. In contrast, recipients of nonvascularized islets remained hyperglycemic. In conclusion, IK allografts cured surgically induced diabetes across allogeneic barriers, whereas nonvascularized islet transplants did not. These data indicate that prevascularization of islet allografts is crucial for their subsequent engraftment and that composite IKs may provide a strategy for successful islet transplantation.

Xenotransplantation of porcine organs carries the risk of reactivation of latent virus in donor and recipient tissues as well as transmission of viruses between species. We have investigated the activation of baboon cytomegalovirus (BCMV) and porcine CMV (PCMV) in a pig-to-primate model of xenotransplantation. Tissues originating from a series of six swine-to-baboon composite thymokidney xenotransplants were investigated. Four immunosuppressed baboons died (survival range, 7 to 27 days) with the graft in situ. Increases in BCMV DNA copy numbers occurred in three (75%) of these baboons and was thought to be responsible for pneumonitis and the death of one animal. In two baboons, disseminated intravascular coagulation was successfully treated by graftectomy and discontinuation of immunosuppression. PCMV was upregulated in five of six xenografts (83%). PCMV infection was associated with ureteric necrosis in one xenograft. Although significantly increased in native tissues, low levels of BCMV and PCMV were also detected in tissues other than that of the native viral host species. The cross-species presence of CMV did not appear to cause clinical or histological signs of invasive disease. Thus, viral infections with clinical disease were restricted to tissues of the native species of each virus. Intensive immune suppression currently required for xenotransplantation results in a significant risk of reactivation of latent infections by BCMV and PCMV. It is not yet known whether viral DNA detected across species lines represents cellular microchimerism, ongoing viral infection, or uptake of free virus. The observation of graft injury by PCMV demonstrates that CMV will be an important pathogen in immunosuppressed xenograft recipients. Strategies must be developed to exclude CMV from porcine organ donors.

Background: Vascularized thymokidney transplants have previously been shown to induce tolerance across major histocompatibility complex barriers. The ability to perform vascularized thymic lobe transplantation could permit such tolerance to be induced with any cotransplanted solid organ or tissue. For this reason, we have developed a technique for vascularized thymic lobe transplantation in miniature swine., Methods: Thymic vessels (n=2) were anastomosed to the carotid artery and the external jugular vein of naïve minor-mismatched recipients treated with a 12-day course of cyclosporine A (10 mg/kg/day). Graft survival and thymopoiesis were assessed by histology, immunohistochemistry, and fluorescence-activated cell sorting. Allele-specific antibodies 74-12-4 and pig allelic antigen (PAA) were used to distinguish donor and recipient cells., Results: Allografts showed intact cortical and medullary structure posttransplantation, without evidence of rejection or ischemia. Recipient thymocytes repopulated the donor cortical thymus by POD30 and increased in the cortex and medulla by POD60., Conclusions: Our study demonstrates the technical feasibility of vascularized thymic lobe transplantation and the support of thymopoiesis by such transplants in a large animal model. This technique may offer a novel strategy to induce transplant tolerance across allogeneic and xenogeneic barriers, and to support long-term thymopoiesis in immunodeficient hosts.