Regulatory B Cell-Dependent Islet Transplant Tolerance Is Also Natural Killer Cell Dependent.

Immunologic tolerance to solid organ and islet cell grafts has been achieved in various rodent models by using antibodies directed at CD45RB and Tim-1. We have shown that this form of tolerance depends on regulatory B cells (Bregs). To elucidate further the mechanism by which Bregs induce tolerance, we investigated the requirement of natural killer (NK) and NKT cells in this model. To do so, hyperglycemic B6, µMT, Beige, or CD1d-/- mice received BALB/c islet grafts and treatment with the tolerance-inducing regimen consisting of anti-CD45RB and anti-TIM1. B6 mice depleted of both NK and NKT cells by anti-NK1.1 antibody and mice deficient in NK activity (Beige) did not develop tolerance after dual-antibody treatment. In contrast, transplant tolerance induction was successful in CD1d-/- recipients (deficient in NKT cells), indicating that NK, but not NKT, cells are essential in B cell-dependent tolerance. In addition, reconstitution of Beige host with NK cells restored the ability to induce transplant tolerance with dual-antibody treatment. Transfer of tolerance by B cells from tolerant mice was also dependent on host Nk1.1+ cells. In conclusion, these results show that regulatory function of B cells is dependent on NK cells in this model of transplantation tolerance.

Prolonged Survival Following Pig-to-Primate Liver Xenotransplantation Utilizing Exogenous Coagulation Factors and Costimulation Blockade.

Since the first attempt of pig-to-primate liver xenotransplantation (LXT) in 1968, survival has been limited. We evaluated a model utilizing α-1,3-galactosyltransferase knockout donors, continuous posttransplant infusion of human prothrombin concentrate complex, and immunosuppression including anti-thymocyte globulin, FK-506, methylprednisone, and costimulation blockade (belatacept, n = 3 or anti-CD40 mAb, n = 1) to extend survival. Baboon 1 remained well until postoperative day (POD) 25, when euthanasia was required because of cholestasis and plantar ulcers. Baboon 2 was euthanized following a seizure on POD 5, despite normal liver function tests (LFTs) and no apparent pathology. Baboon 3 demonstrated initial stable liver function but was euthanized on POD 8 because of worsening LFTs. Pathology revealed C4d positivity, extensive hemorrhagic necrosis, and a focal cytomegalovirus inclusion. Baboon 4 was clinically well with stable LFTs until POD29, when euthanasia was again necessitated by plantar ulcerations and rising LFTs. Final pathology was C4d negative and without evidence of rejection, inflammation, or thrombotic microangiopathy. Thus, nearly 1-mo rejection-free survival has been achieved following LXT in two of four consecutive recipients, demonstrating that the porcine liver can support life in primates for several weeks and has encouraging potential for clinical application as a bridge to allotransplantation for patients with acute-on-chronic or fulminant hepatic failure.

Immune Exhaustion and Transplantation.

Exhaustion of lymphocyte function through chronic exposure to a high load of foreign antigen is well established for chronic viral infection and antitumor immunity and has been found to be associated with a distinct molecular program and characteristic cell surface phenotype. Although exhaustion has most commonly been studied in the context of CD8 viral responses, recent studies indicate that chronic antigen exposure may affect B cells, NK cells and CD4 T cells in a parallel manner. Limited information is available regarding the extent of lymphocyte exhaustion development in the transplant setting and its impact on anti-graft alloreactivity. By analogy to the persistence of a foreign virus, the large mass of alloantigen presented by an allograft in chronic residence could provide an ideal setting for exhausting donor-reactive T cells. The extent of T cell exhaustion occurring with various allografts, the kinetics of its development, whether exhaustion is influenced positively or negatively by different immunosuppressants, and the impact of exhaustion on graft survival and tolerance development remains a fertile area for investigation. Harnessing or encouraging the natural processes of exhaustion may provide a novel means to promote graft survival and transplantation tolerance.

The Effects of Exogenous Administration of Human Coagulation Factors Following Pig-to-Baboon Liver Xenotransplantation.

We sought to determine the effects of exogenous administration of human coagulation factors following pig-to-baboon liver xenotransplantation (LXT) using GalT-KO swine donors. After LXT, baboons received no coagulation factors (historical control, n = 1), bolus administration of a human prothrombin concentrate complex (hPCC; 2.5 mL/kg, n = 2), continuous infusion of hPCC (1.0 mL/h, n = 1) or continuous infusion of human recombinant factor VIIa (1 µg/kg per hour, n = 3). The historical control recipient demonstrated persistent thrombocytopenia despite platelet administration after transplant, along with widespread thrombotic microangiopathy (TMA). In contrast, platelet levels were maintained in bolus hPCC recipients; however, these animals quickly developed large-vessel thrombosis and TMA, leading to graft failure with shortened survival. Recipients of continuous coagulation factor administration experienced either stabilization or an increase in their circulating platelets with escalating doses. Furthermore, transfusion requirements were decreased, and hepatic TMA was noticeably absent in recipients of continuous coagulation factor infusions compared with the historical control and bolus hPCC recipients. This effect was most profound with a continuous, escalating dose of factor VIIa. Further studies are warranted because this regimen may allow for prolonged survival following LXT.

Machine Perfusion of Donor Livers for Transplantation: A Proposal for Standardized Nomenclature and Reporting Guidelines.

With increasing demand for donor organs for transplantation, machine perfusion (MP) promises to be a beneficial alternative preservation method for donor livers, particularly those considered to be of suboptimal quality, also known as extended criteria donor livers. Over the last decade, numerous studies researching MP of donor livers have been published and incredible advances have been made in both experimental and clinical research in this area. With numerous research groups working on MP, various techniques are being explored, often applying different nomenclature. The objective of this review is to catalog the differences observed in the nomenclature used in the current literature to denote various MP techniques and the manner in which methodology is reported. From this analysis, we propose a standardization of nomenclature on liver MP to maximize consistency and to enable reliable comparison and meta-analyses of studies. In addition, we propose a standardized set of guidelines for reporting the methodology of future studies on liver MP that will facilitate comparison as well as clinical implementation of liver MP procedures.

Role of Patient Factors and Practice Patterns in Determining Access to Liver Waitlist.

Geographic variability in access to care is a persistent challenge in transplantation. Little is known about how patients with end-stage liver disease are chosen for referral, evaluation and listing. Utilizing death certificate data from the Centers for Disease Control and Prevention from 2002 to 2009, estimated liver demand (ELD) was measured by aggregating annual deaths from liver disease and liver transplants performed in each donor service area (DSA). In DSAs with higher ELD, more patients per capita were listed for transplantation (p < 0.001). In addition, listing rates per ELD varied fivefold across DSAs, with more patients per ELD being transplanted in DSAs with higher listing rates (p < 0.001). After adjusting for liver donor risk index and MELD at transplant, there was no association between listing rate and posttransplant survival (HR 1.002, p = 0.77). In addition, DSAs with lower listing rates were more likely to export organs (p < 0.001) of lower liver donor risk index (p < 0.001). Listing sicker patients was associated with increased access to the waitlist and transplantation and more efficient organ utilization, but had minimal effect on posttransplant outcomes after adjusting for the resulting organ shortage.

Socioeconomic status and ethnicity of deceased donor kidney recipients compared to their donors.

Public perception and misperceptions of socioeconomic disparities affect the willingness to donate organs. To improve our understanding of the flow of deceased donor kidneys, we analyzed socioeconomic status (SES) and racial/ethnic gradients between donors and recipients. In a retrospective cohort study, traditional demographic and socioeconomic factors, as well as an SES index, were compared in 56,697 deceased kidney donor and recipient pairs transplanted between 2007 and 2012. Kidneys were more likely to be transplanted in recipients of the same racial/ethnic group as the donor (p < 0.001). Kidneys tended to go to recipients of lower SES index (50.5% of the time, p < 0.001), a relationship that remained after adjusting for other available markers of donor organ quality and SES (p < 0.001). Deceased donor kidneys do not appear to be transplanted from donors of lower SES to recipients of higher SES; this information may be useful in counseling potential donors and their families regarding the distribution of their organ gifts.

Pilot Study Evaluating Regulatory T Cell-Promoting Immunosuppression and Nonimmunogenic Donor Antigen Delivery in a Nonhuman Primate Islet Allotransplantation Model.

The full potential of islet transplantation will only be realized through the development of tolerogenic regimens that obviate the need for maintenance immunosuppression. Here, we report an immunotherapy regimen that combines 1-ethyl-3-(3'-dimethylaminopropyl)-carbodiimide (ECDI)-treated donor lymphoid cell infusion (ECDI-DLI) with thymoglobulin, anti-interleukin-6 receptor antibody and rapamycin to achieve prolonged allogeneic islet graft survival in a nonhuman primate (NHP) model. Prolonged graft survival is associated with Treg expansion, donor-specific T cell hyporesponsiveness and a transient absence of donor-specific alloantibody production during the period of graft survival. This regimen shows promise for clinical translation.

Subnormothermic machine perfusion for ex vivo preservation and recovery of the human liver for transplantation.

To reduce widespread shortages, attempts are made to use more marginal livers for transplantation. Many of these grafts are discarded for fear of inferior survival rates or biliary complications. Recent advances in organ preservation have shown that ex vivo subnormothermic machine perfusion has the potential to improve preservation and recover marginal livers pretransplantation. To determine the feasibility in human livers, we assessed the effect of 3 h of oxygenated subnormothermic machine perfusion (21°C) on seven livers discarded for transplantation. Biochemical and microscopic assessment revealed minimal injury sustained during perfusion. Improved oxygen uptake (1.30 [1.11-1.94] to 6.74 [4.15-8.16] mL O2 /min kg liver), lactate levels (4.04 [3.70-5.99] to 2.29 [1.20-3.43] mmol/L) and adenosine triphosphate content (45.0 [70.6-87.5] pmol/mg preperfusion to 167.5 [151.5-237.2] pmol/mg after perfusion) were observed. Liver function, reflected by urea, albumin and bile production, was seen during perfusion. Bile production increased and the composition of bile (bile salts/phospholipid ratio, pH and bicarbonate concentration) became more favorable. In conclusion, ex vivo subnormothermic machine perfusion effectively maintains liver function with minimal injury and sustains or improves various hepatobiliary parameters postischemia.

HCC patients suffer less from geographic differences in organ availability.

It has been suggested that the number of exception model for end-stage liver disease (MELD) points for hepatocellular carcinoma (HCC) overestimates mortality risk. Average MELD at transplant, a measure of organ availability, correlates with mortality on an intent-to-treat basis and varies by donation service area (DSA). We analyzed Scientific Registry of Transplant Recipients data from 2005 to 2010, comparing transplant and death parameters for patients transplanted with HCC exception points to patients without HCC diagnosis (non-HCC), to determine whether the two groups were impacted differentially by DSA organ availability. HCC candidates are transplanted at higher rates than non-HCC candidates and are less likely to die on the waitlist. Overall risk of death trends downward by 1% per MELD point (p = 0.65) for HCC, but increases by 7% for non-HCC patients (p < 0.0001). The difference in the change of mortality with MELD is statistically significant between HCC and non-HCC candidates p < 0.0001. Posttransplant risk of death trends downward by 2% per MELD point (p = 0.28) for HCC patients, but increases by 3% per MELD point in non-HCC patients (p = 0.027), with the difference being statistically significant with p < 0.005. In summary, increasing wait time impacts HCC candidates less than non-HCC candidates and under increased competition for donor organs, HCC candidates' advantage increases.

Human monoclonal antibody MBL-HCV1 delays HCV viral rebound following liver transplantation: a randomized controlled study.

Rapid allograft infection complicates liver transplantation (LT) in patients with hepatitis C virus (HCV). Pegylated interferon-α and ribavirin therapy after LT has significant toxicity and limited efficacy. The effect of a human monoclonal antibody targeting the HCV E2 glycoprotein (MBL-HCV1) on viral clearance was examined in a randomized, double-blind, placebo-controlled pilot study in patients infected with HCV genotype 1a undergoing LT. Subjects received 11 infusions of 50 mg/kg MBL-HCV1 (n=6) or placebo (n=5) intravenously with three infusions on day of transplant, a single infusion on days 1 through 7 and one infusion on day 14 after LT. MBL-HCV1 was well-tolerated and reduced viral load for a period ranging from 7 to 28 days. Median change in viral load (log10 IU/mL) from baseline was significantly greater (p=0.02) for the antibody-treated group (range -3.07 to -3.34) compared to placebo group (range -0.331 to -1.01) on days 3 through 6 posttransplant. MBL-HCV1 treatment significantly delayed median time to viral rebound compared to placebo treatment (18.7 days vs. 2.4 days, p<0.001). As with other HCV monotherapies, antibody-treated subjects had resistance-associated variants at the time of viral rebound. A combination study of MBL-HCV1 with a direct-acting antiviral is underway.

Anti-CD45RB/anti-TIM-1-induced tolerance requires regulatory B cells.

The role of B cells in transplant tolerance remains unclear. Although B-cell depletion often prolongs graft survival, sometimes it results in more rapid rejection, suggesting that B cells may have regulatory activity. We previously demonstrated that tolerance induction by anti-CD45RB antibody requires recipient B cells. Here, we show that anti-CD45RB in combination with anti-TIM-1 antibody has a synergistic effect, inducing tolerance in all recipients in a mouse islet allograft model. This effect depends on the presence of recipient B cells, requires B-cell IL-10 activity, and is antigen-specific. These data suggest the existence of a regulatory B-cell population that promotes tolerance via an IL-10-dependent pathway.

Early onset adenovirus infection after simultaneous kidney-pancreas transplant.

Adenoviruses (AdV) are increasingly recognized as important viral pathogens in immunocompromised hosts. The clinical spectrum ranges from asymptomatic viremia to allograft dysfunction, and death. Most of the medical literature is on AdV infection in children and bone marrow transplant recipients. We report a case of AdV in an adult recipient in the first month after simultaneous kidney-pancreas transplant with thymoglobulin induction. This is a rare report of adenovirus infection after multiorgan transplant, and is unique in that it exhibited tissue invasive disease without any localizing signs or allograft dysfunction, while other cases in medical literature had invasive disease of the allograft with allograft dysfunction, failure, or death. In addition, this is the first report of a radiologic presentation of AdV nephritis.

An unexpected counter-regulatory role of IL-10 in B-lymphocyte-mediated transplantation tolerance.

Monoclonal antibody against the CD45RB protein induces stable transplantation tolerance to multiple types of allograft. We have previously established that this tolerance protocol relies on the regulatory function of B lymphocytes for its effect. B lymphocytes have also been reported to participate in immune regulation in several other settings. In most of these systems, the regulatory function of B lymphocytes depends on the production of IL-10. Therefore, we investigated the role of IL-10 in the anti-CD45RB model of B-cell-mediated transplantation tolerance. Surprisingly, using antibody-mediated neutralization of IL-10, IL-10-deficient recipients and adoptive transfer of IL-10-deficient B lymphocytes, we found that IL-10 actually counter-regulates tolerance induced by anti-CD45RB. Furthermore, neutralization of IL-10 reduced the development of chronic allograft vasculopathy compared to anti-CD45RB alone and reduced the production of graft reactive alloantibodies. These data suggest that the participation of regulatory B lymphocytes in transplantation tolerance may be distinct from how they operate in other systems. Identifying the specific B lymphocytes that mediate transplantation tolerance and defining their mechanism of action may yield new insights into the complex cellular network through which antigen-specific tolerance is established and maintained.

Changing pattern of organ donation at a single center: are potential brain dead donors being lost to donation after cardiac death?

Donation after cardiac death (DCD) has proven effective at increasing the availability of organs for transplantation.We performed a retrospective examination of Massachusetts General Hospital (MGH) records of all 201 donors from 1/1/98 to the 11/2008, including 54 DCD, 115 DBD and 32 DCD candidates that did not progress to donation (DCD-dnp). Comparing three time periods, era 1 (01/98-12/02), era 2 (01/03-12/05) and era 3 (01/06-11/08), DCD's comprised 14.8,48.4% and 60% of donors, respectively (p = 0.002). A significant increase in the incidence of cardiovascular/cerebrovascular as cause of death was evident in era 3 versus eras 1 and 2; 74% versus 57.1% (p<0.001),as was a corresponding decrease in the incidence of traumatic death. Interestingly, we noted an increase in utilization of aggressive neurological management over time, especially in the DCD group.We detected significant changes in the make-up of the donor pool over the past decade. That the changes in diagnosis over time did not differ between DCD and DBD groups suggests this difference is not responsible for the increase in DCD rates. Instead, we suggest that changes in clinical practice, especially in management of patients with severe brain injury may account for the increased proportion of DCD.

Transplant center volume and outcomes after liver retransplantation.

Liver retransplantation surgery has a high rate of allograft failure due to patient comorbidities and technical demands of the procedure. Success of liver retransplantation could depend on surgeon experience and processes of care that relate to center volume. We performed a retrospective cohort study of adult liver retransplantation procedures performed from January 1, 1996 through December 31, 2005 using registry data from the Organ Procurement Transplantation Network. The primary outcome was 1-year allograft failure. Liver transplant centers were categorized as small, intermediate or high volume by dividing overall liver transplants into three tertiles of approximately equal size. Mean annual volume of overall liver transplants was <50 for low-volume centers, 50-88 for intermediate-volume centers and >88 for high-volume centers. The primary analysis consisted of 3977 liver retransplantation patients. The unadjusted risk of 1-year allograft failure was 37.8%. In multivariable logistic regression, the risk of 1-year allograft failure was not significantly different between low- (reference), intermediate- (OR 0.86, CI 0.72-1.03, p = 0.11) and high-volume centers (OR 0.88, CI 0.74-1.04, p = 0.14). Results were similar when the analysis was limited to retransplantation performed >160 days after initial transplantation. Center volume is an imprecise surrogate measure for 1-year outcomes after liver retransplantation.

Regulatory T-cell counter-regulation by innate immunity is a barrier to transplantation tolerance.

Innate immune signals foster adaptive immunity through activation of antigen-presenting cells. Recent in vitro evidence suggests that innate signaling may also contribute to immunity by countering the effects of regulatory T cells (T-regs), counter-regulation. We present in vivo evidence using a transgenic skin allograft model that the function of T-regs is lost in the setting of acute skin transplantation but remains intact when grafts were transplanted 1 month prior to allow surgery-induced inflammation to abate. Our findings identify T-reg counter-regulation as a naturally occurring process that accompanies transplantation and an important barrier to T-reg-mediated tolerance. Our finding further highlights the central role of regulatory cell deactivation in the initiation of the immune response.

ASTS recommended practice guidelines for controlled donation after cardiac death organ procurement and transplantation.

The American Society of Transplant Surgeons (ASTS) champions efforts to increase organ donation. Controlled donation after cardiac death (DCD) offers the family and the patient with a hopeless prognosis the option to donate when brain death criteria will not be met. Although DCD is increasing, this endeavor is still in the midst of development. DCD protocols, recovery techniques and organ acceptance criteria vary among organ procurement organizations and transplant centers. Growing enthusiasm for DCD has been tempered by the decreased yield of transplantable organs and less favorable posttransplant outcomes compared with donation after brain death. Logistics and ethics relevant to DCD engender discussion and debate among lay and medical communities. Regulatory oversight of the mandate to increase DCD and a recent lawsuit involving professional behavior during an attempted DCD have fueled scrutiny of this activity. Within this setting, the ASTS Council sought best-practice guidelines for controlled DCD organ donation and transplantation. The proposed guidelines are evidence based when possible. They cover many aspects of DCD kidney, liver and pancreas transplantation, including donor characteristics, consent, withdrawal of ventilatory support, operative technique, ischemia times, machine perfusion, recipient considerations and biliary issues. DCD organ transplantation involves unique challenges that these recommendations seek to address.