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Background: Liver transplantation stands as the primary treatment for end-stage liver disease, with demand surging in recent decades because of expanded indications. However, hepatic ischemia/reperfusion injury can lead to liver transplant failure in both deceased donor and living donor transplantation. This study explored whether preconditioning donor livers through exercise training (ExT) could mitigate cold ischemic injury posttransplantation., Methods: Donor C57BL/6 mice underwent ExT via treadmill running or remained sedentary. After 4 wk, the donor liver underwent cold storage and subsequent orthotopic liver transplantation or ex vivo warm reperfusion., Results: Donor liver from mice subjected to ExT showed significantly decreased hepatic injury on reperfusion. Tissue histology revealed decreased sinusoidal congestion, vacuolization, and hepatocellular necrosis in livers from ExT mice, and immunofluorescence staining further revealed a decreased number of apoptotic cells in ExT grafts. Livers from ExT donors expressed decreased intragraft inflammatory cytokines cascade, decreased neutrophil infiltration and neutrophil extracellular traps, and increased M2 phenotype of recipient macrophages compared with grafts from sedentary mice. After cold storage, liver grafts from ExT donors showed decreased accumulation of reactive oxygen species and decreased levels of cytochrome c and high mobility group box 1 released in the liver effluent. In addition, ExT grafts showed upregulated peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) and higher levels of mitochondrial content. Similar effects of decreased hepatic injury were observed in wild-type mice when pretreated with a PGC-1α stimulator ZLN005 instead of ExT., Conclusions: These findings suggest that augmenting hepatocytic mitochondrial content through donor exercise or PGC-1α stimulation may offer therapeutic avenues to mitigate postreperfusion inflammation and improve transplant outcomes., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Dual hypothermic oxygenated perfusion (DHOPE) is increasingly being used to extend liver preservation to improve transplant logistics. However, little is known about its benefits in high-risk liver grafts. This study aimed to investigate whether prolonged DHOPE provides benefits other than improved logistics in all liver types. We performed a national retrospective cohort study of 177 liver transplants from 12 Italian centers preserved with DHOPE for ≥4 hours between 2015 and 2022. A control group of 177 DHOPEs of <4 hours during the same period was created using 1:1 propensity score matching. The impact of risk profiles and preservation times on the outcomes was assessed using univariable and multivariable regression models. No significant differences in posttransplant outcomes were found between prolonged and short DHOPEs. However, the prolonged group had a significantly lower incidence of posttransplant acute kidney injury (AKI) compared to the short group (30.5% vs. 44.6%, p = 0.008). Among prolonged DHOPEs, no differences in transplant outcomes were observed according to donor risk index, Eurotransplant definition for marginal grafts, and balance of risk score. DHOPE duration was associated with a lower risk of AKI in multivariable models adjusted for donor risk index, Eutrotransplant marginal grafts, and balance of risk score. Prolonged hypothermic oxygenated perfusion confirmed its protective effect against AKI in a multivariable model adjusted for donor and recipient risk factors [OR: 0.412, 95% CI: 0.200-0.850, p = 0.016]. Prolonged DHOPE is widely used to improve transplant logistics, provides good results with high-risk grafts, and appears to be associated with a lower risk of posttransplant AKI. These results provide further insight into the important role of DHOPE in preventing posttransplant complications., (Copyright © 2024 American Association for the Study of Liver Diseases.)

Marginal liver grafts, such as those from cardiac death donors and donors with steatotic organs, are highly vulnerable to ischemia-reperfusion injury. In addition, ex situ graft alteration, either by reduction or splitting, will prolong the static cold storage time and amplify the ischemia-reperfusion injury. Hypothermic oxygenated machine perfusion has the potential to end the oxygen deprivation during preservation and accordingly improve outcomes in some marginal grafts that have been traditionally discarded. We present a case of a severely steatotic graftfrom a donor after brain death for which the graft was reduced ex situ during hypothermic oxygenated machine perfusion via the portal vein to shorten static cold storage duration. The liver was successfully transplanted into a 67-year-old adult recipient. Despite having a high donor risk index (10.8) and macrosteatosis of 50%, the graft showed early good function, and the recipient was discharged home with no complications and normal liver function tests in less than 2 weeks. Although this represents an anecdotal observation, we believe, in this case,thatthe possibility to provide oxygen during hypothermic organ storage enabled graft reduction without prolonging the cold ischemia time. The endischemic hypothermic oxygenated machine perfusion has the potential to transform the traditional established strategies for organ selection and preservation.

Liver transplantation is the only life-saving procedure for children with end-stage liver disease. The field is however heterogenic with various graft types, recipient age, weight, and underlying diseases. Despite recently improved overall outcomes and the expanded use of living donors, waiting list mortality remains unacceptable, particularly in small children and infants. Based on the known negative effects of elevated donor age, higher body mass index, and prolonged cold ischemia time, the number of available donors for pediatric recipients is limited. Machine perfusion has regained significant interest in the adult liver transplant population during the last decade. Ten randomized controlled trials are published with an overall advantage of machine perfusion techniques over cold storage regarding postoperative outcomes, including graft survival. The concept of hypothermic oxygenated perfusion (HOPE) was the first and only perfusion technique used for pediatric liver transplantation today. In 2018 the first pediatric candidate received a full-size graft donated after circulatory death with cold storage and HOPE, followed by a few split liver transplants after HOPE with an overall limited case number until today. One series of split procedures during HOPE was recently presented by colleagues from France with excellent results, reduced complications, and better graft survival. Such early experience paves the way for more systematic use of machine perfusion techniques for different graft types for pediatric recipients. Clinical reports of pediatric liver transplants with other perfusion techniques are awaited. Strong collaborative efforts are needed to explore the effect of perfusion techniques in this vulnerable population impacting not only the immediate posttransplant outcome but the development and success of an entire life., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

Purpose: Hepatocellular carcinoma (HCC) is one of the malignant tumors responsible for high mortality and recurrence rates. Although liver transplantation (LT) is an effective treatment option for HCC, ischemia-reperfusion injury (IRI) is a contributor to HCC recurrence after LT. Moreover, prolonged cold ischemia time (CIT) is a risk factor for IRI during LT, and there is insufficient clinical evidence regarding the impact of CIT on HCC recurrence after LT., Patients and Methods: This retrospective study analyzed 420 patients who underwent LT for HCC between February 2015 and November 2020 at The First Affiliated Hospital, Sun Yat-sen University. The duration of CIT was defined as the time from clamping of the donor aorta until portal reperfusion., Results: A total of 133 patients (31.7%) experienced tumor recurrence after LT, and CIT > 568 min was the independent risk factor for HCC recurrence (OR, 2.406; 95% CI [1.371-4.220]; p = 0.002). Multivariate Cox's regression analysis revealed that the recipients' gender, exceeding Milan criteria, poor differentiation, and alpha-fetoprotein (AFP) ≥400 ng/ml in CIT > 568 min group were independent risk factors for disease-free survival. The peak 7-day postoperative alanine aminotransferase (ALT) level ( p  < 0.001), the peak 7-day postoperative aspartate aminotransferase (AST) level ( p  < 0.001), the peak 7-day postoperative peak total bilirubin (TBIL) level ( p  = 0.012), and the incidence of early allograft dysfunction (EAD) ( p  = 0.006) were significantly higher in the CIT > 568 min group compared to the CIT ≤ 568 min group. Moreover, the amount of fresh frozen plasma (FFP) infusion during the operation increased ( p  = 0.02), and the time of mechanical ventilation postoperative was longer ( p  = 0.045)., Conclusion: An effective strategy to improve the prognosis is to reduce CIT; this strategy lowers the recurrence of HCC in patients undergoing LT, especially those within the Milan criteria., Competing Interests: The authors declare there are no competing interests., (©2024 Yu et al.)

Background: Liver transplantation (LT) is crucial for end-stage liver disease patients, but organ shortages persist. Donation after circulatory death (DCD) aims to broaden the donor pool but presents challenges. Complications like acute rejection, hepatic artery thrombosis, and biliary issues still impact posttransplant prognosis. Biomarkers, including extracellular vesicles (EVs) and microRNAs (miRNAs), show promise in understanding and monitoring posttransplant events. This study explores the role of EVs and their miRNA cargo in LT, including their potential as diagnostic tools., Methods: EVs from intrahepatic end-ischemic organ preservation solution (eiOPS) in 79 donated livers were detected using different techniques (nanosight tracking analysis, transmission electron microscopy, and flow cytometry). EV-derived miRNAs were identified by quantitative real time-polymerase chain reaction. Bioinformatics analysis was performed using the R platform., Results: Different-sized and origin-specific EVs were found in eiOPS, with significantly higher concentrations in DCD compared with donation after brain death organs. Additionally, several EV-associated miRNAs, including let-7d-5p , miR-28-5p , miR-200a-3p , miR-200b-3p , miR-200c-3p , and miR-429 , were overexpressed in DCD-derived eiOPS. These miRNAs also exhibited differential expression patterns in liver tissue biopsies. Pathway analysis revealed enrichment in signaling pathways involved in extracellular matrix organization and various cellular processes. Moreover, specific EVs and miRNAs correlated with clinical outcomes, including survival and early allograft dysfunction. A predictive model combining biomarkers and clinical variables showed promise in acute rejection detection after LT., Conclusions: These findings provide new insights into the use of EVs and miRNAs as biomarkers and their possible influence on posttransplantation outcomes, potentially contributing to improved diagnostic approaches and personalized treatment strategies in LT., Competing Interests: A.B.-M., P.P., and L.M.-A. are co-founders of Viva In Vitro Diagnostics SL, a company focused on utilizing the NLR Family Pyrin Domain Containing 3 inflammasome as a disease biomarker. A.B.-M. is also co-inventor listed on a provisional patent application for an in vitro method predicting organ transplant rejection. However, it is important to note that this research was conducted independently, without any commercial or financial associations that might be considered a conflict of interest. The other authors declare no conflicts of interest., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Introduction: Biliary complications (BCs) are a well-documented post-liver transplantation concern with potential implications for patient survival. This study aims at identifying risk factors associated with the development of BCs in recipients after liver transplantation (LT) and exploring strategies for their management., Methods: We conducted a retrospective analysis of 1595 adult patients (age > 18 years) who underwent LT surgery between 2019 and 2021. The study assessed the incidence of BCs in this cohort., Results: Of 1595 patients, 178 (11.1%) experienced BCs, while 1417 (88.8%) did not exhibit any signs of such complications. Patients who developed BCs were found to have a significantly lower average age (p < 0.001) and longer cold ischemic times (p < 0.001) compared to those without BCs. Variables such as sex, body mass index (BMI), model for end-stage liver disease (MELD) score, primary diagnosis, type of anastomosis, hepatectomy technique, type of transplanted liver and mortality did not demonstrate statistically significant differences between the two groups (p > 0.05). Univariate logistic regression analysis revealed that a cold ischemic time exceeding 12 hours and duct-to-duct anastomosis were positive predictors for BC development (odds ratios of 6.23 [CI 4.29-9.02] and 1.47 [CI 0.94-2.30], respectively). Conversely, increasing age was associated with a protective effect against BC development, with an odds ratio of 0.64 (CI 0.46-0.89)., Conclusion: Our multi-variate analysis identified cold ischemia time (CIT) as the sole significant predictor of post-liver transplantation biliary complications. Additionally, this study observed that advancing patient age had a protective influence in this context. Notably, no significant disparities were detected between hepatectomy techniques and the etiology of liver disease types in the two study groups., (© 2024. Indian Society of Gastroenterology.)

Purpose of Review: In an attempt to reduce waiting list mortality in liver transplantation, less-than-ideal quality donor livers from extended criteria donors are increasingly accepted. Predicting the outcome of these organs remains a challenge. Machine perfusion provides the unique possibility to assess donor liver viability pretransplantation and predict postreperfusion organ function., Recent Findings: Assessing liver viability during hypothermic machine perfusion remains challenging, as the liver is not metabolically active. Nevertheless, the levels of flavin mononucleotide, transaminases, lactate dehydrogenase, glucose and pH in the perfusate have proven to be predictors of liver viability. During normothermic machine perfusion, the liver is metabolically active and in addition to the perfusate levels of pH, transaminases, glucose and lactate, the production of bile is a crucial criterion for hepatocyte viability. Cholangiocyte viability can be determined by analyzing bile composition. The differences between perfusate and bile levels of pH, bicarbonate and glucose are good predictors of freedom from ischemic cholangiopathy., Summary: Although consensus is lacking regarding precise cut-off values during machine perfusion, there is general consensus on the importance of evaluating both hepatocyte and cholangiocyte compartments. The challenge is to reach consensus for increased organ utilization, while at the same time pushing the boundaries by expanding the possibilities for viability testing., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

Purpose of Review: Machine perfusion has been adopted into clinical practice in Europe since the mid-2010s and, more recently, in the United States (US) following approval of normothermic machine perfusion (NMP). We aim to review recent advances, provide discussion of potential future directions, and summarize challenges currently facing the field., Recent Findings: Both NMP and hypothermic-oxygenated perfusion (HOPE) improve overall outcomes after liver transplantation versus traditional static cold storage (SCS) and offer improved logistical flexibility. HOPE offers additional protection to the biliary system stemming from its' protection of mitochondria and lessening of ischemia-reperfusion injury. Normothermic regional perfusion (NRP) is touted to offer similar protective effects on the biliary system, though this has not been studied prospectively.The most critical question remaining is the optimal use cases for each of the three techniques (NMP, HOPE, and NRP), particularly as HOPE and NRP become more available in the US. There are additional questions regarding the most effective criteria for viability assessment and the true economic impact of these techniques. Finally, with each technique purported to allow well tolerated use of riskier grafts, there is an urgent need to define terminology for graft risk, as baseline population differences make comparison of current data challenging., Summary: Machine perfusion is now widely available in all western countries and has become an essential tool in liver transplantation. Identification of the ideal technique for each graft, optimization of viability assessment, cost-effectiveness analyses, and proper definition of graft risk are the next steps to maximizing the utility of these powerful tools., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Background: Innate immunity plays a fundamental role in solid organ transplantation. Myeloid cells can sense danger signals or DAMPs released after tissue or cell damage, such as during ischemia processes. This study aimed to identify DAMPs released during cold ischemia storage of human liver and analyze their ability to activate the inflammasome in myeloid cells and the possible implications in terms of short-term outcomes of liver transplantation., Methods: 79 samples of organ preservation solution (OPS) from 79 deceased donors were collected after cold static storage. We used different analytical methods to measure DAMPs in these end-ischemic OPS (eiOPS) samples. We also used eiOPS in the human macrophage THP-1 cell line and primary monocyte cultures to study inflammasome activation., Findings: Different DAMPs were identified in eiOPS, several of which induced both priming and activation of the NLRP3 inflammasome in human myeloid cells. Cold ischemia time and donation after circulatory death negatively influenced the DAMP signature. Moreover, the presence of oligomeric inflammasomes and interleukin-18 in eiOPS correlated with early allograft dysfunction in liver transplant patients., Interpretation: DAMPs released during cold ischemia storage prime and activate the NLRP3 inflammasome in liver macrophages after transplantation, inducing a pro-inflammatory environment that will complicate the outcome of the graft. The use of pharmacological blockers targeting DAMPs or the NLRP3 inflammasome in liver ischemia during static cold storage or through extracorporeal organ support could be a suitable strategy to increase the success of liver transplantation., Funding: Fundación Mutua Madrileña and Instituto de Salud Carlos III, Madrid, Spain., Competing Interests: Declaration of interests P.P. is scientific consultant of Glenmak Ltd, FirstThought, Donation and Transplantation Institute and Viva in vitro diagnostics. P.P. is co-inventor on patent application to use NLRP3 inflammasome as biomarker of disease, which have been licensed to Viva in vitro diagnostics S.L, a company co-funded by P.P., A.B.-M. and L.M-A. A.B-M. is co-inventor on provisional patent application of an in vitro method for predicting organ transplant rejection, but declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The rest of authors declare no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

Background: Traditional liver transplant strategies with cold preservation usually result in ischemia-reperfusion injury (IRI) to the donor liver. Regular normothermic machine perfusion (NMP) donor livers suffer IRI twice. Here, we aimed to introduce a novel technique called continuous NMP without recooling to avoid a second IRI and its application in livers from extended criteria donors., Methods: Seven donor livers transplanted following continuous NMP without recooling, 7 donor livers transplanted following standard NMP, and 14 livers under static cold storage (SCS) were included in this study. Perioperative outcomes were recorded and analyzed between groups., Results: During the NMP without a recooling procedure, all livers cleared lactate quickly to normal levels in a median time of 100 min (interquartile range, 60-180) and remained stable until the end of perfusion. In the NMP without recooling and standard NMP groups, posttransplant peak aspartate aminotransferase and alanine aminotransferase levels were both significantly lower than those in the SCS group (P = 0.0015 and 0.016, respectively). The occurrence rate of early allograft dysfunction was significantly lower in the NMP without recooling group than in the SCS group (P = 0.022), whereas there was no difference in the NMP group with or without recooling (P = 0.462)., Conclusions: Our pilot study revealed a novel technique designed to avoid secondary IRI. This novel technique is shown to have at least a comparable effect on the standard NMP, although more data are needed to show its superiority in the future., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc.)

The effectiveness of liver transplantation to cure numerous diseases, alleviate suffering, and improve patient survival has led to an ever increasing demand. Improvements in preoperative management, surgical technique, and postoperative care have allowed increasingly complicated and high-risk patients to be safely transplanted. As a result, many patients are safely transplanted in the modern era that would have been considered untransplantable in times gone by. Despite this, more gains are possible as the science behind transplantation is increasingly understood. Normothermic machine perfusion of liver grafts builds on these gains further by increasing the safe use of grafts with suboptimal features, through objective assessment of both hepatocyte and cholangiocyte function. This technology can minimize cold ischemia, but prolong total preservation time, with particular benefits for suboptimal grafts and surgically challenging recipients. In addition to more physiological and favorable preservation conditions for grafts with risk factors for poor outcome, the extended preservation time benefits operative logistics by allowing a careful explant and complicated vascular reconstruction when presented with challenging surgical scenarios. This technology represents a significant advancement in graft preservation techniques and the transplant community must continue to incorporate this technology to ensure the benefits of liver transplant are maximized., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Hann, Nutu, Clarke, Patel, Sneiders, Oo, Hartog and Perera.)

Background: Cold ischemia time (CIT) is one of the most significant variables affecting graft survival after liver transplantation. The aim of this study was to identify other predictors of worse graft survival depending on the duration of cold ischemia., Methods: This retrospective cohort study included data of liver transplant recipients and donors in the period from 2014 to 2019. A total of 724 patients were analyzed after excluding retransplatations and urgent operations. Using receiver operating characteristic analysis, we identified CIT value which divides into 2 clinically different subgroups with respect to 5-year graft loss. Within those 2 subgroups, we performed Cox proportional hazard analysis with time to graft loss as endpoint., Results: The optimal cut-off point for CIT was identified as 496 minutes. Model of end-stage liver disease score, recipient body mass index, and donor sodium concentration showed no significant effect on time to graft loss in either subgroup. For 3 factors we observed a significant effect on time to graft loss in subgroup CIT ≥496 min: transfused red cell concentrate units (hazard ratio [HR] 1.05; 95% confidence interval [CI] 1.00-1.09; P = .02), transfused fresh frozen plasma units (HR 1.04; 95% CI 1.00-1.08; P = .08), and a recipient age of >60 years (HR 1.81; 95% CI 1.10-2.98; P = .02)., Conclusions: Predictive ability of well-known risk factors for worse outcomes after liver transplantation depend on the length of cold ischemia., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Background: N-acetylcysteine infusions have been widely used to reduce ischemia/reperfusion damage to the liver; however, convincing evidence of their benefits is lacking., Objective: To perform the largest randomized controlled trial to compare the impact of N-acetylcysteine infusion during liver procurement on liver transplant outcomes., Methods: Single center, randomized trial with patients recruited from La Fe University Hospital, Spain, from February 2012 to January 2016. A total of 214 grafts were transplanted and randomized to the N-acetylcysteine group (n = 113) or to the standard protocol without N-acetylcysteine (n = 101). The primary endpoint was allograft dysfunction (Olthoff criteria). Secondary outcomes included metabolomic biomarkers of oxidative stress levels, interactions between cold ischemia time and alanine aminotransferase level and graft and patient survival (ID no. NCT01866644)., Results: The incidence of primary dysfunction was 34% (31% in the N-acetylcysteine group and 37.4% in the control group [P = 0.38]). N-acetylcysteine administration reduced the alanine aminotransferase level when cold ischemia time was longer than 6 h (P = 0.0125). Oxidative metabolites (glutathione/oxidized glutathione and ophthalmic acid) were similar in both groups (P > 0.05). Graft and patient survival rates at 12 mo and 3 y were similar between groups (P = 0.54 and P = 0.69, respectively)., Conclusions: N-acetylcysteine administration during liver procurement does not improve early allograft dysfunction according to the Olthoff classification. However, when cold ischemia time is longer than 6 h, N-acetylcysteine improves postoperative ALT levels., Competing Interests: The authors declare no funding or conflicts of interest., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

Background: A major discrepancy between clinical and most experimental settings of liver ischemia-reperfusion injury (IRI) is the allogenicity., Methods: In the current study, we first established a murine model of allogeneic orthotopic liver transplantation with extended cold ischemia time (18 h). Roles of CD4 T cells in the pathogenesis of IRI in liver allografts were determined using a depleting anti-CD4 antibody. The clinical relevance of CD4 as a marker of liver IRI was analyzed retrospectively in 55 liver transplant patients., Results: CD4 depletion in both donors and recipients resulted in the most effective protection of liver allografts from IRI, as measured by serum transaminase levels and liver histology. CD4 depletion inhibited IR-induced intragraft neutrophil/macrophage infiltration and proinflammatory gene expressions. Quantitative reverse-transcriptase polymerase chain reaction analysis of human liver biopsies (2 h postreperfusion) revealed that posttransplant, rather than pretransplant, CD4 transcript levels correlated positively with proinflammatory gene expression profile. When we divided patients into subgroups according to intragraft CD4 levels, the high CD4 cohort developed a more severe hepatocellular damage than that with low CD4 levels., Conclusions: CD4 T cells play a key pathogenic role in IRI of allogeneic liver transplants, and intragraft CD4 levels in the early postreperfusion phase may serve as a potential biomarker and therapeutic target to ameliorate liver IRI and improve orthotopic liver transplantation outcomes., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

Ischemia-reperfusion injury (IRI) is believed to contribute to graft dysfunction after liver transplantation (LT). However, studies on IRI and the impact of early allograft dysfunction (EAD) in IRI grafts are limited. Histological IRI was graded in 506 grafts from patients who had undergone LT and classified based on IRI severity (no, minimal, mild, moderate, and severe). Of the 506 grafts, 87.4% had IRI (no: 12.6%, minimal: 38.1%, mild: 35.4%, moderate: 13.0%, and severe: 0.8%). IRI severity correlated with the incidence of EAD and graft survival at 6 months. Longer cold/warm ischemia time, recipient/donor hypertension, and having a male donor were identified as independent risk factors for moderate to severe IRI. Among 70 grafts with moderate to severe IRI, 42.9% of grafts developed EAD, and grafts with EAD had significantly inferior survival compared to grafts without EAD. Longer cold ischemia time and large droplet macrovesicular steatosis (≥20%) were identified as independent risk factors for EAD. Our study demonstrated that increased IRI severity was correlated with inferior short-term graft outcomes. Careful consideration of IRI risk factors during donor-recipient matching may assist in optimizing graft utilization and LT outcomes. Furthermore, identification of risk factors of IRI-associated EAD may guide patient management and possible timely graft replacement., (© 2020 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Concepts to ameliorate the continued mismatch between demand for liver allografts and supply include the acceptance of allografts that meet extended donor criteria (ECD). ECD grafts are generally associated with an increased rate of complications such as early allograft dysfunction (EAD). The costs of liver transplantation for the health care system with respect to specific risk factors remain unclear and are subject to change. We analyzed 317 liver transplant recipients from 2013 to 2018 for outcome after liver transplantation and hospital costs in a German transplant center. In our study period, 1-year survival after transplantation was 80.1% (95% confidence interval: 75.8%-84.6%) and median hospital stay was 33 days (interquartile rage: 24), with mean hospital costs of €115,924 (SD €113,347). There was a positive correlation between costs and laboratory Model for End-Stage Liver Disease score (r s  = 0.48, P  < 0.001), and the development of EAD increased hospital costs by €26,229. ECD grafts were not associated with a higher risk of EAD in our cohort. When adjusting for recipient-associated risk factors such as laboratory Model for End-Stage Liver Disease score, recipient age, and split liver transplantation with propensity score matching, only EAD and cold ischemia increased total costs. Conclusion: Our data show that EAD leads to significantly higher hospital costs for liver transplantation, which are primarily attributed to recipient health status. Strategies to reduce the incidence of EAD are needed to control costs in liver transplantation., (© 2020 The Authors. Hepatology Communications published by Wiley Periodicals LLC on behalf of the American Association for the Study of Liver Diseases.)

Liver allograft rejection remains a significant cause of morbidity and graft failure in liver transplant recipients. Rejection is caused by the recognition of non-self donor alloantigens by recipient T-cells. Antigen recognition results in proliferation and activation of T-cells in lymphoid tissue before migration to the allograft. Activated T-cells have a variety of effector mechanisms including direct T-cell mediated damage to bile ducts, endothelium and hepatocytes and indirect effects through cytokine production and recruitment of tissue-destructive inflammatory cells. These effects explain the histological appearances of typical acute T-cell mediated rejection. In addition, donor specific antibodies, most typically against HLA antigens, may give rise to antibody-mediated rejection causing damage to the allograft primarily through endothelial injury. However, as an immune-privileged site there are several mechanisms in the liver capable of overcoming rejection and promoting tolerance to the graft, particularly in the context of recruitment of regulatory T-cells and promotors of an immunosuppressive environment. Indeed, around 20% of transplant recipients can be successfully weaned from immunosuppression. Hence, the host immunological response to the liver allograft is best regarded as a balance between rejection-promoting and tolerance-promoting factors. Understanding this balance provides insight into potential mechanisms for novel anti-rejection therapies., (Copyright © 2020 Ronca, Wootton, Milani and Cain.)

Background: The detrimental role of platelets in sinusoidal endothelial cell (SEC) injury during liver transplantation (LT) has been previously addressed after static cold storage (SCS), however, it is currently unknown after normothermic ex vivo liver perfusion (NEVLP)., Methods: Pig LT was performed with livers from heart-beating donors or donation after circulatory death (DCD) donors subjected to SCS or NEVLP (n = 5/group)., Results: All pigs except for 1 (DCD-SCS-group) survived 4 days. The heart-beating donor- and DCD-NEVLP-groups showed significantly lower aspartate transaminase-levels compared with the SCS-groups 3 hours post-LT (P = 0.006), on postoperative day (POD) 2 (P = 0.005), POD3 (P = 0.007), and on POD4 (P = 0.012). Post-LT total platelet count recovered faster in the NEVLP than in the SCS-groups at 12 hours (P = 0.023) and 24 hours (P = 0.0038). Intrahepatic sequestration of platelets was significantly higher in the SCS-groups 3 hours postreperfusion and correlated with severity of SEC injury. In both SCS-groups, levels of tumor growth factor-β were higher 3 hours post-LT, on POD1 and on POD3. Moreover, platelet factor 4 levels and platelet-derived extracellular vesicles were increased in the SCS-groups. Hyaluronic acid levels were significantly higher in the SCS-groups, indicating a higher grade of endothelial cell dysfunction. Platelet inhibition achieved by pretreatment with clopidogrel (n = 3) partly reversed the detrimental effects on SEC injury and therefore provided further evidence of the important role of platelets in ischemia/reperfusion injury and SEC injury., Conclusions: Normothermic perfusion of liver grafts before transplantation effectively reduced platelet aggregation and SEC injury, which translated into an improved posttransplant organ function.

Introduction: Despite increasing awareness of the negative impact of cold ischemia time (CIT) in liver transplantation, its precise influence in different subgroups of liver transplant recipients has not been analyzed in detail. This study aimed to identify liver transplant recipients with an unfavorable outcome due to prolonged cold ischemia. Methods: 40,288 adult liver transplantations, performed between 1998 and 2017 and reported to the Collaborative Transplant Study were analyzed. Results: Prolonged CIT significantly reduced graft and patient survival only during the first post-transplant year. On average, each hour added to the cold ischemia was associated with a 3.4% increase in the risk of graft loss (hazard ratio (HR) 1.034, P < 0.001). The impact of CIT was strongest in patients with hepatitis C-related (HCV) cirrhosis with a 24% higher risk of graft loss already at 8-9 h (HR 1.24, 95% CI 1.05-1.47, P = 0.011) and 64% higher risk at ≥14 h (HR 1.64, 95% CI 1.30-2.09, P < 0.001). In contrast, patients with hepatocellular cancer (HCC) and alcoholic cirrhosis tolerated longer ischemia times up to <10 and <12 h, respectively, without significant impact on graft survival ( P = 0.47 and 0.42). In HCC patients with model of end-stage liver disease scores (MELD) <20, graft survival was not significantly impaired in the cases of CIT up to 13 h. Conclusion: The negative influence of CIT on liver transplant outcome depends on the underlying disease, patients with HCV-related cirrhosis being at the highest risk of graft loss due to prolonged cold ischemia. Grafts with longer cold preservation times should preferentially be allocated to recipients with alcoholic cirrhosis and HCC patients with MELD <20, in whom the effect of cold ischemia is less pronounced., (Copyright © 2020 Lozanovski, Döhler, Weiss, Mehrabi and Süsal.)

Objectives: In this study, we aimed to investigate the pathologic and ultrastructural changes in transplanted mouse livers after different durations of cold storage by testing indicators of liver function and energy metabolism. We aimed to describe the effects of cold storage on liver function and the mechanisms of cold storage damage., Materials and Methods: We randomly placed 8-weekold male C57BL/6 mice into the following 4 groups to establish a cold-preserved mouse model of liver transplant: a normal control group and 3 cold storage groups, in which livers were stored for 4, 12, and 24 hours. Hepatic morphology, ultrastructural changes, and glycogenolysis were observed by hematoxylin and eosin staining, periodic acid-Schiff staining, and transmission electron microscopy. After different durations of cold storage, livers were reperfused with 4°C University of Wisconsin solution to obtain perfusion fluid, and alanine and aspartate aminotransferase levels were measured. Glycogen synthase, hypoxia-inducible factor-1α, Krüppel-like factor 2, and endothelial nitric oxide synthase mRNA expression levels in liver tissues were detected by real-time polymerase chain reaction, and aquaporin 8 protein expression levels in liver tissues were detected by Western blot., Results: Hematoxylin and eosin staining and electron microscopy ofliver showed signs ofinjury after 12 hours of cold storage, which included mainly cytoplasmic edema characterized by loose liver cell arrangement, increased hepatic sinus fissure, mitochondrial swelling, and nuclear pyknosis. Periodic acid-Schiff staining showed that glycogen content was significantly reduced, with glycogen synthase levels also reduced. Alanine aminotransferase and aspartate aminotransferase levels gradually increasedwith cold storage. Glycogen synthase, Krüppel-like factor 2, endothelial nitric oxide synthase, and aquaporin 8 expression levels also gradually increased in liver tissue. These levels gradually decreased, but hypoxia-inducible factor-1α increased., Conclusions: Mouse livers showed progressive damage to structure and function during cold storage, with mitochondrial damage perhaps showing the earliest damage.

Background: Early allograft dysfunction (EAD) is a severe complication after liver transplantation. The associated risk factors and complications have re-gained recent interest. This study investigated risk factors, survival and complications associated with EAD in a large liver transplant center in Latin America., Methods: Retrospective, unicenter, cohort, based on data from adult patients undergoing first deceased-donor liver transplant from January 2009 to December 2013. EAD was defined by one or more of the following: (i) bilirubin ≥10 mg/dL on postoperative day 7; (ii) international normalized ratio ≥1.6 on postoperative day 7, and (iii) alanine aminotransferase or aspartate aminotransferase >2000 IU/L within the first seven days after transplant., Results: A total of 602 patients were included; of these 34.2% developed EAD. Donor risk factors were male (P = 0.007), age between 50 and 59 years (P = 0.034), overweight (P = 0.028) or grade I obesity (P = 0.012), sodium >157 mmol/L (P = 0.002) and grade IV ischemia/reperfusion injury (P = 0.002). Cold ischemia time ≥10 h (P = 0.008) and warm ischemia time ≥40 min (P = 0.013) were the surgical factors. Male (P <0.001) was the only recipient protective factor. Compared with the non-EAD group, patients with EAD were submitted to more reoperations (24.3% vs. 13.4%, P = 0.001) and had higher graft loss rates (37.9% vs. 21.2%, P <0.001), with similar patient survival rates (P = 0.238)., Conclusions: EAD risk factors are related to donor, surgical procedure and recipient. Donor risk factors for EAD were male, age between 50 and 59 years, donor overweight or grade I obesity, sodium >157 mmol/L and grade IV ischemia/reperfusion injury. Cold ischemia time ≥10 h and warm ischemia time ≥40 min were the surgical risk factors. Male was the only recipient protective factor. Patients with EAD had higher reoperations and graft loss rates., (Copyright © 2019 First Affiliated Hospital, Zhejiang University School of Medicine in China. Published by Elsevier B.V. All rights reserved.)

The use of donation after circulatory death (DCD) liver allografts has been constrained by limitations in the duration of donor warm ischemia time (DWIT), donor agonal time (DAT), and cold ischemia time (CIT). The purpose of this study is to assess the impact of longer DWIT, DAT, and CIT on graft survival and other outcomes in DCD liver transplants. The Scientific Registry of Transplant Recipients was queried for adult liver transplants from DCD donors between 2009 and 2015. Donor, recipient, and center variables were included in the analysis. During the study period, 2107 patients underwent liver transplant with DCD allografts. In most patients, DWIT and DAT were <30 minutes. DWIT was <30 minutes in 1804 donors, between 30 and 40 minutes in 248, and >40 minutes in 37. There was no difference in graft survival, duration of posttransplant hospital length of stay, and readmission rate between DCD liver transplants from donors with DWIT <30 minutes and DWIT between 30 and 40 minutes. Similar outcomes were noted for DAT. In the multivariate analysis, DAT and DWIT were not associated with graft loss. The predictors associated with graft loss were donor age, donor sharing, CIT, recipient admission to the intensive care unit, recipient ventilator dependence, Model for End-Stage Liver Disease score, and low-volume transplant centers. Any CIT cutoff >4 hours was associated with increased risk for graft loss. Longer CIT was also associated with a longer posttransplant hospital stay, higher rate of primary nonfunction, and hyperbilirubinemia. In conclusion, slightly longer DAT and DWIT (up to 40 minutes) were not associated with graft loss, longer posttransplant hospitalization, or hospital readmissions, whereas longer CIT was associated with worse outcomes after DCD liver transplants., (Copyright © 2019 by the American Association for the Study of Liver Diseases.)

Organ transplantation is the treatment of choice against terminal and irreversible organ failure. Optimal preservation of the graft is crucial to counteract cold ischemia effects. As we developed an N,N-bis-2-hydroxyethyl-2-aminoethanesulfonic acid-gluconate-polyethylene glycol (BGP)-based solution (hypothermic machine perfusion [HMP]), we aimed to analyze the use of this solution on static cold storage (SCS) of rat livers for transplantation as compared with the histidine tryptophan ketoglutarate (HTK) preservation solution. Livers procured from adult male Sprague Dawley rats were preserved with BGP-HMP or HTK solutions. Liver total water content and metabolites were measured during the SCS at 0°C for 24 hours. The function and viability of the preserved rat livers were first assessed ex vivo after rewarming (90 minutes at 37°C) and in vivo using the experimental model of reduced-size heterotopic liver transplantation. After SCS, the water and glycogen content in both groups remained unchanged as well as the tissue glutathione concentration. In the ex vivo studies, livers preserved with the BGP-HMP solution were hemodynamically more efficient and the O 2 consumption rate was higher than in livers from the HTK group. Bile production and glycogen content after 90 minutes of normothermic reperfusion was diminished in both groups compared with the control group. Cellular integrity of the BGP-HMP group was better, and the histological damage was reversible. In the in vivo model, HTK-preserved livers showed a greater degree of histological injury and higher apoptosis compared with the BGP-HMP group. In conclusion, our results suggest a better role of the BGP-HMP solution compared with HTK in preventing ischemia/reperfusion injury in the rat liver model., (Copyright © 2019 by the American Association for the Study of Liver Diseases.)

Aim: Acute kidney injury (AKI) commonly occurs in patients after liver transplantation (LT). However, few studies have focused on AKI and its correlation with clinical outcomes under the Kidney Disease Improving Global Outcomes (KDIGO) criteria. This study aimed to identity the incidence, risk factors, and impacts of early AKI on outcomes in LT recipients with normal preoperative renal function, according to the KDIGO criteria., Methods: Clinical and laboratory data of 227 patients with normal preoperative renal function who underwent LT from January 2011 to January 2015 were retrospectively analyzed., Results: During the first week after LT, 106 patients (46.7%) developed AKI based on the KDIGO criteria. A multivariate analysis revealed that BMI of > 25, prolonged inferior vena cava clamping, prolonged cold ischemia time, and post-operative RBC requirements > 10 units were independent risk factors for AKI after LT. The area under the receiver operating characteristic curve for the predictive ability of AKI under these risk factors was 0.748. The occurrence of AKI was associated with longer mechanical ventilation time and post-operative ICU stay, increased post-operative 30-day mortality and decreased long-term patient survival., Conclusions: Even in patients with normal preoperative renal function, AKI was a frequent complication in LT recipients and had both negative short- or long-term effects on patient outcomes, also the severity of AKI had a dose-response relationship with worse outcomes. Patients with BMI > 25, prolonged inferior vena cava clamping, prolonged cold ischemia time, or post-operative RBC requirement > 10 units should be pay particular attention, which may assist in achieving better clinical outcomes., (Copyright © 2018. Published by Elsevier Masson SAS.)

Normothermic machine perfusion (NMP) has been shown to protect livers from injury between procurement and transplantation in a randomized controlled trial, where the machine was transported to and from the donor center. The aim of this study was to determine whether an alternative, more practical back-to-base approach after initial static cold storage would compromise beneficial outcomes. Between February 2015 and June 2018, a nonrandomized pilot study was performed at a single site. Outcomes of back-to-base livers (n = 26) were compared with those of grafts procured locally that underwent immediate NMP (n = 17). The primary outcome measure (safety) was defined as 30-day patient and graft survival. A total of 46 liver grafts were perfused with NMP, of which 3 were discarded based on poor ex situ perfusion function. The 30-day patient and graft survival in the back-to-base and local NMP groups were both 100% (primary outcome: safety). Despite significantly prolonged mean cold ischemia time (6 versus 3.2 hours; P = 0.001), the back-to-base livers demonstrated no difference in graft function, incidence of complications, or graft and patient survival. In conclusion, the back-to-base approach was safe, did not compromise the overall benefit of NMP, and offers a practical alternative to portable normothermic ex situ machine transport., (Copyright © 2019 by the American Association for the Study of Liver Diseases.)

The deep peribiliary glands (DPBG) are a niche of progenitor cells in the wall of the biliary duct (BD) and are the second line of multiplication when severe lesion of the epithelium occurs. Previous studies have identified DPBG injury as a cause of post-liver transplant (LT) biliary stenosis; this complication is a major cause of post-LT morbidity. The incidence of biliary stenosis in our center is high (38.1%). This study evaluates the lesion of DPBG in response to ischemia. Graft BD was collected in adult LT between August 2016-July 2017, from donation after brain death. Samples of 45 grafts were collected at 2 moments: BD1-during graft preparation and BD2-before biliary anastomosis. Histological analysis of the samples was performed and then classified according to degree of lesion (0, ≤50%, and >50%). A comparison was made between the degree of lesion and graft ischemia, graft histology, donor, and procurement variables. The DPBG lesion was more frequent in BD2 (20.9% vs 7%, P = .079). BD2 lesions with DPBG lesions had higher medians and means at all times of ischemia. The difference was greater in the warm ischemia time (0: 43.3 ± 12.53 minutes vs ≤50%: 52.4 ± 14.38 minutes, P = .068). The group of BD1 with DPBG lesion presented superior median cold ischemia time (CIT). In the analysis of the remaining variables there were also no statistically significant differences. We concluded that during the period of CIT there is already lesion of the DPBG, which increases after reperfusion of the graft, in greater association with longer warm ischemia time., (Copyright © 2019. Published by Elsevier Inc.)

Ex situ normothermic machine perfusion (NMP) might minimize ischemia/reperfusion injury (IRI) of liver grafts. In this study, 20 primary liver transplantation recipients of older grafts (≥70 years) were randomized 1:1 to NMP or cold storage (CS) groups. The primary study endpoint was to evaluate graft and patient survival at 6 months posttransplantation. The secondary endpoint was to evaluate liver and bile duct biopsies; IRI by means of peak transaminases within 7 days after surgery; and incidence of biliary complications at month 6. Liver and bile duct biopsies were collected at bench surgery, end of ex situ NMP, and end of transplant surgery. Interleukin (IL) 6, IL10, and tumor necrosis factor α (TNF-α) perfusate concentrations were tested during NMP. All grafts were successfully transplanted. Median (interquartile range) posttransplant aspartate aminotransferase peak was 709 (371-1575) IU/L for NMP and 574 (377-1162) IU/L for CS (P = 0.597). There was 1 hepatic artery thrombosis in the NMP group and 1 death in the CS group. In NMP, we observed high TNF-α perfusate levels, and these were inversely correlated with lactate (P < 0.001). Electron microscopy showed decreased mitochondrial volume density and steatosis and an increased volume density of autophagic vacuoles at the end of transplantation in NMP versus CS patients (P < 0.001). Use of NMP with older liver grafts is associated with histological evidence of reduced IRI, although the clinical benefit remains to be demonstrated., (Copyright © 2018 by the American Association for the Study of Liver Diseases.)

Cold storage is a common procedure for liver preservation in a transplant setting. However, during cold ischemia, the liver suffers molecular alterations that can affect its performance. Also, deleterious mechanisms set forth in the storage phase are exacerbated during reperfusion. This study aimed to identify liver proteins associated with injury during cold storage and/or normothermic reperfusion using the isolated perfused rat liver model. Livers from male rats were subjected to either (1) cold storage for 24 h, (2) ex vivo normothermic reperfusion for 90 min or (3) cold storage for 24 h followed by ex vivo normothermic reperfusion for 90 min. Then, the livers were homogenized and proteins were extracted. Protein expression between each experimental group and the control (freshly resected livers) was compared by two-dimensional (2D) gel electrophoresis. Protein identification was carried out by matrix-assisted laser desorption/ionization time-of-flight spectrometry (MALDI-TOF/TOF) using MASCOT as the search engine. 23 proteins were detected with significantly altered levels of expression among the different treatments, including molecular chaperones, antioxidant enzymes, and proteins involved in energy metabolism. Some of them have been postulated as biomarkers for liver damage while others had been identified in other organs subjected to ischemia and reperfusion injury. The whole data set will be a useful resource for studying deleterious molecular mechanisms that result in diminished liver function during storage and subsequent reperfusion., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background: In liver transplantation, both cold and warm ischemia times are known to impact early graft function. The extraction time is a period during the initial phase of organ cooling which occurs during deceased donor procurement. During this time, the organ is at risk of suboptimal cooling. Whether donor extraction time, the time from donor aortic cross-clamp to removal of the donor organ from the body cavity has an effect on early graft function is not known., Methods: We investigated the effect of donor extraction time on early graft function in 292 recipients of liver grafts procured locally and transplanted at our center between June 2012 and December 2016. Early graft function was assessed using the model of early allograft function score in a multivariable regression model including donor extraction time, cold ischemia time, warm ischemia time, donor risk index, and terminal donor sodium., Results: Donor extraction time had an independent effect on early graft function measured by the model of early allograft function score (coefficient, 0.021; 95% confidence interval, 0.007-0.035; P < 0.01; for each minute increase of donor extraction time). Besides donor extraction time, cold ischemia time, warm ischemia time, and donor risk index had a significant effect on early graft function., Conclusions: We demonstrate an independent effect of donor extraction time on graft function after liver transplantation. Efforts to minimize donor extraction time could improve early graft function in liver transplantation.

Treatment of donation after brain death (DBD) donors with low-dose dopamine improves the outcomes after kidney and heart transplantation. This study investigates the course of liver allografts from multiorgan donors enrolled in the randomized dopamine trial between 2004 and 2007 (clinicaltrials.gov identifier: NCT00115115). There were 264 hemodynamically stable DBDs who were randomly assigned to receive low-dose dopamine. Dopamine was infused at 4 μg/kg/minute for a median duration of 6.0 hours (interquartile range, 4.4-7.5 hours). We assessed the outcomes of 212 liver transplantations (LTs) performed at 32 European centers. Donors and recipients of both groups were very similar in baseline characteristics. Pretransplant laboratory Model for End-Stage Liver Disease score was not different in recipients of a dopamine-treated versus untreated graft (18 ± 8 versus 20 ± 8; P = 0.12). Mean cold ischemia time was 10.6 ± 2.9 versus 10.1 ± 2.8 hours (P = 0.24). No differences occurred in biopsy-proven rejection episodes (14.4% versus 15.7%; P = 0.85), requirement of hemofiltration (27.9% versus 31.5%; P = 0.65), the need for early retransplantation (5.8% versus 6.5%; P > 0.99), the incidence of primary nonfunction (7.7% versus 8.3%; P > 0.99), and in-hospital mortality (15.4% versus 14.8%; P > 0.99). Graft survival was 71.2% versus 73.2% and 59.6% versus 62.0% at 2 and 3 years (log-rank P = 0.71). Patient survival was 76.0% versus 78.7% and 65.4% versus 69.4% at 1 and 3 years (log-rank P = 0.50). In conclusion, donor pretreatment with dopamine has no short-term or longterm effects on outcome after LT. Therefore, low-dose dopamine pretreatment can safely be implemented as the standard of care in hemodynamically stable DBDs., (© 2018 by the American Association for the Study of Liver Diseases.)

Objective: We investigated the protective effect of ulinastatin combined with dexmedetomidine on lung injury after hepatic ischemia-reperfusion in rats., Materials and Methods: A total of 60 healthy and clean male Sprague Dawley (SD) rats were divided into the blank control group (group O), the model control group (group K), the ulinastatin and dexmedetomidine group (group F) according to random number table with 20 rats in each group., Results: The plasma concentrations of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), interleukin-8 (IL-8) and malondialdehyde (MDA) at T1, T2 and T3 time points in rats of the three groups were significantly higher than those of the T0 time point (p<0.05). The superoxide dismutase (SOD) activity in the plasma of rats of the three groups was significantly lower at T1, T2 and T3 time point when compared with that of T0 (p<0.05). The concentrations of TNF-α, IL-6, IL-8 and MDA in group K at T1, T2 and T3 moments were significantly higher than those of group O (p<0.05). However, the concentrations of IL-6, IL-8, TNF-α and MDA in group F at T1, T2, T3 were significantly lower than those of group K (p<0.05). The activities of SOD in group K at T1, T2, T3 were all significantly higher than those of group O (p<0.05). Meanwhile, the activities of SOD in group F at T1, T2, T3 were significantly higher than those of group K (p<0.05)., Conclusions: Ulinastatin combined with dexmedetomidine can reduce the inflammatory response and inhibit lipid peroxidation, eventually alleviating acute lung injury after hepatic ischemia-reperfusion in rats.

BACKGROUND Organ donation from a deceased donor, which is donation after brain death followed by circulatory death, is a unique transplantation practice in China. Pathological features of grafts help guide the utilization of grafts. MATERIAL AND METHODS We retrospectively reviewed our experiences in 188 DBCD allografts from May 2014 to April 2017. We divided 183 transplanted allografts into 3 groups according to pretransplant histology: the good quality graft group (n=62), the preservation injury group (n=27), and the steatotic graft group (n=94). Univariate and multivariate analyses were performed to identify factors in the steatotic graft group predicting the prognoses. RESULTS The prevalence rates of allografts in the good quality, steatotic liver, and preservation injury groups were 33.0% (62/188), 50.0% (94/188), and 14.4%(27/188), respectively, and the discarded rate was 2.7% (5/188). The 1- and 3-year overall survival rates were 92.1% and 88.1%, respectively. There were no differences in 1- and 3-year patient survival among the 3 groups (p=0.615). Some complications occurred: acute rejection in 7 cases, lung infection in 11 recipients, biliary stricture and bile leak in 9 patients, and portal thrombosis in 1 recipient; 17 recipients died of various causes. Cox multivariate analysis revealed that longer cold storage time was associated with worse outcome in the steatotic graft group. CONCLUSIONS Clinical outcomes of adult liver transplantation from deceased donation in China are acceptable.

Background: We previously reported that short oxygenated warm perfusion before cold storage (CS) had improved the graft viability of rat livers from donors after circulatory death (DCD). In this study, we investigated the effectiveness of short-term oxygenated subnormothermic perfusion for different durations after CS in a rat DCD model., Methods: We used an isolated perfused rat liver system. In study 1: the grafts were retrieved from Wistar rats 30 minutes after cardiac arrest (thoracotomy), preserved in CS for 6 hours, and perfused with oxygenated subnormothermic (20-25°C) Krebs-Henseleit buffer for different durations (0, 15, 30, 60, and 90 minutes groups; n = 5 in each). In study 2: in addition to subnormothermic ex vivo liver perfusion (SELP), after 15-minute incubation at room temperature, the grafts were reperfused under normothermic condition for 60 minutes as a model of liver transplantation (0, 30, 60, and 90 minutes groups; n = 5 in each)., Results: In study 1, portal flow, bile production and tissue adenosine triphosphate increased with perfusion duration. In study 2, SELP significantly improved portal flow volume (P <0.05), and bile production (P <0.05), decreased liver enzymes (P <0.05) and cytokines (P <0.0001), and increased tissue adenosine triphosphate (P <0.01). Histological examinations showed that additional SELP ameliorated tissue deterioration, preserved the parenchymal structure, and decreased apoptosis (P <0.01). Furthermore, scanning electron microscopy revealed that additional SELP alleviated sinusoidal endothelial cells and hepatic microvasculature., Conclusions: Even 30 minutes of SELP after CS rescued DCD livers from ischemia-reperfusion injury, which may help the viability of the grafts.

Background & Aims: Primary nonfunction (PNF) is a rare complication after liver transplantation that requires urgent retransplantation. PNF is associated with livers from extended criteria donors. Clinical and biochemical factors have not been identified that reliably associate with graft function after liver transplantation. Serum patterns of N-glycans associate with changes in the liver. We analyzed perfusate from grafted liver to identify protein glycosylation patterns associated with PNF., Methods: We performed a prospective study of consecutive patients who underwent liver transplantation (66 patients, from 1 center, in the derivation set, and 56 patients, from 2 centers, in the validation set) in Belgium, from October 1, 2011, through April 30, 2017. All donor grafts were transported using cold static storage, and perfusate samples were collected from the livers by flushing of hepatic veins before transplantation. Protein-linked N-glycans were isolated from perfusate samples and analyzed with a multicapillary electrophoresis-based ABI3130 sequencer. We compared glycan patterns between patients with vs without PNF of transplanted livers. PNF was defined as the need for urgent retransplantation when a graft had no evidence of function, after exclusion of other causes, such as hepatic artery thrombosis or acute cellular rejection., Results: The relative abundance of a single glycan, agalacto core-alpha-1,6-fucosylated biantennary glycan (NGA2F) was significantly increased in perfusate of livers given to 4 patients who developed PNF after liver transplantation compared with livers given to patients who did not develop PNF. Level of NGA2F identified patients with PNF with 100% accuracy. This glycomarker was the only factor associated with PNF in multivariate analysis in the derivation and the validation sets (P < .0001)., Conclusions: In an analysis of patients who underwent liver transplantation, we associated graft perfusate level of glycan NGA2F present on perfusate proteins with development of PNF with 100% accuracy, and validated this finding in a separate cohort of patients. This biomarker might be used to assess grafts before transplantation, especially when high-risk organs are under consideration., (Copyright © 2018 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Liver transplantation is a therapeutic regimen to treat patients with non-malignant end-stage liver diseases and malignant tumors of hepatic origin. The ischemia/reperfusion (I/R) injury in liver transplantation is associated with disruption of mitochondrial function in the hepatic parenchyma. Several studies have been conducted in animal models to identify pharmacological therapeutic strategies to minimize the injury induced by the cold/warm I/R in liver transplantation. Most of these studies were conducted in unrealistic conditions without the potential to be translated to clinical usage. Berberine (BBR) is a pharmacological compound with a potential protective effect of the mitochondrial function in the context of I/R. For the future clinical application of these pharmacological strategies, it is essential that a close resemblance exists between the methodology used in the animals models and real life. In this study, we have demonstrated that the addition of BBR to the preservation solution in an I/R setting preserves mitochondrial function and bioenergetics, protecting the liver from the deleterious effects caused by I/R. As such, BBR has the potential to be used as a pharmacological therapeutic strategy., Competing Interests: The authors declare no conflict of interest.

It has been shown that combined liver-kidney normothermic machine perfusion (NMP) is able to better maintain the circuit's biochemical milieu. Nevertheless, whether the combined perfusion is superior to liver perfusion alone in protecting livers from donation after circulatory death (DCD) is unclear. We aimed to test the hypothesis and explored the mechanisms. Livers from 15 DCD pig donors were subjected to either static cold storage (group A), liver-alone NMP (group B), or combined liver-kidney NMP (group C). Livers were preserved for 6 hours and reperfused ex vivo for 2 hours to simulate transplantation or were transplanted in situ. During perfusion, group C showed an improved acid-base and biochemical environment in the circuit over group B. After reperfusion, the architecture of the liver grafts was best preserved in group C, followed by group B, then group A, as shown by the histology and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling staining of both hepatocytes and biliary epithelium. Ki-67 staining showed substantial hepatocyte proliferation and biliary epithelial regeneration after perfusion in group B and group C. Group C produced more bile in the reperfusion phase than those in group A and group B, with more physiological bile composition and less severe biliary epithelium injury. Von Willebrand factor-positive endothelial cells and E-selectin expression decreased in both group B and group C. Combined liver-kidney NMP not only produced more adenosine triphosphate, protected the nitric oxide signaling pathway, but also diminished oxidative stress (high mobility group box-1 protein and 8-hydroxy-2-deoxy guanosine levels) and inflammatory cytokine (IL6 and IL8) release when compared with liver-alone NMP and CS. In addition, the 7-day survival rate of liver transplant recipients was higher in group C than that in groups A and B. In conclusion, combined liver-kidney NMP can better protect DCD livers from warm ischemia and reperfusion injury probably by maintaining the stability of the internal environment and by abolishing oxidative stress injury. Liver Transplantation 24 67-79 2018 AASLD., (© 2017 by the American Association for the Study of Liver Diseases.)

Objectives: Biliary complications are common after living-donor liver transplant. This retrospective study reviewed our experience with biliary complications in recipients of living-donor liver transplant., Materials and Methods: Over our 9-year study period, 120 patients underwent living-donor liver transplant. Patients were divided into 2 groups, with group A having biliary complications and group B without biliary complications. Both groups were compared, and different treatment modalities for biliary complications were evaluated., Results: Group A included 45 patients (37.5%), whereas group B included 75 patients (62.5%). Biliary complications included bile leak in 17 patients (14.2%), biliary stricture in 11 patients (9.2%), combined biliary stricture with bile leak in 15 patients (12.5%), and sphincter of Oddi dysfunction and cholangitis in 1 patient each (0.8%). Cold ischemia time was significantly longer in group A (P = .002). External biliary drainage was less frequently used in group A (P = .031). Technical success rates of endoscopic biliary drainage and percutaneous transhepatic biliary drainage were 68.3% and 41.7%. Survival rate following relaparotomy for biliary complications was 62.5%., Conclusions: Graft ischemia is an important risk factor for biliary complications. Bile leaks can predispose to anastomotic strictures. The use of external biliary drainage seems to reduce the incidence of biliary complications. Endoscopic and percutaneous trans-hepatic approaches can successfully treat more than two-thirds of biliary complications. Relaparotomy can improve survival outcomes and is usually reserved for patients with intractable biliary complications.

Objectives: Our goal was to evaluate the predictors of coagulopathic hemorrhage after living-donor liver transplant., Materials and Methods: We retrospectively evaluated 161 patients who had undergone living-donor liver transplant from July 2005 to April 2014 at a single medical institution. Of these patients, 32 developed hemorrhage after transplant. Patients were separated into those with coagulopathy-related hemorrhage (n=15) or noncoagulopathy-related hemorrhage (n=17) based on the results of computed tomography images. Predictors of hemorrhage after living-donor liver transplant evaluated in this study included preoperative, perioperative, and posttransplant factors and hemodynamic status., Results: Patients who developed coagulopathy-related hemorrhage had significantly lower pretransplant platelet counts (P = .040), a longer cold-ischemia time (P = .045), more blood loss (P = .040), and earlier onset of hemorrhage (P = .048) than patients who had noncoagulopathy-related hemorrhage after transplant. Results of the generalized estimating equation analysis showed that heart rate and central venous pressure differed significantly between the 2 groups of patients. Heart rates increased significantly during hemorrhage (P < .010). Central venous pressure was higher in the coagulopathic group (P = .005) than in the noncoagulopathic group., Conclusions: Lower pretransplant platelet counts, longer cold ischemia time, more blood loss, earlier onset of hemorrhage, and higher central venous pressure level are indicators of coagulopathic hemorrhage after living-donor liver transplant.

This study aimed to elucidate the impact of epithelial regenerative responses and immune cell infiltration on biliary complications after liver transplantation. Bile duct (BD) damage after cold storage was quantified by a BD damage score and correlated with patient outcome in 41 patients. Bacterial infiltration was determined by fluorescence in situ hybridization (FISH). BD samples were analyzed by immunohistochemistry for E-cadherin, cytokeratin, CD56, CD14, CD4, CD8, and double-immunofluorescence for cytokine production and by messenger RNA (mRNA) microarray. Increased mRNA levels of adherens junctions (P < 0.01) were detected in damaged BDs from patients without complications compared with damaged BDs from patients with biliary complications. Immunohistochemistry showed increased expression of E-cadherin and cytokeratin in BDs without biliary complications (P = 0.03; P = 0.047). FISH analysis demonstrated translocation of bacteria in BDs. However, mRNA analysis suggested an enhanced immune response in BDs without biliary complications (P < 0.01). Regarding immune cell infiltration, CD4 + and CD8 + cells were significantly increased in patients without complications compared with those with complications (P = 0.02; P = 0.01). In conclusion, following BD damage during cold storage, we hypothesize that the functional regenerative capacity of biliary epithelium and enhanced local adaptive immune cell infiltration are crucial for BD recovery. Such molecular immunological BD analyses therefore could help to predict biliary complications in cases of "major" epithelial damage after cold storage.Liver Transplantation 23 1422-1432 2017 AASLD., (© 2017 by the American Association for the Study of Liver Diseases.)

Transplantation of liver grafts from donation after cardiac death (DCD) is limited. To identify barriers of DCD liver utilization, all active US liver transplant centers (n = 138) were surveyed, and the responses were compared with the United Network for Organ Sharing (UNOS) data. In total, 74 (54%) centers responded, and diversity in attitudes was observed, with many not using organ and/or recipient prognostic variables defined in prior studies and UNOS data analysis. Most centers (74%) believed lack of a system allowing a timely retransplant is a barrier to utilization. UNOS data demonstrated worse 1- and 5-year patient survival (PS) and graft survival (GS) in DCD (PS, 86% and 64%; GS, 82% and 59%, respectively) versus donation after brain death (DBD) recipients (PS, 90% and 71%; GS, 88% and 69%, respectively). Donor alanine aminotransferase (ALT), recipient Model for End-Stage Liver Disease (MELD), and cold ischemia time (CIT) significantly impacted DCD outcomes to a greater extent than DBD outcomes. At 3 years, relisting and retransplant rates were 7.9% and 4.6% higher in DCD recipients. To optimize outcome, our data support the use of DCD liver grafts with CIT <6-8 hours in patients with MELD ≤ 20. In conclusion, standardization of donor and recipient criteria, defining the impact of ischemic cholangiopathy, addressing donor hospital policies, and developing a strategy for timely retransplant may help to expand the use of these organs. Liver Transplantation 23 1372-1383 2017 AASLD., (© 2017 by the American Association for the Study of Liver Diseases.)

Aim: To compare the effect of University of Wisconsin (UW) solution with or without metformin, an AMP-activated protein kinase (AMPK) activator, for preserving standard and marginal liver grafts of young and aged rats ex vivo by hypothermic machine perfusion (HMP)., Methods: Eighteen young (4 mo old) and 18 aged (17 mo old) healthy male SD rats were selected and randomly divided into three groups: control group, UW solution perfusion group (UWP), and UW solution with metformin perfusion group (MUWP). Aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), interleukin-18 (IL-18), and tumor necrosis factor-alpha (TNF-α) in the perfused liquid were tested. The expression levels of AMPK and endothelial nitric oxide synthase (eNOS) in liver sinusoidal endothelial cells were also examined. Additionally, microscopic evaluation of the harvested perfused liver tissue samples was done., Results: AST, ALT, LDH, IL-18 and TNF-α levels in the young and aged liver-perfused liquid were, respectively, significantly lower in the MUWP group than in the UWP group ( P < 0.05), but no significant differences were found between the young and aged MUWP groups. Metformin increased the expression of AMPK and eNOS protein levels, and promoted the extracellular release of nitric oxide through activation of the AMPK-eNOS mediated pathway. Histological examination revealed that in the MUWP group, the extent of liver cells and tissue damage was significantly reduced compared with the UWP group., Conclusion: The addition of metformin to the UW preservative solution for ex vivo HMP can reduce rat liver injury during cold ischemia, with significant protective effects on livers, especially of aged rats., Competing Interests: Conflict-of-interest statement: All the authors declare no conflict of interest related to this publication.

Background: Hepatic artery thrombosis (HAT) is reported in 4%-15% of orthotopic liver transplants. Risk factors include technical error in the anastomosis, vascular anatomic variation, and high microvascular resistance. The aim of this study was to verify the incidence of HAT, early or late, and possible risk factors., Methods: This was a retrospective study from January 2007 to December 2012 at the State University of Campinas. Variables analyzed were age, sex, cold and warm ischemia times, underlying disease, presence of hepatocellular carcinoma, Model for End-Stage Liver Disease (MELD) score, arterial anatomic variation in the graft, cytomegalovirus (CMV) infection, rejection, biliary complications, retransplantation rate, and survival., Results: The incidence of HAT was 21/263, or 7.9%. Pure average MELD score was 22 ± 7.4. There was vascular anatomic variation in the graft in 14.2% of cases, in the majority (66.6%) a right hepatic artery from the superior mesenteric artery, and 4.76% of patients had CMV infection and acute cellular rejection (1 case each). There were biliary complications in 38% of patients, 13.3% of cases in patients with early HAT, and 100% of patients with late HAT (P = .002). Body mass index in late HAT was higher (P = .01)., Conclusions: Late HAT was related to a significant increase in biliary complications (stenosis), and the survival rate was similar at 5 years., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Background: Liver transplantation (LT) is a curative treatment option for hepatocellular carcinoma (HCC); recurrent HCC after liver transplantation (HCC-R) is diagnosed in 9%-16%. The objective of this study was to evaluate which factors are associated with R-HCC after liver transplantation., Methods: This retrospective real-life study analyzed 278 LTs from 3 reference centers (2,093 LTs) in Brazil from 1988 to 2015. HCC-R with histologic confirmation was seen in 40 patients (14.4%)., Results: Most of them were male with cirrhosis secondary to viral hepatitis. Only 37.5% underwent chemoembolization, and 50% had cold ischemia time >8 hours. From the explant analysis, most of the patients were outside Milan criteria and 37.5% had microvascular invasion. The donors were mostly male, and the median intensive care unit time was >3 days. The Kaplan-Meier survival was lower according to alpha-fetoprotein (AFP) >200 ng/dL (P = .02), and older donors and more blood transfusions were risk factors for HCC-R death., Conclusion: AFP >200 ng/mL was associated with lower survival, and older donors and more blood transfusions were risk factors for death after HCC-R. A trend to lower survival was observed in patients who did not have chemoembolization and had cold ischemia times >8 hours., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The use of liver grafts from donation after cardiac death (DCD) has been limited due to the increased rate of graft failure, mostly related to ischemic cholangiopathy (IC). It is our hypothesis that longterm outcomes and quality of life (QOL) similar to patients undergoing liver transplantation (LT) with donation after brain death (DBD) can be achieved. Clinical outcomes of all patients undergoing DCD LT (n = 300) between 1998 and 2015 were compared with a propensity score-matched cohort of patients undergoing DBD LT (n = 300). Patients were contacted for a follow-up questionnaire and short-form (SF)-12 QOL Survey administration. Median follow-up was >5 years. Graft survival at 1-, 3-, and 5-years was 83.8%, 75.5%, and 70.1% in the DCD LT group and 88.4%, 80.3%, and 73.9% in the DBD LT group (P = 0.27). Patient survival at 1-, 3-, and 5-years was 92.3%, 86.1%, and 80.3% in the DCD LT group and 92.3%, 85.1%, and 79.5% in the DBD LT group (P = 0.81). IC developed in 11.7% and 2% of patients in the DCD LT group and DBD LT group, respectively (P < 0.001). DCD LT recipients who developed IC had inferior graft survival compared with both the DCD non-IC group (P < 0.001) and the DBD LT group (P < 0.001); no difference in graft survival was observed between the DCD non-IC group and the DBD LT group (P = 0.50). Physical and Mental Composite Scores on the SF-12 QOL questionnaire were similar between the DCD LT and DBD LT groups (44.0 versus 45.4; P = 0.34 and 51.9 versus 52.2; P = 0.83), respectively. Similar longterm survival and QOL scores can be achieved between DCD LT and DBD LT. Prevention of IC in DCD LT yields excellent graft and patient survival with virtually no difference compared with DBD LT. Liver Transplantation 23 342-351 2017 AASLD., (© 2016 by the American Association for the Study of Liver Diseases.)

Liver transplantation (LT) offers the best chance of survival in selected patients with hepatocellular carcinoma (HCC). Wait-list mortality or dropout due to tumor progression can be significant, and therefore, timely transplantation is critical. Liver grafts discarded by outside organ procurement organizations are a potential source of grafts for low Model for End-Stage Liver Disease tumor patients. The primary aim of this study was to assess the disease-free and overall survival of patients with HCC transplanted with imported liver grafts (ILGs). Review of all patients transplanted for HCC between June 2005 and December 2014 was performed. Data on demographics, survival, and HCC recurrence were analyzed. During this time period, 59 out of 190 (31%) recipients with HCC received ILG. Of these 59 grafts, 54 were imported from within the region and 5 were from national offers (outside the region). The mean cold ischemia time for local liver grafts (LLGs) was 4.1 ± 1.5 hours versus 5.1 ± 1.4 hours for ILG (P < 0.001). The 1-, 3-, and 5-year patient survival was 90%, 85%, and 83% and 85%, 80%, and 79% for LLG and ILG (P = 0.08), respectively. The observed disease recurrence rate for both LLG and ILG recipients was equivalent. The median wait-list time for HCC recipients was 43 days (range, 2-1167 days). In conclusion, with careful graft assessment, the use of ILGs results in comparable outcomes following LT and no increased risk of HCC recurrence. Use of ILGs maximizes the donor pool and results in a higher rate of transplantation for HCC recipients. Liver Transplantation 23 299-304 2017 AASLD., (© 2016 by the American Association for the Study of Liver Diseases.)

Background: Ex situ normothermic machine perfusion (NMP) can be performed after traditional static cold preservation to assess graft function and viability before transplantation. It is unknown whether this results in activation of coagulation and fibrinolysis, as may occur upon graft reperfusion in vivo., Methods: Twelve donor livers declined for transplantation underwent 6 hours of end-ischemic NMP using a heparinized plasma-based perfusion fluid. Concentration of prothrombin fragment F1 + 2 (marker of coagulation activation), D-dimer, plasmin-antiplasmin complex, tissue plasminogen activator and plasminogen activator inhibitor-1 (markers for fibrinolysis) and alanine aminotransferase (ALT) (marker of ischemia-reperfusion [I/R] injury) were measured in perfusion fluid at regular intervals. Liver biopsies were examined for the presence of fibrin, using light microscopy after Maurits, Scarlet and Blue staining., Results: No significant increase in prothrombin F1 + 2 was noted during NMP. D-dimer and plasmin-antiplasmin complex levels increased soon after start of NMP and D-dimer concentrations correlated significantly with levels of tissue plasminogen activator. In livers displaying good function during NMP, perfusate levels of ALT and D-dimers were low (≤3500 ng/mL), whereas significantly higher D-dimer levels (>3500 ng/mL) were in found in livers with poor graft function. Activation of fibrinolysis correlated significantly with the degree of I/R injury, as reflected by ALT levels., Conclusions: End-ischemic ex situ NMP results in activation of fibrinolysis, but not of coagulation. Markers of fibrinolysis activation correlate significantly with markers of I/R injury. High concentrations of D-dimer early after start of NMP can be considered a marker of severe I/R injury and a predictor of poor liver graft function.

Normothermic ex vivo liver perfusion (NEVLP) improves graft preservation by avoiding cold ischemia injury. We investigated whether the protective effects of NEVLP can be further improved by applying strategies targeted on reducing the activation of proinflammatory cytokines during perfusion. Livers retrieved under heart-beating conditions were perfused for 4 hours. Following the preservation period, a pig liver transplantation was performed. In group 1 (n = 5), anti-inflammatory strategies (alprostadil, n-acetylcysteine, carbon monoxide, sevoflurane, and subnormothermic temperature [33°C]) were applied. This was compared with a perfused control group (group 2) where livers (n = 5) were perfused at 37°C without anti-inflammatory agents, similar to the setup used in current European clinical trials, and to a control group preserved with static cold storage (group 3). During 3-day follow-up, markers of reperfusion injury, bile duct injury, and liver function were examined. Aspartate aminotransferase (AST) levels during perfusion were significantly lower in the study versus control group at 1 hour (52 ± 6 versus 162 ± 86 U/L; P = 0.01), 2 hours (43 ± 5 versus 191 ± 111 U/L; P = 0.008), and 3 hours (24 ± 16 versus 218 ± 121 U/L; P = 0.009). During perfusion, group 1 versus group 2 had reduced interleukin (IL) 6, tumor necrosis factor α, and galactosidase levels and increased IL10 levels. After transplantation, group 1 had lower AST peak levels compared with group 2 and group 3 (1400 ± 653 versus 2097 ± 1071 versus 1747 ± 842 U/L; P = 0.47) without reaching significance. Bilirubin levels were significantly lower in group 1 versus group 2 at day 1 (3.6 ± 1.5 versus 6.60 ± 1.5 μmol/L; P = 0.02) and 3 (2 ± 1.1 versus 9.7 ± 7.6 μmol/L; P = 0.01). A trend toward decreased hyaluronic acid, as a marker of improved endothelial cell function, was observed at 1, 3, and 5 hours after reperfusion in group 1 versus group 2. Only 1 early death occurred in each group (80% survival). In conclusion, addition of anti-inflammatory strategies further improves warm perfused preservation. Liver Transplantation 22 1573-1583 2016 AASLD., (© 2016 by the American Association for the Study of Liver Diseases.)