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Direct-acting antivirals have proved to be highly efficacious and safe in monoinfected liver transplant (LT) recipients who experience recurrence of hepatitis C virus (HCV) infection. However, there is a lack of data on effectiveness and tolerability of these regimens in HCV/HIV-coinfected patients who experience recurrence of HCV infection after LT. In this prospective, multicenter cohort study, the outcomes of 47 HCV/HIV-coinfected LT patients who received DAA therapy (with or without ribavirin [RBV]) were compared with those of a matched cohort of 148 HCV-monoinfected LT recipients who received similar treatment. Baseline characteristics were similar in both groups. HCV/HIV-coinfected patients had a median (IQR) CD4 T-cell count of 366 (256-467) cells/µL. HIV-RNA was <50 copies/mL in 96% of patients. The DAA regimens administered were SOF + LDV ± RBV (34%), SOF + SMV ± RBV (31%), SOF + DCV ± RBV (27%), SMV + DCV ± RBV (5%), and 3D (3%), with no differences between the groups. Treatment was well tolerated in both groups. Rates of SVR (negative serum HCV-RNA at 12 weeks after the end of treatment) were high and similar for coinfected and monoinfected patients (95% and 94%, respectively; P = .239). Albeit not significant, a trend toward lower SVR rates among patients with advanced fibrosis (P = .093) and genotype 4 (P = .088) was observed. In conclusion, interferon-free regimens with DAAs for post-LT recurrence of HCV infection in HIV-infected individuals were highly effective and well tolerated, with results comparable to those of HCV-monoinfected patients., (© 2018 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Background & Aims: Subclinical inflammatory changes are commonly described in long-term transplant recipients undergoing protocol liver biopsies. The pathogenesis of these lesions remains unclear. The aim of this study was to identify the key molecular pathways driving progressive subclinical inflammatory liver allograft damage., Methods: All liver recipients followed at Hospital Clínic Barcelona who were >10 years post-transplant were screened for participation in the study. Patients with recurrence of underlying liver disease, biliary or vascular complications, chronic rejection, and abnormal liver function tests were excluded. Sixty-seven patients agreed to participate and underwent blood and serological tests, transient elastography and a liver biopsy. Transcriptome profiling was performed on RNA extracted from 49 out of the 67 biopsies employing a whole genome next generation sequencing platform. Patients were followed for a median of 6.8 years following the index liver biopsy., Results: Median time since transplantation to liver biopsy was 13 years (10-22). The most frequently observed histological abnormality was portal inflammation with different degrees of fibrosis, present in 45 biopsies (67%). Two modules of 102 and 425 co-expressed genes were significantly correlated with portal inflammation, interface hepatitis and portal fibrosis. These modules were enriched in molecular pathways known to be associated with T cell mediated rejection. Liver allografts showing the highest expression levels for the two modules recapitulated the transcriptional profile of biopsies with clinically apparent rejection and developed progressive damage over time, as assessed by non-invasive markers of fibrosis., Conclusions: A large proportion of adult liver transplant recipients who survive long-term exhibit subclinical histological abnormalities. The transcriptomic profile of these patients' liver tissue closely resembles that of T cell mediated rejection and may result in progressive allograft damage., Lay Summary: A large proportion of adult liver transplant recipients who survive for a long time exhibit subclinical histological abnormalities. The expression profile (a measurement of the activity of genes) of liver tissue from a large fraction of these patients closely resembles the profile of T cell mediated rejection. Liver allografts showing the highest expression levels of rejection-related genes developed progressive damage over time., (Copyright © 2018 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Sustained virological response (SVR) improves survival in post-liver transplant (LT) recurrent hepatitis C. However, the impact of SVR on fibrosis regression is not well defined. In addition, the performance of noninvasive methods to evaluate the presence of fibrosis and portal hypertension (PH) post-SVR has been scarcely evaluated. We aimed to investigate the degree of fibrosis regression (decrease ≥1 METAVIR stage) after-SVR and its associated factors in recurrent hepatitis C, as well as the diagnostic capacity of noninvasive methods in the assessment of liver fibrosis and PH after viral clearance. We evaluated 112 hepatitis C virus-infected LT recipients who achieved SVR between 2001 and 2015. A liver biopsy was performed before treatment and 12 months post-SVR. Hepatic venous pressure gradient (HVPG), liver stiffness measurement (LSM), and Enhanced Liver Fibrosis (ELF) score were also determined at the same time points. Sixty-seven percent of the cohort presented fibrosis regression: 43% in recipients with cirrhosis and 72%-85% in the remaining stages (P = 0.002). HVPG, LSM, and ELF significantly decreased post-SVR. Liver function significantly improved, and survival was significantly better in patients achieving fibrosis regression. Baseline HVPG and LSM as well as decompensations before therapy were independent predictors of fibrosis regression. One year post-SVR, LSM had a high diagnostic accuracy to discard the presence of advanced fibrosis (AF) and clinically significant PH (AUROC, 0.902 and 0.888)., Conclusion: In conclusion, SVR post-LT induces fibrosis regression in most patients, leading to significant clinical benefits. Pretreatment HVPG and LSM are significant determinants of the likelihood of fibrosis regression. Finally, LSM accurately predicts the presence of AF and PH 1 year after SVR and thus can be used to determine monitoring strategies. (Hepatology 2018;67:1683-1694)., (© 2017 by the American Association for the Study of Liver Diseases.)

There is a lack of data on incidental hepatocellular carcinoma (iHCC) in the setting of liver transplantation (LT) in human immunodeficiency virus (HIV)-infected patients. This study aims to describe the frequency, histopathological characteristics, and outcomes of HIV+ LT recipients with iHCC from a Spanish multicenter cohort in comparison with a matched cohort of LT patients without HIV infection. A total of 15 (6%) out of 271 patients with HIV infection who received LT in Spain from 2002 to 2012 and 38 (5%) out of the 811 HIV- counterparts presented iHCC in liver explants (P = 0.58). Patients with iHCC constitute the present study population. All patients also had hepatitis C virus (HCV)-related cirrhosis. There were no significant differences in histopathological features of iHCC between the 2 groups. Most patients showed a small number and size of tumoral nodules, and few patients had satellite nodules, microvascular invasion, or poorly differentiated tumors. After a median follow-up of 49 months, no patient developed hepatocellular carcinoma (HCC) recurrence after LT. HIV+ LT recipients tended to have lower survival than their HIV- counterparts at 1 (73% versus 92%), 3 (67% versus 84%), and 5 years (50% versus 80%; P = 0.06). There was also a trend to a higher frequency of HCV recurrence as a cause of death in the former (33% versus 10%; P = 0.097). In conclusion, among LT recipients for HCV-related cirrhosis, the incidence and histopathological features of iHCC in HIV+ and HIV- patients were similar. However, post-LT survival was lower in HIV+ patients probably because of a more aggressive HCV recurrence. Liver Transplantation 23 645-651 2017 AASLD., (© 2017 by the American Association for the Study of Liver Diseases.)

Objectives: IFN-based therapy against hepatitis C recurrence after liver transplantation (LT) has poor effectiveness and tolerability. In HIV/HCV-coinfected liver transplant recipients, the results are even poorer. Here, we report our experience using direct antiviral agents (DAAs) in 11 consecutive coinfected patients within the LT setting., Methods: Four patients with compensated cirrhosis and hepatocellular carcinoma were treated while awaiting LT and seven patients received antiviral therapy due to severe hepatitis C recurrence after LT [fibrosing cholestatic hepatitis (n = 1), fibrosis stage ≥F3 (n = 2) and decompensated cirrhosis (n = 4)]. Patients were treated with different sofosbuvir-based regimens with or without ribavirin for 12 or 24 weeks., Results: Sustained virological response (SVR) was achieved in all patients. Two of the four patients treated while awaiting LT reached the time of transplant with undetectable HCV-RNA that remained undetectable 12 weeks after LT, one patient had detectable HCV-RNA at the time of transplant but achieved SVR after completing 12 weeks of therapy after LT and the last patient is still on the waiting list. Seven patients with severe post-LT hepatitis C recurrence were treated within 11-120 months after LT. In these patients, viral eradication was associated with an improvement in liver function and clinical decompensation. Tolerance to antiviral therapy was good and only four patients reported mild adverse events., Conclusions: IFN-free regimens are effective and well tolerated in HIV/HCV-coinfected patients within the LT setting, but more data are needed to confirm our promising results and to establish the best treatment option in this population., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

The mayor goal still outstanding into the solid organ transplantation field involves the search of surrogate biomarkers able to predict several clinical events, such as acute rejection (AR) or opportunistic infection. In the present multicenter study, a series of interesting surface antigens with important activator or inhibitory immune functions on cultured peripheral T cells were monitored in liver transplant recipients drawn at baseline and up to one year after transplantation. Sixty-four patients were included in the multicenter study during 3 years. Pre- and post-transplantation surface antigens levels displayed significant differences between AR and non acute rejection (NAR) groups, and also this differential expression was used to construct a risk predictive model based on a composite panel of outcome biomarkers (CD38, CD69, CD95 and CD154). The model was able to stratify these patients at high risk of AR. These preliminary results could provide basic information to improve the immunosuppressive treatment and it might better help to predict AR episodes., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

Several studies have analyzed the potential of T regulatory cells (Treg cells) as biomarkers of acute rejection (AR). The aim of the present multicenter study was to correlate the percentage of peripheral Treg cells in liver graft recipients drawn at baseline up to 12 months after transplantation with the presence of AR. The percentage of central memory (cm) Treg cells (CD4(+)CD25(high)CD45RO(+)CD62L(+)) was monitored at pre-transplant and at 1 and 2 weeks, and 1, 2, 3 and 6 months and 1 year post-transplantation. The same validation standard operating procedures were used in all participating centers. Fifteen patients developed AR (23.4%). Hepatitis C virus recurrence was observed in 16 recipients, who displayed low peripheral blood cmTreg levels compared with patients who did not. A steady increase of cmTregs was observed during the first month after transplantation with statistically significant differences between AR and non-AR patients. The high frequency of memory Treg cells allowed us to monitor rejection episodes during the first month post-transplantation. On the basis of these data, we developed a prediction model for assessing risk of AR that can provide clinicians with useful information for managing patients individually and customizing immunosuppressive therapies., (© The Japanese Society for Immunology. 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Unlabelled: The impact of human immunodeficiency virus (HIV) infection on patients undergoing liver transplantation (LT) for hepatocellular carcinoma (HCC) is uncertain. This study aimed to assess the outcome of a prospective Spanish nationwide cohort of HIV-infected patients undergoing LT for HCC (2002-2014). These patients were matched (age, gender, year of LT, center, and hepatitis C virus (HCV) or hepatitis B virus infection) with non-HIV-infected controls (1:3 ratio). Patients with incidental HCC were excluded. Seventy-four HIV-infected patients and 222 non-HIV-infected patients were included. All patients had cirrhosis, mostly due to HCV infection (92%). HIV-infected patients were younger (47 versus 51 years) and had undetectable HCV RNA at LT (19% versus 9%) more frequently than non-HIV-infected patients. No significant differences were detected between HIV-infected and non-HIV-infected recipients in the radiological characteristics of HCC at enlisting or in the histopathological findings for HCC in the explanted liver. Survival at 1, 3, and 5 years for HIV-infected versus non-HIV-infected patients was 88% versus 90%, 78% versus 78%, and 67% versus 73% (P = 0.779), respectively. HCV infection (hazard ratio = 7.90, 95% confidence interval 1.07-56.82) and maximum nodule diameter >3 cm in the explanted liver (hazard ratio = 1.72, 95% confidence interval 1.02-2.89) were independently associated with mortality in the whole series. HCC recurred in 12 HIV-infected patients (16%) and 32 non-HIV-infected patients (14%), with a probability of 4% versus 5% at 1 year, 18% versus 12% at 3 years, and 20% versus 19% at 5 years (P = 0.904). Microscopic vascular invasion (hazard ratio = 3.40, 95% confidence interval 1.34-8.64) was the only factor independently associated with HCC recurrence., Conclusions: HIV infection had no impact on recurrence of HCC or survival after LT. Our results support the indication of LT in HIV-infected patients with HCC., (© 2015 by the American Association for the Study of Liver Diseases.)

Liver transplantation is increasingly performed in selected HIV-infected patients in most developed countries, with excellent results reported in patients with liver diseases unrelated to HCV. In contrast, survival in HCV/HIV-coinfected liver recipients is poorer than in HCV-monoinfected patients, due to more aggressive recurrence of HCV and consequent graft loss and death. Results from American, French, and Spanish cohort studies showed a 5-year survival rate of only 50-55%. Therefore, it is debated whether liver transplantation should be offered to HCV/HIV-coinfected patients. Studies have shown that the variables more consistently associated with poor outcome are: (1) the use of old or HCV-positive donors, (2) dual liver-kidney transplantation, (3) recipients with very low body mass index and (4) less site experience. However, the most effective factor influencing transplantation outcome is the successful treatment of HCV recurrence with anti-HCV. Survival is 80% in patients whose HCV infection resolves. Unfortunately, the rates of sustained virological response with pegylated-interferon plus ribavirin in coinfected recipients are low, particularly for genotype 1 (only 10%). Here we present a non-systematic review of the literature based on our own experience in different liver transplant scenarios. This review covers selection criteria in HIV-infected patients, pre- and post-LT management, donor selection, anti-HCV treatment, drug interactions with antiretrovirals and anti-HCV direct antiviral agents, hepatocellular carcinoma, and liver retransplantation. Recommendations are rated. Finally, we explain how the introduction of new effective and more tolerable direct antiviral agents may improve significantly the outcome of HCV/HIV-coinfected liver recipients., (Copyright © 2014 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Ischemia reperfusion injury (IRI) leads to significant tissue damage in liver surgery. Polyethylene glycols (PEGs) are water soluble nontoxic polymers that have proved their effectiveness against IRI. The objective of our study was to investigate the potential protective effects of intravenous administration of a high molecular weight PEG of 35 kDa (PEG 35) in steatotic livers subjected to cold ischemia reperfusion. In this study, we used isolated perfused rat liver model to assess the effects of PEG 35 intravenous administration after prolonged cold ischemia (24 h, 4°C) and after reperfusion (2 h, 37°C). Liver injury was measured by transaminases levels and mitochondrial damage was determined by confocal microscopy assessing mitochondrial polarization (after cold storage) and by measuring glutamate dehydrogenase activity (after reperfusion). Also, cell signaling pathways involved in the physiopathology of IRI were assessed by western blot technique. Our results show that intravenous administration of PEG 35 at 10 mg/kg ameliorated liver injury and protected the mitochondria. Moreover, PEG 35 administration induced a significant phosphorylation of prosurvival protein kinase B (Akt) and activation of cytoprotective factors e-NOS and AMPK. In conclusion, intravenous PEG 35 efficiently protects steatotic livers exposed to cold IRI.

Background & Aims: The aim of this study was to evaluate the results of treatment with pegylated interferon and ribavirin for the recurrence of hepatitis C after liver transplantation in HCV/HIV-coinfected patients., Methods: This was a prospective, multicentre cohort study, including 78 HCV/HIV-coinfected liver transplant patients who received treatment for recurrent hepatitis C. For comparison, we included 176 matched HCV-monoinfected patients who underwent liver transplantation during the same period of time at the same centres and were treated for recurrent hepatitis C., Results: Antiviral therapy was discontinued prematurely in 56% and 39% (p = 0.016), mainly because of toxicity (22% and 11%, respectively; p=0.034). Sustained virological response (SVR) was achieved in 21% of the coinfected patients and in 36% of monoinfected patients (p = 0.013). For genotype 1, SVR rates were 10% and 33% (p = 0.002), respectively; no significant differences were observed for the other genotypes. A multivariate analysis based on the whole series identified HIV-coinfection as an independent predictor of lack of SVR (OR, 0.17; 95% CI, 0.06-0.42). Other predictors of SVR were donor age, pretreatment HCV viral load, HCV genotype, and early virological response. SVR was associated with a significant improvement in survival: 5-year survival after antiviral treatment was 79% for HCV/HIV-coinfected patients with SVR vs. 43% for those without (p = 0.02) and 92% vs. 60% in HCV-monoinfected patients (p < 0.001), respectively., Conclusions: The response to pegylated interferon and ribavirin was poorer in HCV/HIV-coinfected liver recipients, particularly those with genotype 1. However, when SVR was achieved, survival of coinfected patients increased significantly., (Copyright © 2014 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Aim: To test whether a new rinse solution containing polyethylene glycol 35 (PEG-35) could prevent ischemia-reperfusion injury (IRI) in liver grafts., Methods: Sprague-Dawley rat livers were stored in University of Wisconsin preservation solution and then washed with different rinse solutions (Ringer's lactate solution and a new rinse solution enriched with PEG-35 at either 1 or 5 g/L) before ex vivo perfusion with Krebs-Heinseleit buffer solution. We assessed the following: liver injury (transaminase levels), mitochondrial damage (glutamate dehydrogenase activity), liver function (bile output and vascular resistance), oxidative stress (malondialdehyde), nitric oxide, liver autophagy (Beclin-1 and LCB3) and cytoskeleton integrity (filament and globular actin fraction); as well as levels of metalloproteinases (MMP2 and MMP9), adenosine monophosphate-activated protein kinase (AMPK), heat shock protein 70 (HSP70) and heme oxygenase 1 (HO-1)., Results: When we used the PEG-35 rinse solution, reduced hepatic injury and improved liver function were noted after reperfusion. The PEG-35 rinse solution prevented oxidative stress, mitochondrial damage, and liver autophagy. Further, it increased the expression of cytoprotective heat shock proteins such as HO-1 and HSP70, activated AMPK, and contributed to the restoration of cytoskeleton integrity after IRI., Conclusion: Using the rinse solution containing PEG-35 was effective for decreasing liver graft vulnerability to IRI.

Renal failure is a frequent complication in liver transplant recipients and is associated with increased morbidity and mortality. A variety of risk factors for the development of renal failure in the pre- and post-transplantation periods have been described, as well as at the time of surgery. To reduce the negative impact of renal failure in this population, an active approach is required for the identification of those patients with risk factors, the implementation of preventive strategies, and the early detection of progressive deterioration of renal function. Based on published evidence and on clinical experience, this document presents a series of recommendations on monitoring RF in LT recipients, as well as on the prevention and management of acute and chronic renal failure after LT and referral of these patients to the nephrologist. In addition, this document also provides an update of the various immunosuppressive regimens tested in this population for the prevention and control of post-transplantation deterioration of renal function., (Copyright © 2013 Elsevier España, S.L.U. and AEEH y AEG. All rights reserved.)

Pathogen-induced immune responses prevent the establishment of transplantation tolerance in experimental animal models. Whether this occurs in humans as well remains unclear. The development of operational tolerance in liver transplant recipients with chronic hepatitis C virus (HCV) infection allows us to address this question. We conducted a clinical trial of immunosuppression withdrawal in HCV-infected adult liver recipients to elucidate (i) the mechanisms through which allograft tolerance can be established in the presence of an ongoing inflammatory response and (ii) whether anti-HCV heterologous immune responses influence this phenomenon. Of 34 enrolled liver recipients, drug withdrawal was successful in 17 patients (50%). Tolerance was associated with intrahepatic overexpression of type I interferon and immunoregulatory genes and with an expansion of exhausted PD1/CTLA4/2B4-positive HCV-specific circulating CD8(+) T cells. These findings were already present before immunosuppression was discontinued and were specific for HCV infection. In contrast, the magnitude of HCV-induced proinflammatory gene expression and the breadth of anti-HCV effector T cell responses did not influence drug withdrawal outcome. Our data suggest that in humans, persistent viral infections exert immunoregulatory effects that could contribute to the restraining of alloimmune responses, and do not necessarily preclude the development of allograft tolerance., (Copyright © 2014, American Association for the Advancement of Science.)

Ischemia-reperfusion (IR) injury is an important problem in liver surgery especially when steatosis is present. Ischemic preconditioning (PC) is the only surgical strategy that has been applied in patients with steatotic livers undergoing warm ischemia. Silent information regulator 1 (SIRT1) is a histone deacetylase that regulates various cellular processes. This study evaluates the SIRT1 implication in PC in fatty livers. Homozygous (Ob) Zucker rats were subjected to IR and IR + PC. An additional group treated with sirtinol or EX527 (SIRT1 inhibitors) before PC was also realized. Liver injury and oxidative stress were evaluated. SIRT1 protein levels and activity, as well as other parameters involved in PC protective mechanisms (adenosine monophosphate protein kinase, eNOS, HSP70, MAP kinases, apoptosis), were also measured. We demonstrated that the protective effect of PC was due in part to SIRT1 induction, as SIRT1 inhibition resulted in increased liver injury and abolished the beneficial mechanisms of PC. In this study, we report for the first time that SIRT1 is involved in the protective mechanisms induced by hepatic PC in steatotic livers., (© 2014 Steunstichting ESOT.)

Transplant recipients receiving immunosuppressive therapy are at increased risk of active cytomegalovirus (CMV) infection and disease. Without appropriate prophylaxis, as many as 80% of solid organ transplant recipients may experience CMV infection. In addition to the direct effects of CMV, infection may be associated with a range of indirect effects, including an increase in risk of other infections, as well as a higher incidence of rejection, graft loss and death. The indirect effects of CMV infection can vary depending on the transplanted organ. For example, CMV-infected kidney transplant recipients may be at increased risk of cardiovascular disease and diabetes, while CMV infection in liver transplant recipients may potentiate hepatitis C infection and increase the risk of post-transplant lymphoproliferative disease. Indirect effects result from a number of pathological processes, including immune modulation and immunosuppression, generation of cytotoxic, pro-inflammatory responses, and smooth muscle proliferation. Prophylactic treatment with antiviral medication can reduce the risk of CMV disease, thereby improving graft survival and overall outcomes, particularly in kidney and heart transplant recipients. Antiviral prophylaxis should be considered for all patients at risk of CMV infection after solid organ transplantation. In this paper we review the main indirect effects of CMV infection in solid organ transplant recipients, and the impact of CMV prophylaxis on these effects., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Objectives: The aim of this study is to investigate the protective mechanisms induced by bortezomib added to Institut George Lopez (IGL)-1 preservation solution to protect steatotic livers against cold ischaemia reperfusion injury and to examine whether these mechanisms occur through the activation of adenosine monophosphate activated protein kinase (AMPK), Akt/mTOR pathways., Methods: Steatotic livers from obese rats were preserved for 24 h (at 4 °C) in IGL-1 solution with or without bortezomib (100 nM) or pretreated with AMPK inhibitor adenine 9-α-D-arabinofuranoside and preserved in IGL-1 + bortezomib. Livers were then perfused for 2 h at 37 °C. Liver injury (alanine aminotransferase/aspartate aminotransferase) and function (bile production and vascular resistance) were measured. Also, Akt/mTOR, phosphorylated AMPK (pAMPK) and apoptosis were determined by Western blot analyses., Key Findings: Bortezomib addition to IGL-1 solution significantly reduced steatotic liver injury, improved graft function and decreased liver apoptosis. These benefits were diminished by the pretreatment of obese rats with AMPK inhibitor Ara. Western blot analyses showed a significant increase in pAMPK after ischaemia and reperfusion. We also observed a significant phosphorylation of Akt in IGL-1 +bortezomib group that, in turn, induced the phosphorylation of mTOR and glycogen synthase kinase 3β., Conclusions: Bortezomib, at low and non toxic concentration, is a promising additive to IGL-1 solution for steatotic liver preservation. Its protective effect is due to the activation of AMPK and Akt/mTOR pathways., (© 2013 Royal Pharmaceutical Society.)

Unlabelled: Lifelong immunosuppression increases morbidity and mortality in liver transplantation. Discontinuation of immunosuppressive drugs could lessen this burden, but the safety, applicability, and clinical outcomes of this strategy need to be carefully defined. We enrolled 102 stable liver recipients at least 3 years after transplantation in a single-arm multicenter immunosuppression withdrawal trial. Drugs were gradually discontinued over a 6 to 9-month period. The primary endpoint was the development of operational tolerance, defined as successful immunosuppressive drug cessation maintained for at least 12 months with stable graft function and no histopathologic evidence of rejection. Out of the 98 recipients evaluated, 57 rejected and 41 successfully discontinued all immunosuppressive drugs. In nontolerant recipients rejection episodes were mild and resolved over 5.6 months (two nontolerant patients still exhibited mild gradually improving cholestasis at the end of follow-up). In tolerant recipients no progressive clinically significant histological damage was apparent in follow-up protocol biopsies performed up to 3 years following drug withdrawal. Tolerance was independently associated with time since transplantation (odds ratio [OR] 1.353; P = 0.0001), recipient age (OR 1.073; P = 0.009), and male gender (OR 4.657; P = 0.016). A predictive model incorporating the first two clinical variables identified subgroups of recipients with very high (79%), intermediate (30%-38%), and very low (0%) likelihood of successful withdrawal., Conclusion: When conducted at late timepoints after transplantation, immunosuppression withdrawal is successful in a high proportion of carefully selected liver recipients. A combination of clinical parameters could be useful to predict the success of this strategy. Additional prospective studies are now needed to confirm these results and to validate clinically applicable diagnostic biomarkers., (© 2013 by the American Association for the Study of Liver Diseases.)

Despite the increase in long-term survival, liver transplant recipients still exhibit higher morbidity and mortality than the general population. This is in part attributed to the lifelong administration of immunosuppression and its associated side effects. Several studies reported in the last decades have evaluated the impact of immunosuppression minimization in liver transplant recipients, but results have been inconsistent due to the heterogeneity of study designs and insufficient sample sizes. On the other hand, complete immunosuppression withdrawal has proven to be feasible in approximately 20% of carefully selected liver transplant recipients, especially in older patients and those with longer duration after transplantation. The long-term risks and clinical benefits of this strategy, however, also need to be clarified. As a consequence, and despite the general perception that a large proportion of liver recipients are over-immunosuppressed, it is currently not possible to derive evidence-based guidelines on how to manage long-term immunosuppression to improve clinical outcomes. Large clinical trials of drug minimization and/or withdrawal focused on clinically-relevant long-term outcomes are required. Development of personalized medicine tools and a deeper understanding of the pathogenesis of idiopathic inflammatory graft lesions will be pre-requisites to achieve these goals., (Copyright © 2013. Published by Elsevier B.V.)

Background: The dramatic shortage of organs leads to consider the steatotic livers for transplantation although their poor tolerance against ischemia reperfusion injury (IRI). Ubiquitin proteasome system (UPS) inhibition during hypothermia prolongs myocardial graft preservation. The role of UPS in the liver IRI is not fully understood. Bortezomib (BRZ) treatment at non-toxic doses of rats fed alcohol chronically has shown protective effects by increasing liver antioxidant enzymes. We evaluated and compared both proteasome inhibitors BRZ and MG132 in addition to University of Wisconsin preservation solution (UW) at low and non-toxic dose for fatty liver graft protection against cold IRI., Experimental: Steatotic and non-steatotic livers have been stored in UW enriched with BRZ (100 nM) or MG132 (25 μM), for 24h at 4°C and then subjected to 2-h normothermic reperfusion (37 °C). Liver injury (AST/ALT), hepatic function (bile output; vascular resistance), mitochondrial damage (GLDH), oxidative stress (MDA), nitric oxide (NO) (e-NOS activity; nitrates/nitrites), proteasome chymotrypsin-like activity (ChT), and UPS (19S and 20S5 beta) protein levels have been measured., Results: ChT was inhibited when BRZ and MG132 were added to UW. Both inhibitors prevented liver injury (AST/ALT), when compared to UW alone. BRZ increased bile production more efficiently than MG132. Only BRZ decreased vascular resistance in fatty livers, which correlated with an increase in NO generation (through e-NOS activation) and AMPK phosphorylation. GLDH and MDA were also prevented by BRZ. In addition, BRZ inhibited adiponectin, IL-1, and TNF alpha, only in steatotic livers., Conclusion: MG132 and BRZ, administrated at low and non toxic doses, are very efficient to protect fatty liver grafts against cold IRI. The benefits of BRZ are more effective than those of MG132. This evidenced for the first time the potential use of UPS inhibitors for the preservation of marginal liver grafts and for future applications in the prevention of IRI., (Copyright © 2013 Elsevier Inc. All rights reserved.)

In this descriptive study, we examined the role of single-operator cholangioscopy (SOC) in the evaluation of biliary complications after liver transplantation (LT). We prospectively included adult recipients of deceased donor LT who were referred for endoscopic retrograde cholangiopancreatography between June 2009 and July 2011. All patients underwent SOC with biopsy of the biliary anastomosis. Sixteen patients were included: 12 with biliary anastomotic strictures (ASs), 2 with common bile duct stones, 1 with a bile leak, and 1 with sphincter of Oddi dysfunction. Patients with ASs displayed 1 of 2 patterns: (A) mild erythema (n = 9) or (B) edema, ulceration, and sloughing (n = 3). Those without ASs displayed a pale mucosa with mild edema at the anastomosis. Patients with ASs and pattern B required a longer period of stenting than patients with pattern A (457 versus 167 days, P = 0.02). In addition, patients with pattern A had a better response and better resolution of their strictures with endoscopic therapy than those with pattern B (66% versus 33%, P = 0.13). Histological examinations of ASs showed nonspecific intraepithelial inflammation in patients with patterns A and B. Biopsy samples from patients without ASs showed normal columnar epithelial bile duct cells. The total cholangioscopy time for all procedures was 26.8 ± 10.1 minutes. In conclusion, SOC in LT recipients is feasible and allows adequate visualization and tissue sampling of ASs and bile ducts. Two distinct visual patterns that are easily identified with SOC may help to predict the outcomes of endoscopic therapy in patients with biliary complications after LT., (Copyright © 2012 American Association for the Study of Liver Diseases.)

Introduction: Living donor liver transplantation (LDLT) is an effective treatment for patients with terminal chronic liver disease, despite the high incidence of biliary complications. The objective is to evaluate the results and long-term impact of biliary complications after THDV., Patients and Methods: From 2000 to 2010, 70 right lobe LDLT were performed. Biliary complications (leakage and stenosis) of the 70 LDLT recipients were collected prospectively and analyzed retrospectively., Results: A total of 39 patients (55.7%) had some type of biliary complication. Twenty nine presented a leak, and of these, 14 subsequently developed a stricture. In addition, 10 patients had a stenosis without prior leakage. The median time to onset of stenosis was almost a year. Patients with previous biliary leakage were more likely to develop stenosis (58% vs. 29.5% at 5 years, P=.05). With a median follow up of 80 months, 70.8% of patients were successfully treated by interventional radiology. After excluding early mortality, there were no differences in survival according to biliary complications. A decrease of biliary complications was observed in the last 35 patients compared with the first 35., Conclusions: LDLT is associated with a high incidence of biliary complications. However, long-term outcome of patients is not affected. After a median follow-up time of nearly seven years, no differences were found in survival according to the presence of biliary complications., (Copyright © 2012 AEC. Published by Elsevier España, S.L. All rights reserved.)

Background: Few studies have studied the effects of graft quality on non-urgent liver retransplantation (ReLT) outcomes. We aimed to analyze graft characteristics and survival in non-urgent ReLT and the effect of using grafts with extended criteria on survival., Methods: Eighty non-urgent ReLT were performed from June 1988 to June 2010. The whole series was divided by identical time periods to study time-related effects. We assessed graft quality with donor risk index (DRI) and Briceño scores and recipient status with the Model for End-stage Liver Diseases and Rosen scores. Low and high-risk grafts were defined by a DRI cutoff of 1.8., Results: Graft survival was similar in both periods (1-, 5-, and 10-year graft survivals: 73.5, 46.9, and 40.8 versus 71, 47.7, and 47.7%, p=0.935) although donor quality was worse in the second period (DRI: 1.35±0.32 vs. 1.66±0.34, p<0.001). In the first period high-risk grafts did worse than low-risk grafts (5-year survival: 0 vs. 54.5%, p=0.002) while in the second period outcomes were similar (5-year survival: 48.6 vs. 56.7%, p=0.660). Donor age was the only independent donor factor for graft survival, with lower survival when using grafts from donors over 60-years-old., Conclusions: Graft quality in ReLT has worsened with time mainly because of older donors but nowadays the use of high-risk grafts in non-urgent ReLT is not associated with worse graft survival because of better perioperative management. Moreover of being selective on recipient conditions, care should be taken when using grafts from donors over 60-years-old for non-urgent ReLT.

Steatotic livers show increased hepatic damage and impaired regeneration after partial hepatectomy (PH) under ischemia/reperfusion (I/R), which is commonly applied in clinical practice to reduce bleeding. The known function of retinol-binding protein 4 (RBP4) is to transport retinol in the circulation. We examined whether modulating RBP4 and/or retinol could protect steatotic and nonsteatotic livers in the setting of PH under I/R. Steatotic and nonsteatotic livers from Zucker rats were subjected to PH (70%) with 60 minutes of ischemia. RBP4 and retinol levels were measured and altered pharmacologically, and their effects on hepatic damage and regeneration were studied after reperfusion. Decreased RBP4 levels were observed in both liver types, whereas retinol levels were reduced only in steatotic livers. RBP4 administration exacerbated the negative consequences of liver surgery with respect to damage and liver regeneration in both liver types. RBP4 affected the mobilization of retinol from steatotic livers, and this revealed actions of RBP4 independent of simple retinol transport. The injurious effects of RBP4 were not due to changes in retinol levels. Treatment with retinol was effective only for steatotic livers. Indeed, retinol increased hepatic injury and impaired liver regeneration in nonsteatotic livers. In steatotic livers, retinol reduced damage and improved regeneration after surgery. These benefits of retinol were associated with a reduced accumulation of hepatocellular fat. Thus, strategies based on modulating RBP4 could be ineffective and possibly even harmful in both liver types in the setting of PH under I/R. In terms of clinical applications, a retinol pretreatment might open new avenues for liver surgery that specifically benefit the steatotic liver., (Copyright © 2012 American Association for the Study of Liver Diseases.)

Solid-organ transplant recipients are at increased risk of developing cancer compared with the general population. Tumours can arise de novo, as a recurrence of a preexisting malignancy, or from the donated organ. The ATOS (Aula sobre Trasplantes de Órganos Sólidos; the Solid-Organ Transplantation Working Group) group, integrated by Spanish transplant experts, meets annually to discuss current advances in the field. In 2011, the 11th edition covered a range of new topics on cancer and transplantation. In this review we have highlighted the new concepts and best practices for managing cancer in the pre-transplant and post-transplant settings that were presented at the ATOS meeting. Immunosuppression plays a major role in oncogenesis in the transplant recipient, both through impaired immunosurveillance and through direct oncogenic activity. It is possible to transplant organs obtained from donors with a history of cancer as long as an effective minimization of malignancy transmission strategy is followed. Tumour-specific wait-periods have been proposed for the increased number of transplantation candidates with a history of malignancy; however, the patient's individual risk of death from organ failure must be taken into consideration. It is important to actively prevent tumour recurrence, especially the recurrence of hepatocellular carcinoma in liver transplant recipients. To effectively manage post-transplant malignancies, it is essential to proactively monitor patients, with long-term intensive screening programs showing a reduced incidence of cancer post-transplantation. Proposed management strategies for post-transplantation malignancies include viral monitoring and prophylaxis to decrease infection-related cancer, immunosuppression modulation with lower doses of calcineurin inhibitors, and addition of or conversion to inhibitors of the mammalian target of rapamycin., (Copyright © 2012. Published by Elsevier Inc.)

Background: Livers from donation after circulatory determination-of-death (DCD) donors suffer ischemic injury during a preextraction period of cardiac arrest and are infrequently used for transplantation; they have the potential, however, to considerably expand the donor pool. We aimed to determine whether hypothermic oxygenated machine perfusion would improve or further deteriorate the quality of these livers using a clinically relevant porcine model., Methods: Donor livers were subjected to 90 min of cardiac arrest and preserved at 4°C with either static cold storage using University of Wisconsin solution (CS, n=6) or oxygenated machine perfusion using University of Wisconsin machine perfusion solution and 25% physiological perfusion pressures (HMP, n=5). After 4 hr of preservation, livers were transplanted into recipient pigs, which were followed intensively for up to 5 days., Results: Five-day survival was 0 in CS and 20% in HMP. Immediately after reperfusion, hepatocellular injury and function were improved in HMP versus CS. However, HMP grafts also demonstrated significant endothelial and Kupffer cell injury, and a progressive lesion developed 24 to 48 hr after reperfusion that led to death in all but one of the recipient animals., Conclusions: Although hypothermic oxygenated machine perfusion performed using subphysiological perfusion pressures seems to offer some advantages over cold storage in the preservation of ischemically damaged livers, it simultaneously conditions endothelial and Kupffer cell injury that may ultimately lead to the failure of these grafts.

Background: Increased blood pressure (BP) is common after liver transplantation. However, there is scarce information on its control., Methods: In this prospective, cross-sectional, multicenter study, we determined BP according to the recommended international standards in 921 liver transplant patients during one routine outpatient visit to assess their grade of control of BP. At the time of the study, 490 patients had been previously diagnosed with arterial hypertension and were receiving antihypertensive treatment, and 431 were not previously diagnosed as hypertensive., Results: In the hypertensive group, arterial hypertension was uncontrolled (BP >140/90 mm Hg [>130/80 in diabetics]) in 158 (32%) patients and controlled in 332 (68%) patients. In a multivariate analysis, only diabetes was identified as a significant predictor of uncontrolled hypertension. Among patients not previously diagnosed as hypertensive, BP was increased in 106 (25%) and normal in 325 (75%). On multivariate analysis, the only variable independently associated with increased BP in this group was metabolic syndrome., Conclusion: BP is not adequately controlled in a noticeable percentage of liver transplant patients, especially in subjects with diabetes or metabolic syndrome.

Anastomotic strictures (ASs) of the biliary duct after liver transplantation (LT) are primarily managed with endoscopic retrograde cholangiopancreatography (ERCP), but in some cases, this fails because of difficulties in passing the strictures. The aim of this case-control study was to examine specific risk factors for initial ERCP failure and the outcomes of percutaneous transhepatic cholangiography (PTC) as a second-line approach in LT recipients with ASs. Between January 2002 and December 2010, we identified LT recipients with ASs who experienced initial ERCP failure (which was defined as the inability to traverse the AS with guidewires in 2 or more consecutive procedures). A period-matched control group (ratio = 1:2) with ASs and initial ERCP success was analyzed. Preoperative, intraoperative, postoperative, and endoscopic variables were evaluated as risk factors. The outcomes of PTC and the need for hepaticojejunostomy (HJ) or retransplantation were evaluated. Seventeen cases who experienced initial ERCP failure were compared with 34 controls. The median times from LT to ERCP were similar (8.7 months for cases and 8.6 months for controls, P = not significant). A multivariate analysis revealed that previous bile leaks [odds ratio (OR) = 6.07, 95% confidence interval (CI) = 1.0-36.5] and more than 4 U of intraoperatively transfused red blood cells (OR = 11.51, 95% CI = 1.9-71.2) were independent risk factors for failure. PTC was an effective second-line treatment in only 3 of 12 cases (25%). The need for HJ was more frequent for the cases (13/17 or 76.5%) versus the controls (7/34 or 20.6%, P < 0.001). One patient in each group underwent retransplantation (P = not significant). In conclusion, previous bile leaks and high packed red blood cell transfusion requirements during surgery are risk factors for initial ERCP failure in LT recipients with ASs. A high proportion of these patients will need surgery as their final therapy., (Copyright © 2012 American Association for the Study of Liver Diseases.)

Following organ transplantation, lifelong immunosuppressive therapy is required to prevent the host immune system from destroying the allograft. This can cause severe side effects and increased recipient morbidity and mortality. Complete cessation of immunosuppressive drugs has been successfully accomplished in selected transplant recipients, providing proof of principle that operational allograft tolerance is attainable in clinical transplantation. The intra-graft molecular pathways associated with successful drug withdrawal, however, are not well defined. In this study, we analyzed sequential blood and liver tissue samples collected from liver transplant recipients enrolled in a prospective multicenter immunosuppressive drug withdrawal clinical trial. Before initiation of drug withdrawal, operationally tolerant and non-tolerant recipients differed in the intra-graft expression of genes involved in the regulation of iron homeostasis. Furthermore, as compared with non-tolerant recipients, operationally tolerant patients exhibited higher serum levels of hepcidin and ferritin and increased hepatocyte iron deposition. Finally, liver tissue gene expression measurements accurately predicted the outcome of immunosuppressive withdrawal in an independent set of patients. These results point to a critical role for iron metabolism in the regulation of intra-graft alloimmune responses in humans and provide a set of biomarkers to conduct drug-weaning trials in liver transplantation.

Objective: Unexpected donation after cardiac death (DCD) donors suffer cardiac arrest suddenly and are maintained with normothermic extracorporeal membrane oxygenation (NECMO) while consent for donation is obtained. The objective of this study was to determine whether ex vivo normothermic machine perfusion (NMP) improves upon the benefits of NECMO in a large-animal model of unexpected DCD liver transplant., Methods: Donor pigs underwent 90-minute cardiac arrest and were divided in to 3 groups. In the first, livers were preserved immediately with cold storage (CS, n = 6). In the other 2 groups, donors underwent 60-minute NECMO followed by CS (NECMO+CS, n = 6) or NMP (NECMO+NMP, n = 6). After 4-hour preservation, livers were transplanted into recipient pigs., Results: Five-day survival was 0 in CS, 83% in NECMO+CS, and 100% in NECMO+NMP. After reperfusion, injury, and inflammatory markers rose significantly among CS grafts, all of which developed primary nonfunction. Sixty minutes of NECMO, however, resulted in only 1 death, whereas NECMO followed by NMP led to no deaths and significant improvements in injury, inflammation, and synthetic function in comparison to NECMO and CS., Conclusion: Although 60 minutes recuperative NECMO is better than CS alone, NMP improves further on NECMO and may have a role in preserving DCD livers in the clinical setting.

Numerous steatotic livers are discarded as unsuitable for transplantation (TR) because of their poor tolerance of ischemia/reperfusion (I/R). Cyclic adenosine 3',5'-monophosphate (cAMP)-elevating agents protect against I/R injury both in nonsteatotic livers that have been removed from non-heart-beating donors and subjected to warm ischemia or cold ischemia (CIS) and in perfused, isolated livers. Ischemic preconditioning (PC), which is based on brief periods of I/R, protects steatotic liver grafts, but the mechanism that is responsible is poorly understood. This study examines the role of cAMP in the vulnerability shown by steatotic livers to TR-associated I/R injury and the benefits of PC in this situation. Steatotic livers with or without PC were transplanted into Zucker rats. The hepatic levels of cAMP were measured and altered pharmacologically. Our results indicate that the cAMP levels in the nonsteatotic liver grafts were similar to those found in a sham group. However, high cAMP levels were observed in steatotic liver grafts. The blockage of cAMP generation by adenylate cyclase inhibitor pre-treatment or PC had the following results: reduced hepatic injury and increased survival of steatotic graft recipients; greater preservation of adenosine triphosphate (ATP) and reduced lactate accumulation throughout CI. This blockade of cAMP by a nitric oxide-dependent mechanism protected steatotic liver grafts against oxidative stress and microvascular disorders after reperfusion. In conclusion, cAMP blocking-based strategies could protect patients against the inherent risk of steatotic liver failure after TR., (Copyright © 2011 American Association for the Study of Liver Diseases.)

Numerous steatotic livers are discarded for transplantation because of their poor tolerance of ischemia-reperfusion (I/R). The injurious effects of retinol-binding protein 4 (RBP4) in various pathologies are well documented. RBP4 levels are reduced by peroxisome proliferator-activated receptor-γ (PPARγ) agonists. Strategies aimed at increasing PPARγ protect steatotic livers under warm ischemia. Ischemic preconditioning (PC) based on brief periods of I/R protects steatotic liver grafts against I/R injury, but the responsible mechanism is poorly understood. We examined the roles of RBP4 and PPARγ in I/R injury associated with steatotic liver transplantation and the benefits of PC in such situations. We report that RBP4 and PPARγ expression levels in nonsteatotic livers were similar to those found in the sham group. However, reduced RBP4 and increased PPARγ levels were observed in steatotic livers. Treatment with either RBP4 or a PPARγ antagonist was effective only in steatotic livers. PC, which increased RBP4 levels, and RBP4 treatment both reduced PPARγ levels and hepatic injury in steatotic livers. When PPARγ was activated, neither RBP4 treatment nor PC (despite RBP4 induction) protected steatotic livers. In conclusion, steatotic liver grafts are more predisposed to down-regulate RBP4 and overexpress PPARγ. RBP4 treatment and PC, through RBP4 induction, reduced PPARγ levels in steatotic liver grafts, thus protecting them from the PPARγ detrimental effects.

Background: In partial liver transplant, a reduction in the intrahepatic vascular bed produces a rise in the portal vein flow and the portal venous pressure gradient, leading to endothelial and, thereby, hepatocellular injury and death in a process known as "small-for-size" (SFS) syndrome., Objective: To demonstrate that a calibrated portocaval shunt prevents superfluous inflow in a porcine model of SFS transplant., Methods: Donor pigs (15-20 kg) underwent 70% hepatectomy. In 2 groups, a 6 mm (S6) (n = 6) or 12 mm (S12) (n = 6) Gore-Tex shunt was placed between the portal vein and infrahepatic inferior vena cava. In a third group, no portocaval shunt was placed (SFS) (n = 17). Grafts were stored for 5 hours at 4°C and then transplanted into recipients (30-35 kg)., Results: Five-day survival was 29% in SFS, 100% in S6, and 0 in S12. Postreperfusion portal vein flow was 4-, 2-, and 1-times flow at baseline in SFS, S6, and S12, respectively. With respect to portal venous pressure gradient, both the 6- and 12-mm shunts effectively decompressed the portal bed. Aspartate aminotransferase and bilirubin rose and the Quick prothrombin time fell in all animals after reperfusion but improved significantly by day 5 in S6. Serum levels of endothelin-1 remained elevated in SFS and S12 but returned to baseline by 12 hours in S6: 2.76 (2.05-4.08) and 2.04 (1.97-2.12) versus 0.43 (0.26-0.50) pg/mL, respectively (P < 0.05 for both comparisons)., Conclusions: A calibrated portocaval shunt that maintains portal vein flow about twice its baseline value produces a favorable outcome after SFS liver transplantation, avoiding endothelial injury due to portal hyperperfusion or to hypoperfusion because of excess shunting.

Background & Aims: Liver steatosis enhances ischemia/reperfusion (I/R) injury and is considered a primary factor in graft failure after liver transplantation. Although previous reports have shown a role for qualitative steatosis (macrovesicular vs. microvesicular) in hepatic I/R injury, no studies have compared side by side the specific contribution of individual lipids accumulating in fatty liver to I/R damage., Methods: We used nutritional and genetic models of micro and macrovesicular fatty livers exhibiting specific lipid profiles to assess their susceptibility to normothermic I/R injury., Results: Unlike choline-deficient (CD) diet-fed mice, characterized by predominant liver triglycerides/free fatty acids (TG/FFA) accumulation, mice fed a cholesterol-enriched (HC) diet, which exhibited enhanced hepatic cholesterol loading in mitochondria, were highly sensitive to I/R-induced liver injury. In vivo two-photon confocal imaging revealed enhanced mitochondrial depolarization and generation of reactive oxygen species following hepatic I/R in HC-fed but not in CD-fed mice, consistent with decreased mitochondrial GSH (mGSH) observed in HC-fed mice. Moreover, ob/ob mice, characterized by increased hepatic TG, FFA, and cholesterol levels, were as sensitive to I/R-mediated liver injury as mice fed the HC diet. Livers from ob/ob mice displayed increased StAR expression and mitochondrial cholesterol accumulation, resulting in mGSH depletion. Interestingly, atorvastatin therapy or squalene synthase inhibition in vivo attenuated StAR overexpression, mitochondrial cholesterol loading, and mGSH depletion, protecting ob/ob mice from I/R-mediated liver injury., Conclusions: Cholesterol accumulation, particularly in mitochondria, sensitizes to hepatic I/R injury, and thus represents a novel target to prevent the enhanced damage of steatotic livers to I/R-mediated damage., (Copyright © 2010 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Aim: To investigate the benefits of insulin like growth factor-1 (IGF-1) supplementation to serum-free institut georges lopez-1 (IGL-1) solution to protect fatty liver against cold ischemia reperfusion injury., Methods: Steatotic livers were preserved for 24 h in IGL-1 solution supplemented with or without IGF-1 and then perfused "ex vivo" for 2 h at 37degrees C. We examined the effects of IGF-1 on hepatic damage and function (transaminases, percentage of sulfobromophthalein clearance in bile and vascular resistance). We also studied other factors associated with the poor tolerance of fatty livers to cold ischemia reperfusion injury such as mitochondrial damage, oxidative stress, nitric oxide, tumor necrosis factor-α (TNF-α) and mitogen-activated protein kinases., Results: Steatotic livers preserved in IGL-1 solution supplemented with IGF-1 showed lower transaminase levels, increased bile clearance and a reduction in vascular resistance when compared to those preserved in IGL-1 solution alone. These benefits are mediated by activation of AKT and constitutive endothelial nitric oxide synthase (eNOS), as well as the inhibition of inflammatory cytokines such as TNF-α. Mitochondrial damage and oxidative stress were also prevented., Conclusion: IGL-1 enrichment with IGF-1 increased fatty liver graft preservation through AKT and eNOS activation, and prevented TNF-α release during normothermic reperfusion.

Biomarkers that reflect immune response recovery and predict clinical events during withdrawal or minimization of immunosuppressive therapy have not been evaluated. This study aimed to evaluate whether immune response recovers after withdrawal of long-term immunosuppressive treatment in stable liver transplant patients and to determine whether specific biomarkers reflect immune response reactivity and predict rejection. Pharmacokinetic-pharmacodynamic profiles were determined in 24 patients and 80 healthy donors before immunosuppressive treatment reduction began, at 50%, and at complete withdrawal. In patients who rejected, effector-T-cell response mediated by soluble IFN-γ, %CD4(+)IFN-γ and %CD8(+)IL-2/IFN-γ were significantly increased, while TGF-β1 production and the TGF-β1/IFN-γ ratio were significantly decreased. In patients with rejection, soluble IFN-γ and %CD8(+)IL-2 were significantly higher before immunosuppressive treatment was reduced. Further studies are required, but this battery of biomarkers performed in whole blood could be a useful tool to monitor immunosuppressive treatment minimization or withdrawal protocols and identify patients at increased risk of rejection., (Copyright © 2010 Elsevier Inc. All rights reserved.)

During the few last years, after the introduction of high activity antiretroviral therapy (HAART), liver diseases, particularly those related to HCV infection, have emerged as one of the most important causes of mortality in patients with HIV infection. Consequently, liver transplantation is increasingly indicated in this population. Post-transplantation survival in HIV-positive patients with non-hepatitis C virus (HCV) liver diseases is adequate and similar to that in HIV-negative patients. In contrast, survival in patients coinfected with HIV and HCV is only moderate (around 50% at 5 years after transplantation). The main cause of mortality in these patients is HCV recurrence. In almost all patients, HIV infection remains controlled with HAART after liver transplantation. Other issues of interest in this setting are the selection of liver transplantation candidates and the frequent interactions between HAART and immunosuppressive drugs., (2010 Elsevier España, S.L. All rights reserved.)

γδ T cells are innate-type lymphocytes that preferentially act as regulators of local effector immune responses. Recent reports found an altered distribution of the two main subpopulations of blood γδ T cells (Vδ1 and Vδ2) in operationally tolerant liver transplant recipients. Based on this, γδ T cells subset quantification was proposed as a biomarker of immunologic risk in liver transplantation. The specific characteristics of γδ T cell subsets in transplantation remain however unknown. We have investigated here the phenotype, repertoire and functional properties of γδ T cell subsets in a large population of allograft recipients. Our results indicate that alterations in the γδ T cell compartment are not restricted to tolerant liver recipients. In fact, most immunosuppressed liver and kidney recipients also display an enlarged peripheral blood γδ T cell pool mainly resulting from an expansion of Vδ1 T cells exhibiting an oligoclonal repertoire and different phenotypic and cytokine production traits than Vδ2 T cells. We propose that persistent viral infections are likely to contribute to these alterations. Our data provide novel insight in the biology of γδ T cells and a rationale for exploring these lymphocytes in more depth into the pathogenesis of viral infections in transplantation., (© 2010 The Authors. Journal compilation © 2010 European Society for Organ Transplantation.)

This study examined the effects of epidermal growth factor (EGF) and insulin-like growth factor-I (IGF-I) supplementation to University of Wisconsin solution (UW) in steatotic and nonsteatotic livers during cold storage. Hepatic injury and function were evaluated in livers preserved for 24 hours at 4 degrees C in UW and in UW with EGF and IGF-I (separately or in combination) and then perfused ex vivo for 2 hours at 37 degrees C. AKT was inhibited pharmacologically. In addition, hepatic injury and survival were evaluated in recipients who underwent transplantation with steatotic and nonsteatotic livers preserved for 6 hours in UW and UW with EGF and IGF-I (separately or in combination). The results, based on isolated perfused liver, indicated that the addition of EGF and IGF-I (separately or in combination) to UW reduced hepatic injury and improved function in both liver types. A combination of EGF and IGF-I resulted in hepatic injury and function parameters in both liver types similar to those obtained by EGF and IGF-I separately. EGF increased IGF-I, and both additives up-regulated AKT in both liver types. This was associated with glycogen synthase kinase-3beta (GSK3(beta)) inhibition in nonsteatotic livers and PPAR gamma overexpression in steatotic livers. When AKT was inhibited, the effects of EGF and IGF-I on GSK3(beta), PPAR gamma, hepatic injury and function disappeared. The benefits of EGF and IGF-I as additives in UW solution were also clearly seen in the liver transplantation model, because the presence of EGF and IGF-I (separately or in combination) in UW solution reduced hepatic injury and improved survival in recipients who underwent transplantation with steatotic and nonsteatotic liver grafts. In conclusion, EGF and IGF-I may constitute new additives to UW solution in steatotic and nonsteatotic liver preservation, whereas a combination of both seems unnecessary., ((c) 2010 AASLD.)

Aim: To examine the relevance of hypoxia inducible factor (HIF-1) and nitric oxide (NO) on the preservation of fatty liver against cold ischemia-reperfusion injury (IRI)., Methods: We used an isolated perfused rat liver model and we evaluated HIF-1alpha in steatotic and non-steatotic livers preserved for 24 h at 4 degrees C in University of Wisconsin and IGL-1 solutions, and then subjected to 2 h of normothermic reperfusion. After normoxic reperfusion, liver enzymes, bile production, bromosulfophthalein clearance, as well as HIF-1alpha and NO [endothelial NO synthase (eNOS) activity and nitrites/nitrates] were also measured. Other factors associated with the higher susceptibility of steatotic livers to IRI, such as mitochondrial damage and vascular resistance were evaluated., Results: A significant increase in HIF-1alpha was found in steatotic and non-steatotic livers preserved in IGL-1 after cold storage. Livers preserved in IGL-1 showed a significant attenuation of liver injury and improvement in liver function parameters. These benefits were enhanced by the addition of trimetazidine (an anti-ischemic drug), which induces NO and eNOS activation, to IGL-1 solution. In normoxic reperfusion, the presence of NO favors HIF-1alpha accumulation, promoting also the activation of other cytoprotective genes, such as heme-oxygenase-1., Conclusion: We found evidence for the role of the HIF-1alpha/NO system in fatty liver preservation, especially when IGL-1 solution is used.

Understanding the pathogenesis of small-for-size (SFS) syndrome is critical to expanding the applicability of partial liver transplantation. We aimed to characterize its acute presentation and association with alterations in hepatic hemodynamics, microstructure, and regeneration in a porcine model. Eighteen SFS liver transplants were performed. Donors underwent 70% hepatectomy. Partial grafts were implanted into larger recipients. Whole liver transplants were also performed (n = 6). Recipients were followed until death or for 5 days. Hemodynamics were measured, and tissue was sampled intraoperatively and at the study end. Serum was sampled regularly during follow-up. Seventeen SFS transplants and 6 whole liver transplants were included. SFS grafts represented 23.2% (19.3%-25.3%) of the recipients' standard liver volume. The survival rate was 29% and 100% in the SFS and whole liver groups, respectively. The portal venous flow, pressure gradient, and resistance were significantly higher in recipients of SFS grafts versus whole livers after portal and arterial reperfusion. Arterial flow as a percentage of the total liver blood flow was significantly lower after reperfusion in SFS grafts and remained so when measured again after 5 days. Markers of endothelial cell injury increased soon after reperfusion, and those of hepatocellular injury increased later; both predicted the appearance of either graft failure or histological recovery. Proliferative activity peaked earlier and higher among nonsurvivors in the SFS group. Surviving grafts demonstrated a slower but maintained rise in regenerative activity, although metabolic activity failed to improve. In SFS transplantation in the acute setting, portal hyperperfusion is a stimulus for regeneration but may simultaneously cause irreparable endothelial injury. This porcine model not only helps to elucidate the inciting factors in SFS pathogenesis but also offers a clinically relevant means to study its prevention.

Here we examine the effect of adding carvedilol (CVD) to University of Wisconsin (UW) solution on the preservation of steatotic and nonsteatotic livers during cold ischemia and after normothermic reperfusion. We used an isolated perfused rat liver model. The following protocols were evaluated. Protocol 1 concerned the effect of CVD after cold ischemia. Steatotic and nonsteatotic livers were preserved for 24 hours in UW solution alone or with CVD. Livers without cold ischemia were used as controls. Transaminases were evaluated in the flushing effluent. Protocol 2 involved the effect of CVD after reperfusion. Both liver types were preserved for 24 hours in UW solution alone or with CVD and then perfused ex vivo for 2 hours at 37 degrees C. Livers flushed and perfused without ischemia were used as controls. Hepatic injury and functionality [transaminases, bile production, and hepatic clearance of sulfobromophthalein (BSP)] were evaluated after reperfusion. In addition, factors potentially involved in hepatic ischemia-reperfusion injury, including oxidative stress (malondialdehyde and superoxide anion levels), mitochondrial damage (glutamate dehydrogenase activity), microcirculatory disorders (flow rate and vascular resistance), and adenosine triphosphate (ATP) depletion, were evaluated after reperfusion. After cold ischemia, steatotic livers preserved in UW solution showed higher transaminase levels than nonsteatotic livers. After reperfusion, steatotic livers preserved in UW solution showed higher transaminase levels and lower bile production and BSP clearance than nonsteatotic livers. Alterations in the perfusion flow rate and vascular resistance, mitochondrial damage, and reduced ATP content were more evident in steatotic livers preserved in UW solution. The addition of CVD to UW solution reduced hepatic injury, obstructed its mechanisms, and improved hepatic functionality in both liver types. We conclude that CVD is a useful additive for UW solution that improves the preservation of steatotic and nonsteatotic livers subjected to prolonged cold ischemia.

This open-label, randomized study compared the efficacy of a regimen of corticosteroids and tacrolimus (standard therapy group, n = 79) with a regimen of daclizumab induction therapy in combination with mycophenolate mofetil and tacrolimus (modified therapy group, n = 78) in primary liver transplant recipients. The primary endpoint was biopsy-proven acute rejection (BPAR) at 24 weeks. Secondary endpoints included time to rejection and patient and graft survival. The incidence of BPAR was significantly reduced in the modified therapy group compared to the standard therapy group (11.5% versus 26.6%, respectively, P = 0.017). The time to rejection was significantly shorter in the standard therapy group compared with the modified therapy group (P = 0.044). There was no significant difference between groups in patient or graft survival. Hepatitis C virus-positive patients exhibited no differences from hepatitis C virus-negative patients with respect to the incidence of BPAR. A steroid-sparing regimen of daclizumab, mycophenolate mofetil, and tacrolimus was effective and well tolerated in the prevention of BPAR in adult liver transplant recipients in comparison with a standard regimen of tacrolimus and steroids.

Background: In liver transplantation, macrovesicular steatosis is a major determinant of graft outcome. Visual assessment of steatosis by the donor surgeon is highly inaccurate, whereas hepatic biopsy is user dependent and cumbersome. Our objective was to validate a novel bioelectrical impedance sensor as a means of objectively quantifying macrovesicular hepatic steatosis and to correlate the results with another surrogate measure of macrosteatosis, hepatic microcirculation., Methods: Fatty (n=36) and lean (n=18) male Zucker rats, 250 to 450 g, were used to achieve varying degrees of steatosis. After a bilateral subcostal incision, hepatic microcirculation was measured using laser Doppler microflowmetry. Low-frequency bioelectrical impedance (LF-BEI) was measured at 1 kHz using a custom-made sensor and instrumentation system. Complete hepatectomy was performed. Hepatic tissue was preserved and stained with hematoxylin-eosin for histology., Results and Conclusion: Both microflow and LF-BEI correlated well with macrosteatosis and each other: Pearson correlation coefficients -0.71, 0.73, and -0.81, respectively. Livers were grouped according to the degree of macrosteatosis: mild (<30%), moderate (30%-60%), and severe (>60%). Both LF-BEI and microflow varied significantly among groups on one-way analysis of variance, although only LF-BEI was capable of discriminating between mild and moderate macrosteatosis on post hoc analysis. Regarding their individual capacities to detect the presence of severe macrosteatosis, both tests were excellent classifiers: receiver operating curve area under the curve 0.885 and 0.9 for LF-BEI and microflow, respectively. However, the bioimpedance apparatus is more rapid and less susceptible to local factors and background noise and could more easily be used in the clinical liver transplantation setting.

Background: Intracellular ATP (iATP) production in CD4(+)-T cells has been proposed as a biomarker for routine personalized monitoring in transplant recipients. The aim of our study was to measure this biomarker in adult renal and liver transplant recipients receiving moderate immunosuppressive treatment (IST) during the maintenance period and to evaluate whether this biomarker can effectively predict clinical status (stability, rejection, or infection) and drug effect., Methods: iATP concentration and trough blood levels of IST were evaluated in 84 adult transplant recipients (50 renal and 34 liver) during the maintenance period. One hundred and fifty healthy donors were also recruited as a control group. In addition, 22 of 34 liver transplant recipients were included in a clinical study in which IST was withdrawn gradually until total elimination., Results: Most of the patients evaluated had iATP levels in the moderate immune response zone during a posttransplantation maintenance period with no relevant clinical events. iATP levels were significantly lower in transplant patients with infection (n=3) than in those free of infection. In the IST withdrawal study, 10 of 22 liver transplant recipients have achieved complete withdrawal and 12 receive 50% of IST. So far, there have been eight rejections once IST began to be withdrawn. None of these patients had iATP in the strong immune response zone (risk of rejection)., Conclusions: iATP seems to be more effective in identifying overimmunosuppressed patients with a high risk of adverse events (IST-safety) than in identifying those with a high risk of rejection (IST-efficacy) and should be used in combination with other biomarkers of immunosuppressive effect to improve the efficacy of IST.

Hepatic steatosis is a major risk factor in ischemia-reperfusion (I/R). IGF-binding proteins (IGFBPs) modulate IGF-I action by transporting circulating IGF-I to its sites of action. Epidermal growth factor (EGF) stimulates IGF-I synthesis in vitro. We examined the effect of IGF-I and EGF treatment, separately or in combination, on the vulnerability of steatotic livers to I/R. Our results indicated that I/R impaired IGF-I synthesis only in steatotic livers. Only when a high dose of IGF-I (400 microg/kg) was given to obese animals did they show high circulating IGF-I:IGFBP levels, increased hepatic IGF-I levels, and protection against damage. In lean animals, a dose of 100 microg/kg IGF-I protected nonsteatotic livers. Our results indicated that the combined administration of IGF-I and EGF resulted in hepatic injury parameters in both liver types similar to that obtained by IGF-I and EGF separately. IGF-I increased egf expression in both liver types. The beneficial role of EGF on hepatic I/R injury may be attributable to p38 inhibition in nonsteatotic livers and to PPAR gamma overexpression in steatotic livers. In conclusion, IGF-I and EGF may constitute new pharmacological strategies to reduce the inherent susceptibility of steatotic livers to I/R injury.

A rapid, sensitive, and specific method was developed and validated using liquid chromatography-tandem mass spectrometry for the simultaneous quantitation of atorvastatin (ATV) and its major metabolite ortho-hydroxyatorvastatin (o-HATV) in human plasma. The sample preparation involved a liquid-liquid extraction without chlorinated solvents and an on-line solid-phase extraction exploring the possibilities that anion exchange offers. The analytical method presented intraday and day-to-day variation below 10%; intraday and day-to-day accuracy stood between 94% and 105%; the limit of quantification was 0.1 ng/mL for ATV and 0.5 ng/mL for o-HATV; and the recovery was above 75% for both molecules. This method was applied successfully to quantitate ATV and o-HATV concentrations in an unstudied renal transplant recipient cohort treated with an immunosuppressive regime of tacrolimus and mycophenolic acid and a cohort of hypercholesterolemic patients included in the study as a control group. It can be used to evaluate patient adherence, drug-drug interactions, and pharmacokinetic/pharmacodynamic relationships. The results in our study showed that ATV and o-HATV levels in the renal transplant group were significantly increased (p < 0.001), compared to the hypercholesterolemic group.

We examined whether angiotensin (Ang) II receptor antagonists could be considered a therapeutic strategy in steatotic and nonsteatotic livers in conditions of partial hepatectomy under ischemia-reperfusion (I/R), which is commonly applied in clinical practice to reduce blood loss. We report that Ang II type I receptor (AT1R) antagonist, but not Ang II type II receptor (AT2R) antagonist, increased regeneration in nonsteatotic livers. In the presence of steatosis, both AT1R and AT2R antagonists increased liver regeneration. This effect was stronger when the two were combined. Neither of the Ang II receptor antagonists protected nonsteatotic livers against damage. Only the AT1R antagonist, through nitric oxide inhibition, reduced damage in steatotic livers. The combination of the AT1R and AT2R antagonists in steatotic livers conferred a similar degree of protection to AT1R antagonist alone. Herein, we show that p38 mitogen-activated protein kinase (p38) was a key mechanism in the regeneration induced by the Ang II receptor antagonists in both liver types because when this signaling pathway was inhibited, the beneficial effects of the Ang II receptor antagonists on liver regeneration disappeared, regardless of hepatocyte growth factor or transforming growth factor beta-hepatic levels. In conclusion, in conditions of partial hepatectomy under I/R, the AT1R antagonist for nonsteatotic livers and the AT1R and AT2R antagonists for steatotic livers improved regeneration in the remnant liver through p38 activation. In addition, the combination of the AT1R and AT2R antagonists in steatotic livers led to stronger liver regeneration than either antagonists used separately and also provided the same protection against damage as that afforded by AT1R antagonist alone.

This study examined whether angiotensin II (Ang II) blockers [Ang II type I receptor antagonist, Ang II type II receptor antagonist, and angiotensin converting enzyme (ACE) inhibitor] could reduce hepatic injury and improve regeneration in reduced-size orthotopic liver transplantation (ROLT) and whether the beneficial effects of ischemic preconditioning (PC) in ROLT could be explained by changes in Ang II. We show that small liver grafts generated Ang II after ROLT and that this was associated with increased angiotensinogen and ACE messenger RNA expression. Furthermore, inhibition of Ang II did not contribute to PC-induced protection in ROLT. All Ang II blockers reduced hepatic injury, but none of them promoted liver regeneration. Bradykinin (BK) receptor antagonist improved liver regeneration but did not reduce hepatic injury in ROLT. Finally, the combination of Ang II blockers and BK receptor antagonists in ROLT reduced hepatic injury and improved liver regeneration. In conclusion, treatments with either Ang II blockers or BK receptor antagonists cannot, on their own, improve the outcome of ROLT. Although Ang II blockers can reduce hepatic ischemia-reperfusion injury and BK receptor antagonists can promote liver regeneration, neither confers both benefits at the same time. Consequently, it may be of clinical interest to apply both treatments simultaneously.