Liver Perfusion Research Articles

Continuous Blood Gas Control Based on Active Disturbance Rejection Control During ExVivo Porcine Liver Perfusion.

Membrane oxygenators facilitate extracorporeal gas exchange, necessitating the monitoring of blood gas. Recent advances in normothermic machine perfusion (NMP) for exvivo liver offer solutions to the shortage of donor liver. However, maintaining physiological blood gas levels during prolonged NMP is complex and costly. We introduce a noninvasive and economical approach for regulating the blood gas during NMP of exvivo porcine livers. By monitoring gas fractions at the outlet of oxygenator, real-time adjustments of blood gas can be made without the online blood gas analyzer. The method involves constructing multivariate linear regression (MLR) models, aligning target setpoints of gas, and employing active disturbance rejection control (ADRC) to achieve closed-loop regulation. Ex vivo porcine liver perfusion experiments demonstrated the effectiveness of the method, maintaining blood gas within physiological levels over 24 h (oxygen partial pressure: 150.36 ± 3.33 mmHg, carbon dioxide partial pressure: 41.34 ± 0.91 mmHg). ADRC-based continuous regulation of gas fraction at the outlet of oxygenator is a feasible and effective approach for managing blood gas during exvivo porcine liver perfusion.

Model-based perfusion reconstruction with time separation technique in cone-beam CT dynamic liver perfusion imaging.

The success of embolization, a minimally invasive treatment of liver cancer, could be evaluated in the operational room with cone-beam CT by acquiring a dynamic perfusion scan to inspect the contrast agentflow. The reconstruction algorithm must address the issues of low temporal sampling and higher noise levels inherent in cone-beam CT systems, compared to conventional CT. Therefore, a model-based perfusion reconstruction based on the time separation technique (TST) was applied. TST uses basis functions to model time attenuation curves. These functions are either analytical or based on prior knowledge (PK), extracted using singular value decomposition of the classical CT perfusion data of animal subjects. To explore how well the PK can model perfusion dynamics and what the potential limitations are, the dynamic cone-beam CT (CBCT) perfusion scan was simulated from a dynamic CT perfusion scan under different noise levels. The TST method was compared to staticreconstruction. It was demonstrated on this simulated dynamic CBCT perfusion scan that a set consisting of only four basis functions results in perfusion maps that preserve relevant information, denoise the data, and outperform static reconstruction under higher noise levels. TST with PK would not only outperform static reconstruction but also the TST with analytical basis functions. Furthermore, it has been shown that only eight CBCT rotations, unlike previously assumed ten, are sufficient to obtain the perfusion maps comparable to the reference CT perfusion maps. This contributes to saving dose and reconstruction time. The real dynamic CBCT perfusion scan, reconstructed under the same conditions as the simulated scan, shows potential for maintaining the accuracy of the perfusion maps. By visual inspection, the embolized region was matching to that in corresponding CT perfusionmaps. CBCT reconstruction of perfusion scan data using the TST method has shown promising potential, outperforming static reconstructions and potentially saving dose by reducing the necessary number of acquisition sweeps. Further analysis of a larger cohort of patient data is needed to draw final conclusions regarding the expected advantages of theTST.

Reduced fetal ductus venosus shunt fraction is associated with adverse perinatal outcomes in pregnancy with pregestational diabetes mellitus.

We aimed to determine fetal liver perfusion in PGDM and GDM pregnancies and to assess the relation of ductus venosus (DV) shunt fraction with adverse pregnancy outcomes. We conducted a prospective longitudinal observational study including 188 pregnant women: group I-patients with pregestational DM (PGDM, n = 86), group II-patients with gestational DM (GDM, n = 44), group III-control (n = 58). The patients included in the study underwent ultrasound examination at 30+0-40+0weeks of pregnancy. We evaluated volumetric blood flow adjusted to EFW (Q, ml/min/kg) for umbilical vein, DV, left and main portal vein. The relative risk was calculated for adverse pregnancy outcomes. In PGDM pregnancies, umbilical blood flow was redistributed to the fetal liver, increasing left portal and total liver volumetric blood flow (p < 0.001) compared with GDM and control groups. Pathological reduction in the DV shunt fraction (≤ 16.5%) was associated with an increased relative risk of preterm delivery (3.61 [95%CI 1.68; 7.71]), LGA-birth (1.64 [95% CI 1.26; 2.12]), neonatal adiposity (1.53 [95%CI 1.18; 1.98]), fetal hypoxia (3.47 [95%CI 1.34; 9.05]), emergency cesarean Sect.(1.93 [95%CI 1.26; 2.97]), and neonatal intensive care unit stay of more than 5days (1.78 [95%CI 1.08; 2.93]). Decreased DV shunt fraction reflects changes in fetal hemodynamics in PGDM-pregnancies and associated with an increased risk of adverse perinatal outcomes.

A simplified computational liver perfusion model, with applications to organ preservation

Advanced liver preservation strategies could revolutionize liver transplantation by extending preservation time, thereby allowing for broader availability and better matching of transplants. However, developing new cryopreservation protocols requires exploration of a complex design space, further complicated by the scarcity of real human livers to experiment upon. We aim to create computational models of the liver to aid in the development of new cryopreservation protocols. Towards this goal, we present an approach for generating 3D models of the liver vasculature by building upon the space colonization algorithm. Additionally, we introduce the concept of a super lobule which enables a computational abstraction of biological liver lobules. User-tunable parameters allow for vasculatures of varying depth and topology to be generated. In each model, we solve for a common lumped resistance value assigned to the super lobules, allowing the overall physiological blood pressure and flow rate through the liver to be preserved. We demonstrate our approach’s ability to maintain consistency between models of varying depth. Finally, we simulate steady state machine perfusion of the generated models and demonstrate how they can be used to quickly test the effect of different boundary conditions when designing organ preservation protocols.

Real-Time Biomarkers of Liver Graft Quality in Hypothermic Oxygenated Machine Perfusion.

Background: Hypothermic oxygenated machine perfusion has emerged as a strategy to alleviate ischemic-reperfusion injury in liver grafts. Nevertheless, there is limited data on the effectiveness of hypothermic liver perfusion in evaluating organ quality. This study aimed to introduce a readily accessible real-time predictive biomarker measured in machine perfusate for post-transplant liver graft function. Methods: The study evaluated perfusate analytes over a 90-day postoperative period in 26 patients randomly assigned to receive a liver graft following dual hypothermic machine perfusion in a prospective randomized controlled trial. Machine perfusion was consistently conducted end-ischemically for at least 120 min, with real-time perfusate assessment at 30-min intervals. Graft functionality was assessed using established metrics, including Early Allograft Dysfunction (EAD). Results: Perfusate lactate concentration after 120 min of machine perfusion demonstrated significant predictive value for EAD (AUC ROC: 0.841, p = 0.009). Additionally, it correlated with post-transplant peak transaminase levels and extended hospital stays. Subgroup analysis revealed significantly higher lactate accumulation in livers with post-transplant EAD. Conclusions: Liver graft quality can be effectively assessed during hypothermic machine perfusion using simple perfusate lactate measurements. The reliability and accessibility of this evaluation support its potential integration into diverse transplant centers.

Artificial Liver Support Systems in Acute Liver Failure and Acute-on-Chronic Liver Failure: Systematic Review and Meta-Analysis.

To systematically review the safety and efficacy of nonbiological (NBAL) or biological artificial liver support systems (BAL) and whole-organ extracorporeal liver perfusion (W-ECLP) systems, in adults with acute liver failure (ALF) and acute-on-chronic liver failure (ACLF). Eligible NBAL/BAL studies from PubMed/Embase searches were randomized controlled trials (RCTs) in adult patients with ALF/ACLF, greater than or equal to ten patients per group, reporting outcomes related to survival, adverse events, transplantation rate, and hepatic encephalopathy, and published in English from January 2000 to July 2023. Separately, we searched for studies evaluating W-ECLP in adult patients with ALF or ACLF published between January1990 and July 2023. Two researchers independently screened citations for eligibility and, of eligible studies, retrieved data related to study characteristics, patients and interventions, outcomes definition, and intervention effects. The Cochrane Risk of Bias 2 tool and Joanna Briggs Institute checklists were used to assess individual study risk of bias. Meta-analysis of mortality at 28-30 days post-support system initiation and frequency of at least one serious adverse event (SAE) generated pooled risk ratios (RRs), based on random (mortality) or fixed (SAE) effects models. Of 17 trials evaluating NBAL/BAL systems, 11 reported 28-30 days mortality and five reported frequency of at least one SAE. Overall, NBAL/BAL was not statistically associated with mortality at 28-30 days (RR, 0.85; 95% CI, 0.67-1.07; p = 0.169) or frequency of at least one SAE (RR, 1.15; 95% CI, 0.99-1.33; p = 0.059), compared with standard medical treatment. Subgroup results on ALF patients suggest possible benefit for mortality (RR, 0.67; 95% CI, 0.44-1.03; p = 0.069). From six reports of W-ECLP (12 patients), more than half (58%) of severe patients were bridged to transplantation and survived without transmission of porcine retroviruses. Despite no significant pooled effects of NBAL/BAL devices, the available evidence calls for further research and development of extracorporeal liver support systems, with larger RCTs and optimization of patient selection, perfusion durability, and treatment protocols.

CT perfusion and MRI: A combined approach for hepatocellular carcinoma diagnosis and follow-up after locoregional treatment.

CT liver perfusion (CTLP) has been well validated for hepatocellular carcinoma (HCC) detection, characterization, and treatment response evaluation. However, its role in HCC management algorithms remains unclear. This study aims to assess the diagnostic performance of CTLP alone or as an adjunct to MRI in patients considered for- or undergoing locoregional treatment for HCC. Thirty-nine patients under HCC surveillance (36 male, 31 cirrhotic, 16 pretreatment, 19 post-transarterial chemoembolization, 2 post-ablation) underwent MRI and CTLP in a single center within a 45-day interval. Two readers identified and characterized all observations on MRI using Liver Imaging Reporting and Data System (LI-RADS) v2018 criteria. CTLP assessment was based on Mean Slope of Increase (MSI), Time To Peak (TTP), Hepatic arterial Blood Flow (HaBF) and Hepatic Arterial Fraction (HAF) maps and established cut-offs. Diagnostic performance of MRI, CTLP, and their combination was evaluated for treated and untreated lesions using imaging or pathology as reference standard. Of the total 33 treated and 61 untreated lesions, 13 and 41 were considered viable HCCs. CTLP demonstrated 75.9 % sensitivity and 95 % specificity compared to 72.2 % and 100 % for MRI (p > 0.05). Combining both modalities increased sensitivity to 85.2 % (p < 0.05) and maintained specificity at 97.5 % (p > 0.05). The combined approach led to an LR category change in 5 treated and 19 untreated lesions and affected management in 5 cases. CTLP and MRI have comparable diagnostic performance for HCC. A combined approach improves sensitivity, without sacrificing specificity. This approach might enable more efficient patient selection for early and individualized loco-regional treatment.

Incessant ventricular tachycardia and Multi-organ failure after inferior STEMI due to Right coronary artery occlusion

Inferior wall myocardial infarction (STEMI) is commonly associated with hemodynamic instability and conduction disturbances; however, incessant ventricular tachycardia (VT) as a complication is rare. We report a case of a 66-year-old male with inferior STEMI due to right coronary artery (RCA) occlusion, who developed incessant monomorphic VT, cardiogenic shock, and subsequent multi-organ failure. Despite initial medical therapy, including dual antiplatelet therapy (DAPT), statins, and anticoagulation, the patient’s arrhythmia persisted, necessitating urgent percutaneous coronary intervention (PCI). Post-PCI, the patient experienced cardiogenic shock complicated by acute renal failure, ischemic hepatitis, disseminated intravascular coagulation (DIC), and required hemodialysis. Intensive critical care measures, including mechanical ventilation and renal replacement therapy, led to clinical improvement, and the patient was discharged after 18 days, hemodynamically stable with restored organ function.

Cytokine adsorption during normothermic machine perfusion of porcine livers

Cytokine adsorption during normothermic machine perfusion of porcine livers

Pictorial review of ischaemic hepatitis and its mimics.

Pictorial review of ischaemic hepatitis and its mimics.

A Novel Machine Perfusion System for Enhancing Hepatic Microcirculation Perfusion

ABSTRACTBackgroundMachine perfusion is a promising strategy for safeguarding liver transplants donated after cardiac death (DCD). In this study, we developed and validated a novel machine perfusion approach for mitigating risk factors and salvaging severe DCD livers.MethodsA novel hypothermic oxygenated perfusion (HOPE) system was developed, incorporating two pumps and an elastic water sac to emulate the functionality of the cardiac cycle. Compared to conventional systems (HOPE S1 and S2), the novel HOPE system (HOPE S3) was evaluated in rats, utilizing healthy livers perfused with methylene blue diluted using Histidine‐tryptophan‐ketoglutarate (HTK) solution or DCD livers subjected to 60 min of warm ischemia without heparin administration. Liver perfusion outcomes were assessed through macroscopic and microscopic evaluations, molecular analyses, and orthotopic liver transplantation (OLT).ResultsDCD livers subjected to HOPE systems' perfusion exhibited decreased injury and enhanced survival rates compared to static cold storage following 60 min of warm ischemia (DCD + SCS). The 4‐week post‐transplantation survival rates were 0%, 20%, and 33% in the DCD + SCS, HOPE S1, and HOPE S2 groups, respectively. HOPE S3 conferred protection against hepatocyte and non‐parenchymal cell injury, resulting in a 67% animal survival rate following 60 min of warm donor ischemia (HOPE S3). Assessments of hepatic sinusoidal microcirculation, morphological changes, and molecular alterations in preserved livers further confirmed these findings.ConclusionsThe newly devised machine perfusion system can enhance and uniform liver perfusion and may become a promising tool for revitalizing DCD liver grafts afflicted with severe warm ischemic injuries.

The caveat of biliary pH as biomarker of bile duct viability during normothermic machine perfusion of donor livers.

The caveat of biliary pH as biomarker of bile duct viability during normothermic machine perfusion of donor livers.

Rat Liver Perfusion and Primary Hepatocytes Isolation: An Old Procedure Crucial for Cutting-Edge 3D Organoids Culture.

Primary hepatocytes are a commonly used tool for in vitro liver-related studies. However, the maintenance of these cells has always been a challenge due to the rapid loss of morphology, viability, and functionality in culture. A recent approach to long-term culture is the generation of three-dimensional (3D) organoids, an in vitro tool that can recapitulate tissues in a dish based on the marvelous ability of the liver to regenerate itself. Published protocols have been designed to obtain long-term functional 3D organoids from primary adult hepatocytes (Hep-Orgs). The 3D organoid cutting-edge tool requires the ability to isolate cells from adult tissue, and this initial step is crucial for a high-quality final result. The two-step collagenase perfusion, introduced in the 1970s, is still a valid procedure to obtain single hepatocytes. The present article aims to describe all the crucial steps of the surgical procedure, thereby optimizing the primary hepatocytes isolation procedure in the rat model. Moreover, particular attention is paid to the PREPARE guidelines to increase the likelihood of successful procedures and ensure high-quality results. A detailed protocol allows researchers to speed up and optimize the downstream work to establish 3D organoids from primary adult rat hepatocytes. Compared to 2D hepatocytes, Hep-Orgs were still viable and in active proliferation at Day 15, demonstrating a long-term potential.

Unlocking the Promise of Liver Perfusion Technologies for Pediatric Transplantation: A State-of-the-Art Review.

Outcomes after pediatric liver transplantation are generally excellent, but the limited avavailability of suitable, size-matched liver allografts remains a significant barrier. Machine perfusion technology has emerged as a promising approach to expand the donor pool, enabling the use of less ideal whole liver grafts, such as livers donated after circulatory death, and enhancing the execution of split liver transplantation. This review examines the application of machine perfusion in pediatric liver transplantation, focusing on two primary techniques: hypothermic oxygentaed perfusion and normothermic machine perfusion. These methods optimize storage, resuscitation, and assessment of liver grafts before transplantation, potentially expanding the range of usable donor organs. The use of machine perfusion allows for the consideration of suboptimal donor livers and facilitates split liver transplantation, both of which could increase organ availability for pediatric patients. Implementation of machine perfusion could also help reduce waiting list mortality by enabling the safe use of a broader spectrum of donor organs. Adoption of machine perfusion in pediatric liver transplantation will require collaborative, multidisciplinary efforts across transplant centers. By fostering cooperative learning and sharing resources. the integration of machine perfusion into clinical practice has the potential to reduce mortality among children awaiting liver transplantation.

Mouse liver blood flow is regulated by hepatic stellate cells in response to the sympathetic neurotransmitter norepinephrine

BackgroundThere is no clear information on the regulation of liver blood flow by the autonomic nervous system. We conducted this study to investigate whether quiescent hepatic stellate cells (qHSCs) regulate liver blood flow in response to the sympathetic neurotransmitter norepinephrine (NE). MethodsqHSCs isolated from mice were cultured in Dulbecco's modified Eagle medium without fetal bovine serum for 1 day on collagen gel. NE-induced qHSC contraction was evaluated using the quantitative single-cell contraction measurement method that we had developed previously. For the measurement of liver perfusion pressure in situ, a buffer solution was perfused from the portal vein in mice. ResultsNE-induced a reversible contraction of qHSCs. This contraction was suppressed by the nonmuscle myosin II inhibitor blebbistatin, the myosin light chain kinase inhibitor ML-9, the Rho kinase inhibitor H-1152, the calmodulin inhibitor W-7, the store-operated calcium channel inhibitor YM-58483, and the IP3 receptor inhibitor xestospongin C. In contrast, the transient receptor potential C channel inhibitor SKF96365 did not affect the NE-induced contraction. ConclusionThese results suggest that qHSCs contract in response to NE. New & noteworthyThe present study provides direct evidence for the first time that norepinephrine (NE) induces a reversible contraction of isolated single quiescent hepatic stellate cells (qHSCs) and further suggests that the NE-mediated qHSC contraction participates in the regulation of liver blood flow in vivo.

Gastrointestinal complications and visceral circulation changes after intentional celiac artery embolization during complex endovascular aortic repair

Intentional celiac artery embolization (CAE) is an oft-used strategy to extend proximal or distal seal during complex endovascular aortic repair. Prior reports document a wide range of gastointestinal-related complications. However, associated changes in collateral circulation are poorly defined. We sought to report the long-term outcomes and adaptive changes in collateral visceral circulation following CAE during complex endovascular aortic repair. All patients undergoing complex endovascular aortic procedures (endovascular aortic repair [EVAR] and thoracic EVAR) with CAE at a single institution over a 12-year period were included. Pre- and postoperative clinical, radiologic, and laboratory data were reviewed to identify mesenteric complications related to CAE and to assess long-term survival and radiologic changes in collateral anatomy. Multivariable logistic regression was used to determine the association between collateral vessel diameter change and mesenteric complications. From 2011 to 2023, 70 patients underwent planned CAE during complex EVAR. With regards to mesenteric complications not attributable to the superior mesenteric artery (SMA) stent, 11.4% had 30-day mesenteric complications, including upper gastrointestinal bleed or perforated ulcer (n= 3), cholecystitis (n= 2), pancreatitis (n= 2), and ischemic hepatitis (n= 1). During 31 to 90days after CAE, two additional patients (2.9%) had upper gastrointestinal bleed. With regards to 90-day mesenteric complications related to the SMA stent, four additional patients (5.7%) had SMA stent complications leading to mesenteric ischemia. On Kaplan-Meier analysis, patients with any 90-day mesenteric complication had significantly lower overall 2-year survival (42.5% vs 75.0%; P= .002). On preoperative imaging, 20% of patients had variant SMA anatomy with the gastroduodenal artery as the dominant SMA-celiac collateral pathway in 68.6%. Postoperatively, patients without mesenteric complications had a greater increase in the collateral diameter at both SMA and celiac junctions at 1, 3 to 6, 12, and 24months, with a statistically significant difference in diameter at 1month compared with patients with complications (median: 16.2% vs-2.1% at celiac; P= .006 and 20.8% vs 7.7% at SMA; P= .021). On adjusted multivariate regression, increase in collateral diameter at the SMA junction on first postoperative computed tomography was significantly protective of 90-day mesenteric complications (odds ratio, 0.93; 95% confidence interval, 0.87-0.96; P= .046). CAE during complex EVAR is a useful adjunct to increase seal zone in select patients; however, mesenteric complications occur in 14% of the patients over a 90-day postoperative period, and patients with mesenteric complications have a higher long-term all-cause mortality. CAE should be a technique within the toolbox of vascular surgeons for urgent circumstances that do not allow for celiac preservation. Careful selection of candidates for CAE and early postoperative surveillance of collateral pathways may help with prevention and early identification of long-term visceral complications.

Routine end-ischemic hypothermic machine perfusion in liver transplantation from donors after brain death: results of 2-year follow-up of a randomized controlled trial.

Data on routine hypothermic machine perfusion of livers procured from donors after brain death (DBD) are scarce, and the benefits of the method have only been demonstrated in extended criteria grafts. This study aimed to assess if end-ischemic dual hypothermic oxygenated machine perfusion (dHOPE) is superior to static cold storage (SCS) in preservation of livers procured from DBD donors with respect to long-term outcomes. Existing data on short-term outcomes favours dHOPE in patients receiving high-risk grafts. This prospective randomized controlled trial included 104 recipients of DBD livers randomly assigned to SCS arm (78 patients) and the dHOPE arm (26 patients). Endpoints of interest were the occurrence of biliary complications (biliary fistula, anastomotic, and nonanastomotic strictures) and overall patient and graft survival (GS) during the 2-year follow-up. A total of 36 patients developed biliary complications (at least one event) - six events in dHOPE arm and 30 in SCS arm. There was no significant difference in biliary complications between groups (23.7 vs. 43.4%, P =0.11). No differences were found significant with respect to anastomotic (19.9 vs. 33.7%, P =0.20) and nonanastomotic strictures (0 vs. 11.1%, P =0.10) as well as biliary fistulas (11.7 vs. 12.2%, P =0.93). Survival analysis did not show significantly different results in the study population - overall survival: 92.3% in dHOPE and 83.9% in SCS ( P =0.35), and GS: 92.3 and 81.4% ( P =0.23), respectively. However, a significant difference in GS was noted in recipients of high-risk grafts - 100% in dHOPE and 73.1% in SCS, respectively ( P =0.038). The long-term outcome data suggest that the routine use of dHOPE may be beneficial for recipients of high-risk grafts from DBD donors. The present study does not provide any evidence for the benefits of dHOPE in low-risk grafts.

The Primary Cause of Markedly Elevated Aminotransferases in Hospitalized Patients with Cirrhosis in Ischemic Hepatitis: Erratum.

The Primary Cause of Markedly Elevated Aminotransferases in Hospitalized Patients with Cirrhosis in Ischemic Hepatitis: Erratum.

Hepatic Stellate Cell-mediated Increase in CCL5 Chemokine Expression after X-ray Irradiation Determined In Vitro and In Vivo.

Radiation exposure causes hepatitis which induces hepatic steatosis and fibrosis. Although hepatic stellate cells (HSCs) have been considered potential pathological modulators for the development of hepatitis due to viral and microbial infections, their involvement in radiation-induced hepatitis is yet to be determined. This study aimed to clarify the relationship between radiation exposure and expressions of inflammatory cytokines and chemokines in HSCs in vitro and invivo. HSCs were obtained from 1-week-old mice, known to be highly sensitive to radiation-induced hepatocellular carcinoma, using a newly established method combining liver perfusion, cell dissociation, and density gradient centrifugation, followed by magnetic negative selection of hematopoietic and endothelial cells with anti-CD45.2 and CD146 antibodies. The isolated HSCs were confirmed by the expression of desmin and glial fibrillary acidic protein (GFAP). We demonstrated that primary cultured HSCs, exposed to X-ray irradiation (0, 1.9, and 3.8 Gy) and cultured for 3 and 7 days, produced elevated levels of C-C motif chemokine ligand 5 (CCL5, also known as RANTES) inflammatory chemokine in a dose-dependent manner. An invivo immunofluorescence method confirmed that increased CCL5 signals were observed in GFAP-positive HSCs in mouse livers 7 days after whole-body X-ray irradiation (1.9 and 3.8 Gy). Adequate expression of C-C motif chemokine receptor 5 (Ccr5), a receptor for CCL5, was also detected using real-time PCR in the liver of both irradiated and non-irradiated mice. Taken together, our data suggest that HSCs may drive hepatitis via CCL5/CCR5 axis in the liver under radiation-induced stress. Furthermore, this newly established experimental protocol can help evaluate the expression of other inflammatory cytokines in primary cultures of HSCs isolated from infant mice.

A reproducible extended ex-vivo normothermic machine liver perfusion protocol utilising improved nutrition and targeted vascular flows

BackgroundNormothermic machine perfusion of donor livers has become standard practice in the field of transplantation, allowing the assessment of organs and safe extension of preservation times. Alongside its clinical uses, there has been expanding interest in extended normothermic machine perfusion (eNMP) of livers as a potential vehicle for medical research. Reproducible extended normothermic machine perfusion has remained elusive due to its increased complexity and monitoring requirements. We set out to develop a reproducible protocol for the extended normothermic machine perfusion of whole human livers.MethodsHuman livers declined for transplantation were perfused using a blood-based perfusate at 36 °C using the Liver Assist device (XVIVO, Sweden), with continuous veno-venous haemofiltration in-parallel. We developed the protocol in a stepwise fashion.ResultsPerfusion techniques utilised included: targeted physiological vascular flows, phosphate replacement (to prevent hypophosphataemia), N-acetylcysteine (to prevent methaemoglobin accumulation), and the utilisation of sodium lactate as both a nutritional source and real-time measure of hepatocyte function. All five human livers perfused with the developed protocol showed preserved function with a median perfusion time of 168 h (range 120–184 h), with preserved viability throughout.ConclusionsLivers can be reproducibly perfused in excess of 120 (range 121–184) hours with evidence of preserved hepatocyte and cholangiocyte function.