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Background: We have previously reported the efficacy of post-reperfusion H 2 gas treatment in cold storage (CS) and subsequent reperfusion of the rat liver. The present study aimed to evaluate the effect of H 2 gas treatment during hypothermic machine perfusion (HMP) in rat livers retrieved from donation after circulatory death (DCD) and elucidate the mechanism of action of H 2 gas., Methods: Liver grafts were procured from rats after 30 min of cardiopulmonary arrest. The graft was subjected to HMP for 3 hours at 7°C using Belzer MPS with or without dissolved H 2 gas. The graft was reperfused using an isolated perfused rat liver apparatus at 37°C for 90 minutes. Perfusion kinetics, liver damage, function, apoptosis, and ultrastructure were evaluated., Results: Portal venous resistance, bile production, and oxygen consumption rates were identical in the CS, MP, and MP-H 2 groups. Liver enzyme leakage was suppressed by MP (vs control), whereas H 2 treatment did not show a combination effect. Histopathology revealed poorly stained areas with a structural deformity just below the liver surface in the CS and MP groups, whereas these findings disappeared in the MP-H 2 group. The apoptotic index in the CS and MP groups was high but decreased in the MP-H 2 group. Mitochondrial cristae were damaged in the CS group but preserved in the MP and MP-H 2 groups., Conclusions: In conclusion, HMP and H 2 gas treatment are partly effective in DCD rat livers but insufficient. Hypothermic machine perfusion can improve focal microcirculation and preserve mitochondrial ultrastructure., (Copyright © 2023 Elsevier Inc. All rights reserved.)

We previously reported the efficacy of cold storage (CS) using a heavy water-containing solution (Dsol) and post-reperfusion hydrogen gas treatment separately. This study aimed to clarify the combined effects of these treatments. Rat livers were subjected to 48-hour CS and a subsequent 90-minute reperfusion in an isolated perfused rat liver system. The experimental groups were the immediately reperfused control group (CT), the CS with University of Wisconsin solution (UW) group, the CS with Dsol group, the CS with UW and post-reperfusion H 2 treatment group (UW-H 2 ), and the CS with Dsol and post-reperfusion H 2 group (Dsol-H 2 ). We first compared the Dsol-H 2 , UW, and CT groups to evaluate this alternative method to conventional CS. The protective potential of the Dsol-H 2 group was superior to that of the UW group, as evidenced by lower portal venous resistance and lactate dehydrogenase leakage, a higher oxygen consumption rate, and increased bile production. Multiple comparison tests among the UW, Dsol, UW-H 2 , and Dsol-H 2 groups revealed that both treatments, during CS and after reperfusion, conferred a similar extent of protection and showed additive effects in combination therapy. Furthermore, the variance in all treatment groups appeared smaller than that in the no-treatment or no-stress groups, with excellent reproducibility. In conclusion, combination therapy with Dsol during CS and hydrogen gas after reperfusion additively protects against graft injury., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Objective: The aim of this study was to evaluate sequential hypothermic and normothermic machine perfusion (NMP) as a tool to resuscitate and assess viability of initially declined donor livers to enable safe transplantation., Summary Background Data: Machine perfusion is increasingly used to resuscitate and test the function of donor livers. Although (dual) hypothermic oxygenated machine perfusion ([D]HOPE) resuscitates livers after cold storage, NMP enables assessment of hepatobiliary function., Methods: In a prospective clinical trial, nationwide declined livers were subjected to ex situ NMP (viability assessment phase), preceded by 1-hour DHOPE (resuscitation phase) and 1 hour of controlled oxygenated rewarming (COR), using a perfusion fluid containing an hemoglobin-based oxygen carrier. During the first 2.5 hours of NMP, hepatobiliary viability was assessed, using predefined criteria: perfusate lactate <1.7 mmol/L, pH 7.35 to 7.45, bile production >10 mL, and bile pH >7.45. Livers meeting all criteria were accepted for transplantation. Primary endpoint was 3-month graft survival., Results: Sixteen livers underwent DHOPE-COR-NMP. All livers were from donors after circulatory death, with median age of 63 (range 42-82) years and median Eurotransplant donor risk index of 2.82. During NMP, all livers cleared lactate and produced sufficient bile volume, but in 5 livers bile pH remained <7.45. The 11 (69%) livers that met all viability criteria were successfully transplanted, with 100% patient and graft survival at 3 and 6 months. Introduction of DHOPE-COR-NMP increased the number of deceased donor liver transplants by 20%., Conclusions: Sequential DHOPE-COR-NMP enabled resuscitation and safe selection of initially declined high-risk donor livers, thereby increasing the number of transplantable livers by 20%., Trial Registration: www.trialregister.nl; NTR5972.

Background: Machine perfusion of donor livers is typically performed via the portal vein main stem. Instead, cannulation of a reopened umbilical vein could allow machine perfusion during organ procurement and subsequent implantation in the recipient without interruption of the portal venous circulation. We aimed to assess the feasibility of portal venous machine perfusion via the umbilical vein., Methods: During back table inspection of 5 human livers declined for transplantation, the umbilical vein was surgically reopened, dilated, and cannulated. Hypothermic and normothermic oxygenated machine perfusion (NMP) were performed using the umbilical vein for portal inflow. Three livers were perfused with hypothermic machine perfusion, 1 full liver graft underwent NMP for 4 hours, and 1 left lateral split procedure was performed under continuous NMP with portal perfusion via the umbilical vein., Results: In all livers, access to the portal venous system via the umbilical vein was successfully achieved with good portal flows and macroscopically homogeneous perfusion. The full liver graft that underwent NMP via the umbilical vein for 4 hours showed good lactate clearance, normalized pH, and achieved good bile production with pH >7.55. During the split procedure under continuous NMP via the umbilical vein, the left lateral segment and extended right lobe remained equally perfused, as demonstrated by Doppler ultrasound., Conclusions: Machine perfusion with portal perfusion via the umbilical vein is feasible. Portal venous flows were similar to those obtained after cannulation of the portal vein main stem. This technique enables continuous oxygenated perfusion of liver grafts during procurement, splitting, and implantation.

Background and Purpose: We reported previously that hydrogen gas (H 2 ) reduced hepatic ischemia and reperfusion injury (IRI) after prolonged cold storage (CS) of livers retrieved from heart-beating donors. The present study was designed to assess whether H 2 reduced hepatic IRI during donation of a cardiac death (DCD) graft with subsequent CS., Methods: Rat livers were harvested after 30-min cardiac arrest and stored for 4 h in University of Wisconsin solution. The graft was reperfused with oxygenated buffer, with or without H 2 (H 2 or NT groups, respectively), at 37° for 90 min on isolated perfused rat liver apparatus., Results: In the NT group, liver enzyme leakage, apoptosis, necrosis, energy depletion, redox status, impaired microcirculation, and bile production were indicative of severe IRI, whereas in the H 2 group these impairments were significantly suppressed. The phosphorylation of cytoplasmic MKK4 and JNK were enhanced in the NT group and suppressed in the H 2 group. NFkB-p65 and c-Fos in the nucleus were unexpectedly unchanged by IRI regardless of H 2 treatment, indicating the absence of inflammation in this model., Conclusion: H 2 was observed to ameliorate IRI in the DCD liver by maintaining microcirculation, mitochondrial functions, and redox status, as well as suppressing the cytoplasmic MKK4-JNK-mediated cellular death pathway.