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Background: Cell‐free microRNAs (miRs) have emerged as early and sensitive biomarkers for tissue injury and function. This study aimed to investigate whether the release of hepatocyte‐derived microRNAs (HDmiRs) and cholangiocyte‐derived miRs (CDmiRs) correlates with hepato‐cholangiocellular injury and function during oxygenated, normothermic machine perfusion (NMP) of human liver grafts. Methods: Donor livers (n = 12), declined for transplantation, were subjected to oxygenated NMP (6 hours) after a period of static cold storage (median 544 minutes (IQR 421‐674)). Perfusate and bile samples were analyzed by qRT‐PCR for HDmiR‐122 and CDmiR‐222. Spearman correlations were performed between miR levels and currently available indicators and classic markers. Results: Both HDmiR‐122 and CDmiR‐222 levels in perfusate at 30 minutes of NMP strongly correlated with hepatocyte injury (peak perfusate AST) and cholangiocyte injury (peak biliary LDH). In bile, only CDmiR‐222 correlated with these injury markers. For hepato‐cholangiocellular function, both miRs in perfusate correlated with total bilirubin, while HDmiR‐122 (in perfusate) and CDmiR‐222 (in bile) correlated with bicarbonate secretion. Both the relative ratio of HDmiR‐122/CDmiR‐222 and AST in perfusate at 30 minutes significantly correlated with cumulative bile production, but only the relative ratio was predictive of histopathological injury after 6 hours NMP. Conclusion: Early levels of HDmiR‐122 and CDmiR‐222, in perfusate and/or bile, are predictive of excretory functions and hepato‐cholangiocellular injury after 6 hours NMP. These miRs may represent new biomarkers for graft viability and function during machine perfusion. [ABSTRACT FROM AUTHOR]

Background: The major concern in liver transplantation of grafts from donation after circulatory death (DCD) donors remains the high incidence of non-anastomotic biliary strictures (NAS). Machine perfusion has been proposed as an alternative strategy for organ preservation which reduces ischemia-reperfusion injury (IRI). Experimental studies have shown that dual hypothermic oxygenated machine perfusion (DHOPE) is associated with less IRI, improved hepatocellular function, and better preserved mitochondrial and endothelial function compared to conventional static cold storage (SCS). Moreover, DHOPE was safely applied with promising results in a recently performed phase-1 study. The aim of the current study is to determine the efficacy of DHOPE in reducing the incidence of NAS after DCD liver transplantation.Methods: This is an international multicenter randomized controlled trial. Adult patients (≥18 yrs. old) undergoing transplantation of a DCD donor liver (Maastricht category III) will be randomized between the intervention and control group. In the intervention group, livers will be subjected to two hours of end-ischemic DHOPE after SCS and before implantation. In the control group, livers will be subjected to care as usual with conventional SCS only. Primary outcome is the incidence of symptomatic NAS diagnosed by a blinded adjudication committee. In all patients, magnetic resonance cholangiography will be obtained at six months after transplantation.Discussion: DHOPE is associated with reduced IRI of the bile ducts. Whether reduced IRI of the bile ducts leads to lower incidence of NAS after DCD liver transplantation can only be examined in a randomized controlled trial.Trial Registration: The trial was registered in Clinicaltrials.gov in September 2015 with the identifier NCT02584283 . [ABSTRACT FROM AUTHOR]

• Six hours of ex vivo subnormothermic machine perfusion (SNMP) of rodent vascularized composite allografts (VCA) is feasible using acellular oxygen carrier HBOC-201 • SNMP results in superior tissue preservation compared to conventional static cold storage (SCS) preservation • Successful transplantation of VCA grafts preserved with 6 h of SNMP is feasible and 30-d • survival rates are comparable to SCS preserved graft Machine perfusion is gaining interest as an efficient method of tissue preservation of Vascularized Composite Allografts (VCA). The aim of this study was to develop a protocol for ex vivo subnormothermic oxygenated machine perfusion (SNMP) on rodent hindlimbs and to validate our protocol in a heterotopic hindlimb transplant model. In this optimization study we compared three different solutions during 6 h of SNMP (n = 4 per group). Ten control limbs were stored in a preservation solution on Static Cold Storage [SCS]). During SNMP we monitored arterial flowrate, lactate levels, and edema. After SNMP, muscle biopsies were taken for histology examination, and energy charge analysis. We validated the best perfusion protocol in a heterotopic limb transplantation model with 30-d follow up (n = 13). As controls, we transplanted untreated limbs (n = 5) and hindlimbs preserved with either 6 or 24 h of SCS (n = 4 and n = 5). During SNMP, arterial outflow increased, and lactate clearance decreased in all groups. Total edema was significantly lower in the HBOC-201 group compared to the BSA group (P = 0.005), 4.9 (4.3-6.1) versus 48.8 (39.1-53.2) percentage, but not to the BSA + PEG group (P = 0.19). Energy charge levels of SCS controls decreased 4-fold compared to limbs perfused with acellular oxygen carrier HBOC-201, 0.10 (0.07-0.17) versus 0.46 (0.42-0.49) respectively (P = 0.002). Six hours ex vivo SNMP of rodent hindlimbs using an acellular oxygen carrier HBOC-201 results in superior tissue preservation compared to conventional SCS. [ABSTRACT FROM AUTHOR]

Background Lack of oxygen and biomechanical stimulation during static cold storage (SCS) of donor livers compromises endothelial cell function. We investigated the effect of end-ischemic oxygenated hypothermic machine perfusion (HMP) on endothelial cell function of extended criteria donor (ECD) livers. Methods Eighteen livers, declined for transplantation, were transported to our center using static cold storage (SCS). After SCS, 6 livers underwent two hours of HMP, and subsequent normothermic machine perfusion (NMP) to assess viability. Twelve control livers underwent NMP immediately after SCS. mRNA expression of transcription factor Krüppel-like-factor 2 (KLF2), endothelial nitric oxide synthase (eNOS), and thrombomodulin (TM) was quantified by RT-PCR. Endothelial cell function and injury were assessed by nitric oxide (NO) production and release of TM into the perfusate. Results In HMP livers, mRNA expression of KLF2 (p = 0.043), eNOS (p = 0.028), and TM (p = 0.028) increased significantly during NMP. In parallel, NO levels increased during NMP in HMP livers but not in controls. At the end of NMP cumulative TM release was significantly lower HMP livers, compared to controls (p = 0.028). Conclusion A short period of two hours oxygenated HMP restores endothelial cell viability after SCS and subsequent normothermic reoxygenation of ECD livers. [ABSTRACT FROM AUTHOR]