Hypothermic oxygenated machine perfusion enables 24-hour Ex situ preservation of porcine donation after circulatory death livers

Abstract

Background: End-ischemic hypothermic oxygenated machine perfusion reduces ischemia-reperfusion injury of donor livers during storage and transplantation. However, it is unknown whether this technique also supports an extension of the ex situ preservation time. We aimed to determine whether liver grafts from donation after circulatory death (DCD) porcine donors can be effectively preserved up to 24-hours using dual hypothermic oxygenated machine perfusion (DHOPE). Methods Porcine DCD livers were subjected to 2h static cold storage (SCS), followed by 2, 6, or 24 hours DHOPE (n=6 per group), using Belzer UW solution. Hepatocellular and bile duct viability were tested during 4h normothermic ex situ reperfusion with autologous blood. DCD livers preserved by 24h SCS (n=2) served as controls. Results: In all three study groups, portal venous and arterial flows remained stable during DHOPE. After normothermic reperfusion, there were no significant differences in lactate clearance, blood pH, glucose, and alanine aminotransferase levels among the DHOPE-preserved levels. All DHOPE-preserved livers produced bile and there were no significant differences in biliary pH, HCO3-, and LDH between the groups at 4h after reperfusion. Moreover, levels of malondialdehyde and HMGB-1 (markers for oxidative stress) in serum and liver parenchyma were similar for all DHOPE-preserved livers. Levels of cell-free DNA, a marker of cell death, were similar between the groups. Histological analysis of bile ducts and liver parenchyma also revealed no differences. In contrast, liver preserved by 24h SCS did not produce bile after reperfusion and turned bluish with reducing portal and arterial flows, representative for a non-viable liver. Histology revealed massive cellular necrosis. Conclusion: While 24h SCS preservation of DCD liver grafts lead to non-viable livers after warm reperfusion, DHOPE enabled successful ex situ preservation of donor livers up to 24h. If confirmed with human liver grafts, this technique can facilitate logistics of allocation and transplantation. Illustrative example of a porcine liver at the end of 24-hour DHOPE (A), and at the end of reperfusion after 24-hour DHOPE (B) or 24-hourSCS (C)