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Although machine perfusion is a hot topic today, we are just at the beginning of understanding the underlying mechanisms of protection. Recently, the first randomized controlled trial reported a significant reduction of ischemic cholangiopathies after transplantation of livers donated after circulatory death, provided the grafts were treated with an endischemic hypothermic oxygenated perfusion (HOPE). This approach has been known for more than fifty years, and was initially mainly used to preserve kidneys before implantation. Today there is an increasing interest in this and other dynamic preservation technologies and various centers have tested different approaches in clinical trials and cohort studies. Based on this, there is a need for uniform perfusion settings (perfusion route and duration), and the development of general guidelines regarding the duration of cold storage in context of the overall donor risk is also required to better compare various trial results. This article will highlight how cold perfusion protects organs mechanistically, and target such technical challenges with the perfusion setting. Finally, the options for viability testing during hypothermic perfusion will be discussed.

Simple Summary Little is known about the functions and intracellular mechanisms of the endothelial cells in liver grafts. In particular, we still know little about the effect of the most recent machine perfusion techniques currently applied to improve liver transplant outcomes. In this study we analyzed different endothelial markers of both immunohistochemical and gene expression in two different biopsies (for each donor). We observed a severe depression of endothelial trophism in liver grafts, which is restored after reperfusion. This is interesting for further studies on liver grafts, especially considering that the execution of HOPE seems to improve this functional recovery. We propose that our results may help improve the knowledge on graft tissues in order to customize the perfusion techniques prior to transplant and, therefore, improve liver transplant outcomes. The aim of the present study was to evaluate the homeostasis and trophism of liver sinusoidal endothelial cells (LSECs) in vivo in different stages of liver graft donation, in order to understand the effects of graft ischemia and perfusion on LSEC activity in liver grafts. Special attention was paid to grafts that underwent hypothermic oxygenated perfusion (HOPE). Forty-seven donors were prospectively enrolled, and two distinct biopsies were performed in each case: one allocation biopsy (at the stage of organ allocation) and one post-perfusion biopsy, performed after graft implant in the recipients. In all biopsies, immunohistochemistry and RT-PCR analyses were carried out for the endothelial markers CD34, ERG, Nestin, and VEGFR-2. We observed an increase in CD34 immunoreactivity in LSEC during the whole preservation/perfusion period (p < 0.001). Nestin and ERG expression was low in allocation biopsies, but increased in post-perfusion biopsies, in both immunohistochemistry and RT-PCR (p < 0.001). An inverse correlation was observed between ERG positivity and donor age. Our results indicate that LSEC trophism is severely depressed in liver grafts, but it is restored after reperfusion in standard conditions. The execution of HOPE seems to improve this recovery, confirming the effectiveness of this machine perfusion technique in restoring endothelial functions.

In the face of a critical organ shortage in the Western world, various strategies are employed to expand the donor pool for orthotopic liver transplantation (OLT). Among them is the transplantation of organs from extended criteria donors, a valuable source of liver allografts, however, characterized by potential risks for post-OLT complications and inferior outcomes. In recent years, machine perfusion (MP) of the explanted donor liver as well as regional perfusion techniques has witnessed significant advancements. Here, we aim to discuss different modes of dynamic organ preservation in OLT. These include hypothermic and normothermic MP, hypothermic oxygenated machine perfusion (HOPE), controlled oxygenated rewarming as well as regional perfusion protocols. Over recent years, multiple feasibility trials have demonstrated the clinical prospects of MP. In the context of OLT using organs from extended criteria donors, MP has numerous advantages compared to conventional cold storage, some of which include the preservation and reconditioning of borderline transplantable organs and the viability assessment of high-risk donor allografts. This review aims to address the topic of liver allograft MP, highlighting particularly the current trends in clinical applications and future perspectives. Furthermore, different approaches of liver storage and reconditioning are reviewed in the context of ongoing research.

Biliary complications are the most frequent cause of morbidity, re-transplantation, and even mortality after liver transplantation. In general, biliary leakage and anastomotic and non-anastomotic biliary strictures (NAS) can be recognized. There is no consensus on the exact definition of NAS and different names and criteria have been used in literature. We propose to use the term post-transplant cholangiopathy for the spectrum of abnormalities of large donor bile ducts, that includes NAS, but also intraductal casts and intrahepatic biloma formation, in the presence of a patent hepatic artery. Combinations of these manifestations of cholangiopathy are not infrequently found in the same liver and ischemia-reperfusion injury is generally considered the common underlying mechanism. Other factors that contribute to post-transplant cholangiopathy are biliary injury due to bile salt toxicity and immune-mediated injury. This review provides an overview of the various types of post-transplant cholangiopathy, the presumed pathogenesis, clinical implications, and preventive strategies.

Background Mitochondrial succinate accumulation has been suggested as key event for ischemia reperfusion injury in mice. No specific data are however available on behavior of liver mitochondria during ex situ machine perfusion in clinical transplant models. Methods We investigated mitochondrial metabolism of isolated perfused rat livers before transplantation. Livers were exposed to warm and cold ischemia to simulate donation after circulatory death (DCD) and organ transport. Subsequently, livers were perfused with oxygenated Belzer-MPS for 1h, at hypothermic or normothermic conditions. Various experiments were performed with supplemented succinate and/or mitochondrial inhibitors. The perfusate, liver tissues, and isolated mitochondria were analyzed by mass-spectroscopy and fluorimetry. Additionally, rat DCD livers were transplanted after 1h hypothermic or normothermic oxygenated perfusion. In parallel, perfusate samples were analysed during HOPE-treatment of human DCD livers before transplantation. Findings Succinate exposure during rat liver perfusion triggered a dose-dependent release of mitochondrial Flavin-Mononucleotide (FMN) and NADH in perfusates under normothermic conditions. In contrast, perfusate FMN was 3-8 fold lower under hypothermic conditions, suggesting less mitochondrial injury during cold re-oxygenation compared to normothermic conditions. HOPE-treatment induced a mitochondrial reprogramming with uploading of the nucleotide pool and effective succinate metabolism. This resulted in a clear superiority after liver transplantation compared to normothermic perfusion. Finally, the degree of mitochondrial injury during HOPE of human DCD livers, quantified by perfusate FMN and NADH, was predictive for liver function. Interpretation Mitochondrial injury determines outcome of transplanted rodent and human livers. Hypothermic oxygenated perfusion improves mitochondrial function, and allows viability assessment of liver grafts before implantation. Funding detailed information can be found in Acknowledgments., Graphical Abstract Image, graphical abstract

Donation after circulatory death (DCD) liver transplantation is known for potentially worse outcomes because of higher rates of graft non-function or irreversible cholangiopathy. The impact of machine liver perfusion techniques on these complications remains elusive. We aimed to provide data on 5-year outcomes in patients receiving DCD liver transplants, after donor organs had been treated by hypothermic oxygenated perfusion (HOPE).Fifty HOPE-treated DCD liver transplants performed in Zurich between 2012 and 3/2017 were matched with 50 primary donation after brain death (DBD) liver transplants, and with 50 untreated DCD liver transplants in Birmingham. Match factors focussed on short cold ischaemia, comparable recipient age and low recipient laboratory model for end-stage liver disease scores. Primary endpoints were post-transplant complications, and non-tumour-related patient death or graft loss.Despite extended donor warm ischaemia, HOPE-treated DCD liver transplants achieved similar overall graft survival, compared to standard DBD liver transplants. Particularly, graft loss due to any non-tumour-related causes occurred in 8% (4/50) of cases. In contrast, untreated DCD livers resulted in non-tumour-related graft failure in one-third (16/50) of cases (p = 0.005), despite significantly (p 0.001) shorter functional donor warm ischaemia. Five-year graft survival, censored for tumour death, was 94% for HOPE-treated DCD liver transplants vs. 78% in untreated DCD liver transplants (p = 0.024).The 5-year outcomes of HOPE-treated DCD liver transplants were similar to those of DBD primary transplants and superior to those of untreated DCD liver transplants, despite much higher risk. These results suggest that a simple end-ischaemic perfusion approach is very effective and may open the field for safe utilisation of extended DCD liver grafts.Machine perfusion techniques are currently being introduced into the clinic, with the aim of optimising injured grafts prior to implantation. While short-term effects of machine liver perfusion have been frequently reported in terms of hepatocellular enzyme release and early graft function, the long-term benefit on irreversible graft loss has been unclear. Herein, we report on 5-year graft survival in donation after cardiac death livers, treated either by conventional cold storage, or by 1-2 h of hypothermic oxygenated perfusion (HOPE) after cold storage. Graft loss was significantly less in HOPE-treated livers, despite longer donor warm ischaemia times. Therefore, HOPE after cold storage appears to be a simple and effective method to treat high-risk livers before implantation.

Dynamic preservation strategies such as hypothermic machine perfusion are increasingly discussed to improve liver graft quality before transplantation. This review summarizes current knowledge of this perfusion technique for liver preservation. We discuss optimization of perfusion conditions and current strategies to assess graft quality during cold perfusion. Next, we provide an overview of possible pathways of protection from ischemia-reperfusion injury. Finally, we report on recent clinical applications of human hypothermic machine liver perfusion.