Effects of hypothermic machine perfusion on rat liver function depending on the route of perfusion.

BACKGROUND AND METHODS: The aim of this study was to evaluate the efficacy of hypothermic machine perfusion (HMP) to preserve rat livers according to the route of perfusion, i.e., via portal vein, hepatic veins (retrograde), or hepatic artery. Livers were preserved for 24 or 48 hr by simple cold storage (SCS) or by HMP. Preservation solution was supplemented with (HMP) or without (SCS) hydroxyethyl starch. After preservation, grafts were reperfused for 2 hr with an oxygenated Krebs-Henseleit bicarbonate buffer. RESULTS: After 24 hr of preservation, total glutathione concentrations in HMP livers were similar (1287+/-37, 1418+/-118, and 1471+/-62 nmol/g in hepatic artery, portal vein, and hepatic vein HMP livers, respectively) and higher than in the SCS (833+/-118 nmol/g, P<0.05) group. These higher total glutathione values were due to higher reduced glutathione concentrations. ATP concentrations in the liver tissue were similar in HMP groups (0.75+/-0.4, 0.64+/-0.1, and 0.77+/-0.1 micromol/g in hepatic artery, portal vein, and hepatic vein HMP livers, respectively) and higher than in SCS (0.32+/-0.06 micromol/g, P<0.05). After 2 hr of normothermic reperfusion, bile production in the HMP portal and HMP retrograde groups were similar (391+/-29 ml and 372+/-25 ml) and higher than in the HMP artery or SCS groups (275+/-25 ml and 277+/-32 ml, respectively; P<0.05). Aspartate transaminase, alanine transaminase, lactate dehydrogenase, and purine nucleoside phosphorylase release into the perfusate of HMP portal and HMP retrograde perfused livers was similar and significantly lower compared to the HMP artery and SCS groups. At the end of reperfusion, no statistical differences were found for glutathione concentration and energetic reserves in the livers of each group. After 48 hr of preservation, livers from the HMP portal and HMP retrograde groups did significantly better than livers from the HMP artery or SCS groups. CONCLUSIONS: This study confirms the superiority of HMP over SCS to preserve the liver graft. It shows that retrograde perfusion is similar to PV perfusion and that perfusion by HA is less beneficial.     

Influence of flow and addition of oxygen during porcine liver hypothermic machine perfusion

INTRODUCTION: In contrast with kidneys, data on hypothermic machine perfusion (HMP) of livers remain scarce. Optimal liver HMP is poorly defined. Superiority of liver HMP over simple cold storage (SCS), the current standard preservation, must be proven before HMP is applied clinically. In this study, morphology and adenosine triphosphate (ATP) contents of HMP livers at different flows and with versus without O(2) studied in a porcine ex vivo model were compared to SCS. METHODS: Pig livers were procured, flushed with HTK and preserved via SCS or HMP at 3 HMP settings: high flow (HF); low flow (LF); low flow + O(2) (300 mm Hg) (LFO). HMP livers were perfused via the hepatic artery (HA) and portal vein (PV) with KPS-1 TM at 4 degrees C to 6 degrees C for 24 hours with HF: PV: 3 to 5 mm Hg, 1 mL/g liver/min for HA and 25 mm Hg; LF: PV: 3 to 5 mm Hg, 0.5 ml/g liver/min with HA: 20 mm Hg. Morphology and ATP levels were measured in preserved liver tissues. RESULTS: Throughout the SCS preservation, livers remained intact. In HMP livers, vacuoles appeared after 4 hours of preservation in the HF group and after 12 hours in the LF livers. LFO livers remained intact with limited vacuoles. Compared to SCS, HMP livers showed dilated sinusoids, particularly in the HF group. ATP remained relatively constant or even increased during HMP, particularly in the LF group, whereas ATP decreased after SCS. CONCLUSION: Among the various HMP settings, HMP with LFO was superior. ATP levels were the highest in LF. In contrast with all HMP groups, SCS showed the lowest ATP levels, indicating that HMP has the potential to better preserve energy stores.     

Determination of an adequate perfusion pressure for continuous dual vessel hypothermic machine perfusion of the rat liver

Hypothermic machine perfusion (HMP) provides better protection against ischemic damage of the kidney compared to cold-storage. The required perfusion pressures needed for optimal HMP of the liver are, however, unknown. Rat livers were preserved in University of Wisconsin organ preservation solution enriched with acridine orange (AO) to stain viable cells and propidium iodide (PI) to detect dead cells. Perfusion pressures of 12.5%, 25% or 50% of physiologic perfusion pressures were compared. Intravital fluorescence microscopy was used to assess liver perfusion by measuring the percentage of AO staining. After 1-h, the perfusion pressure of 12.5% revealed 72% +/- 3% perfusion of mainly the acinary zones one and two. The perfusion pressure of 25% and 50% showed complete perfusion. Furthermore, 12.5% showed 14.7 +/- 3.6, 25% showed 3.7 +/- 0.9, and 50% showed 11.2 +/- 1.4 PI positive cells. One hour was followed by another series of experiments comprising 24-h preservation. In comparison with 24-h cold-storage, HMP at 25% showed less PI positive cells and HMP at 50% showed more PI positive cells. In summary, perfusion at 25% showed complete perfusion, demonstrated by AO staining, with minimal cellular injury, shown with PI. This study indicates that fine-tuning of the perfusion pressure is crucial to balance (in)complete perfusion and endothelial injury.     

Attempt to rescue discarded human liver grafts by end ischemic hypothermic oxygenated machine perfusion

In a porcine liver transplant model, a brief period of oxygenated hypothermic machine perfusion (HMP) at the end of simple cold storage (SCS) has been shown to improve the viability of damaged liver grafts. To test the clinical validity of this strategy, we randomized SCS-discarded human liver grafts to either 4 hours of HMP (n = 13) or an additional 4 hours of SCS (n = 14). All livers were then warm reperfused to mimic ischemia-reperfusion injury ex vivo. The settings for HMP were: portal vein: 3 mm Hg, 300 mL/min and hepatic artery: 20 mm Hg, po₂: 300 mm Hg. Perfusion used Kidney Machine Perfusion Solution at 4°C to 8°C. During warm reperfusion, aspartate aminotransferase (AST) and lactate dehydrogenase (LDH) values were higher (P < .015) among the SCS versus HMP methods at all times. The AST slope was lower in HMP versus SCS (P = .01). The LDH slope tended to be lower for HMP versus SCS (P = .07). Morphological scores were not different between HMP and SCS. At the start of warm reperfusion, MAPK was lower in HMP versus SCS (P = .02). Endothelin-1 (EDN1) and ICAM-1 tended to be lower in HMP versus SCS (P = .1 and .07, respectively). No difference was noted in MAPK, EDN1, and ICAM-1 after 60 or 120 minutes of warm reperfusion. In conclusion, HMP down-regulated MAPK and tended to reduce EDN1 and ICAM-1 mRNA in human liver grafts. During warm reperfusion, HMP versus SCS livers showed reduced AST and LDH release but no morphological difference. Further optimization of liver HMP may require different timing/duration of perfusion and/or an higher perfusion temperature.     

Seventy-two-hour preservation of the canine liver by machine perfusion

The UW solution effectively preserves the dog liver for up to 48 hr by simple cold storage. This solution contains lactobionate as the primary impermeant. Another solution developed for machine perfusion of the kidney is similar to the UW solution but contains gluconate in place of lactobionate. In this study the UW gluconate solution was used for the continuous hypothermic machine perfusion of dog livers for 72 hr. Dog livers were continuously perfused at 5 degrees C through the portal vein at a pressure of 16-18 mm Hg and transplanted. Seven of 8 dogs survived for 7 or more days following orthotopic transplantation. The livers functioned as well as those preserved for 48 hr by cold storage in the UW solution as indicated by various liver-function tests. Successful machine perfusion was only achieved when the perfusate contained a high concentration of potassium (125 mM) but not with a high concentration of sodium (125 mM). This study demonstrates the feasibility of machine-perfusion preservation of the liver that yields longer preservation of equal quality compared to simple cold storage. For the development of truly long-term preservation (5 or more days) and better quality short-term preservation, machine perfusion may be the method of choice.     

Improved kidney graft function after preservation using a novel hypothermic machine perfusion device

OBJECTIVE: To study graft function and ischemia/reperfusion injury of porcine kidneys after preservation with the new Groningen Machine Perfusion (GMP) system versus static cold storage (CS). INTRODUCTION: The increasing proportion of marginal and nonheart beating donors necessitates better preservation methods to maintain adequate graft viability. Hypothermic machine preservation (HMP) is a promising alternative to static CS. We have therefore developed and tested an HMP device, which is portable and actively oxygenates the perfusate via an oxygenator. The aim of the present study was to examine the efficacy of the GMP system in a transplantation experiment. MATERIALS AND METHODS: In a porcine autotransplantation model, kidneys were retrieved and either cold stored in University of Wisconsin CS for 20 hours at 4 degrees C or subjected to HMP using University of Wisconsin machine perfusion at 4 degrees C with 2 different pressure settings: 30/20 mm Hg or 60/40 mm Hg. RESULTS: HMP at 30/20 mm Hg was found to better preserve the viability of kidneys reflected by improved cortical microcirculation, less damage to the proximal tubule, less damage mediated by reactive oxygen species, less proinflammatory cytokine expression, and better functional recovery after transplantation. However, high perfusion pressures (60/40 mm Hg) resulted in higher expression of von Willebrand factor and monocyte chemotactic peptide-1 in postpreservation biopsies and subsequent graft thrombosis in 2 kidneys. CONCLUSIONS: It is concluded that the GMP system improves kidney graft viability and perfusion pressures are critically important for outcome.     

Controlled Oxygenated Rewarming of Cold Stored Liver Grafts by Thermally Graduated Machine Perfusion Prior to Reperfusion

The quality of cold‐stored livers declines with the extension of ischemic time, increasing the risk of primary dys or nonfunction. A new concept to rescue preserved marginal liver grafts by gentle oxygenated warming‐up prior to blood reperfusion was investigated. Porcine livers were preserved by cold storage (CS) in modified HTK‐solution for 18 h. Some grafts were subsequently subjected to 90 min of controlled oxygenated rewarming (COR) by machine perfusion with gradual increase of perfusate temperature up to 20°C or simple oxygenated machine perfusion in hypothermia (HMP) or subnormothermia (SNP). Graft viability was assessed thereafter by 4 h of normothermic blood reperfusion ex vivo. Endischemic tissue energetics were significantly improved by COR or SNP and to a notably lesser extent by HMP. COR significantly reduced cellular enzyme loss, gene expression and perfusate activities of TNF‐alpha, radical mediated lipid peroxidation (LPO) and increase of portal vascular perfusion resistance upon reperfusion, while HMP or SNP were less protective. Only COR resulted in significantly more bile production than after CS. Histological injury score and caspase 3‐activation were significantly lower after COR than after CS. Oxygenated rewarming prior to reperfusion seems to be a promising technique to improve subsequent organ recovery upon reperfusion of long preserved liver grafts.     

Hypothermic machine perfusion of liver grafts for transplantation: technical development in human discard and miniature swine models

INTRODUCTION: Cold storage (CS) is the standard preservation technique for liver transplantation (LTx). Hypothermic machine perfusion (HMP) is an alternative preservation technique that provides a continuous supply of substrates and removes waste products. HMP improves early graft function in kidney transplantation, especially for marginal organs: To our knowledge there have been no reports HMP in human LTx. The aim of this study was to develop a reproducible technique for liver HMP prior to initiating a clinical trial. METHODS: For the discard protocol, between May 2001 and March 2002, 10 nontransplantable human livers were obtained. We designed a model of atraumatic, centrifugal HMP of the portal vein (PV) and hepatic artery (HA) via donor vascular conduit. Livers were perfused at 3 degrees C to 5 degrees C with Vasosol solution for 5 to 10 hours using a modified Medtronic Portable Bypass System. Perfusion variables (temp, flow, pressure) where recorded every 30 minutes. During the study, we also validated our techniques in an animal model. For the animal protocol; six swine were used as liver donors and randomized to 12 hours of CS in UW (n = 3) or 12 hours of HMP using Vasosol solution (n = 3). LTx was performed in six swine. Animals survived until postoperative day 5. RESULTS: For the discard protocol, mean HMP time was 6.7 +/- 1.8 hours. Target flow was 0.7 mL/g liver/min. PV and HA pressure ranged from 3 to 5 and 12 to 18 mm Hg, respectively. All grafts were maintained at 3 degrees C to 5 degrees C during HMP. For the animal protocol, all recipients had good liver function and survived to postoperative day 5. AST and TBili were similar between CS and HMP. CONCLUSIONS: Our method of liver HMP appears to be a safe and reliable method to preserve livers. A clinical trial is now underway to evaluate this technique in human LTx.     

Hepatic function in hypothermically stored porcine livers: comparison of hypothermic machine perfusion vs cold storage

Hypothermic machine perfusion (HMP) has a potential to relieve the current donor liver crisis by providing an improved and extended preservation method. This study examined the effect of HMP on hepatocellular functions, using a prototype liver transporter capable of preserving livers for 24 hours. Livers obtained from adult farm pigs (28 to 32 kg body weight) were divided into three groups: fresh control, HMP, and simple cold storage (n = 4 each). A 4-hour normothermic reperfusion of livers was conducted to assess hepato-metabolic and cellular functions. The hepatic transport function, as indicated by canalicular excretion of indocyanine green, was improved in the HMP group than in the SCS group. The overall tissue viability, as indicated by oxygen consumption levels, was notably improved in HMP and control livers as compared to the SCS group. Higher bile production in both the preserved groups as compared to the fresh control livers could be a result of biliary edema and leakage of plasma into the canaliculus. The hepato-cellular injury, measured by ALT, release was significantly greater in the SCS group as compared to the HMP and control groups. These findings suggest that HMP could be a better method to preserve hepatic function and overall tissue viability as compared to SCS. Improved hepatic functions are indirect indicators of superior microcirculation and sinusoidal endothelial cell functions. Further studies in progress will evaluate these functions to confirm the significance of these observations.     

Functional recovery of preserved livers following warm ischemia: improvement by machine perfusion preservation

BACKGROUND: Hypothermic machine perfusion preservation has the potential to relieve the current donor shortage problem by reclaiming and preserving marginal donor organs including those from viable non-heart-beating donors. A number of problems exist with the current machine perfusion technology for preserving livers, and much research is needed to determine the clinical impact of this technology in preserving non-heart-beating donor livers. METHODS: This study was conducted to compare the poststorage function and microcirculation of simple cold stored and machine perfusion preserved livers that had experienced 30 min of warm ischemia followed by a 10 hr preservation period. In an isolated rat liver perfusion model, lactate dehydrogenase activity, indocyanine green secretion, and portal pressure values were determined at major time points. An intravital microscopy was conducted to assess microcirculation. RESULTS: The results showed an increase in flow homogeneity of machine perfused livers, which correlated with the reduction in portal pressure when compared with simple cold storage (5.4+/-0.4 vs. 8.7+/-0.6 mm Hg). A reduction in lactate dehydrogenase levels in the perfusate (333+/-22 vs.103+/-8 U/L) and an increase in bile production of the machine perfused livers (4.9+/-0.5 vs. 33.2+/-1.7 microg/min/g liver) and indocyanine green secretion (11.7+/-1.7 vs. 21.2+/-2.1 Abs/g bile) were observed at all time points (mean+/-SE of final point given). Intravital microscopic examination indicated that large regions of non flow, as indicated by the absence of fluorescein isothiocyanate-labeled albumin, were observed in the simple cold stored tissue, whereas machine perfused liver showed increase flow homogeneity. Values of bile production, indocyanine secretion, and cellular damages were comparable with controls. Histologic examination confirmed that simple cold stored tissue displayed increased vacuolization, and machine perfused tissue showed regions of normal hepatic structure. CONCLUSION: These results suggest that machine perfusion for 10 hr improves both poststorage function and microcirculation while reducing cellular damage of liver tissue that has experienced 30 min of warm ischemia, when compared with simple cold storage. Further studies need to be conducted, but this study suggests that machine perfusion preservation has the potential to reclaim and preserve liver tissues after warm ischemic insult.     

Sequential cold storage and normothermic perfusion of the ischemic rat liver

Extending transplant criteria to include livers obtained from donors after cardiac death (DCD) could increase the liver donor pool, but conventional simple cold storage of these ischemic organs can lead to poor graft function after transplantation. Experimental normothermic machine perfusion has previously proven to be useful for the recovery and preservation of DCD livers, but it is more complicated than conventional cold storage, and, therefore, is perhaps not practical during the entire preservation period. In clinical situations, the combined use of simple cold storage and normothermic perfusion preservation of DCD livers might be more realistic, but even a brief period of cold storage prior to normothermic preservation has been suggested to have a negative impact on graft viability. In this study we show that rat livers subjected to 45 minutes of ex vivo warm ischemia followed by 2 hours of simple cold storage can be reclaimed by 4 hours of normothermic machine perfusion. These livers could be orthotopically transplanted into syngeneic recipients with 100% survival after 4 weeks (N = 10), similar to the survival of animals that received fresh livers that were stored on ice in University of Wisconsin (UW) solution for 6 hours (N = 6). On the other hand, rats that received ischemic livers preserved on ice in UW solution for 6 hours (N = 6) all died within 12 hours after transplantation. These results suggest that normothermic perfusion can be used to reclaim DCD livers subjected to an additional period of cold ischemia during hypothermic storage.     

Advantages of normothermic perfusion over cold storage in liver preservation

BACKGROUND: To minimize the ischemia-reperfusion injury that occurs to the liver with the current method of preservation and transplantation, we have used an extracorporeal circuit to preserve the liver with normothermic, oxygenated, sanguineous perfusion. In this study, we directly compared preservation by the standard method of simple cold storage in University of Wisconsin (UW) solution with preservation by perfusion. METHODS: Porcine livers were harvested from large white sows weighing between 30 and 50 kg by the standard procedure for human retrieval. The livers were preserved for 24 hr by either cold storage in UW solution (n=5) or by perfusion with oxygenated autologous blood at body temperature (n=5). The extracorporeal circuit used included a centrifugal pump, heat exchanger, and oxygenator. Both groups were then tested on the circuit for a 24 hr reperfusion phase, analyzing synthetic function, metabolic capacity, hemodynamics, markers of hepatocyte and reperfusion injury, and histology. RESULTS: Livers preserved with normothermic perfusion were significantly superior (P=0.05) to cold-stored livers in terms of bile production, factor V production, glucose metabolism, and galactose clearance. Cold-stored livers showed significantly higher levels of hepatocellular enzymes in the perfusate and were found to have significantly more damage by a blinded histological scoring system. CONCLUSIONS: Normothermic sanguineous oxygenated perfusion is a superior method of preservation compared with simple cold storage in UW solution. In addition, perfusion allows the possibility to assess viability of the graft before transplantation.     

Ex vivo machine perfusion for renal graft preservation

Kidney transplantation is the treatment of choice for end-stage renal disease. Despite its superiority over dialysis, the persisting organ shortage remains a major drawback. Additional sources to increase the donor pool are grafts recovered from extended criteria donors (ECD) and donation after circulatory death (DCD). Although transplantation of marginal grafts demonstrates promising outcomes, increased rates of primary non-function, delayed graft function, and reduced graft survival have been reported. Cold ischemic injury, caused by static cold storage is a significant risk factor for poor outcome. Machine perfusion (MP) at various temperatures bears the potential to improve organ preservation, assessment, and repair. While hypothermic machine perfusion (HMP) is well established in clinical practice, modified HMP, subnormothermic machine perfusion (SMP), and normothermic machine perfusion (NMP) are novel emerging strategies with the potential to significantly improve the outcome of marginal kidney grafts. This review summarizes findings and recent advances from pre-clinical and clinical machine perfusion studies, organized by temperature, and discusses potential future developments for graft assessment and repair.     

Hemodynamic and biochemical changes during normothermic and hypothermic sanguinous perfusion of the porcine hepatic graft

Using an ex vivo liver sanguinous perfusion system, hemodynamic and biochemical changes of the porcine livers were studied, which were preserved cold (4 degrees C) for 24 hr in University of Wisconsin solution and reperfused with normothermic (37 degrees C) (n = 8) or hypothermic (32 degrees C) (n = 8) blood for 3 hr. Six more livers were reperfused with normothermic blood (37 degrees C) immediately after procurement as controls. The total hepatic blood flow was adjusted to 1 ml/min/g liver weight, in which hepatic artery and portal vein flows were administered at a 1:2 ratio. In livers stored cold for 24 hr in UW solution and perfused normothermically, a statistically higher hepatic artery resistance was exhibited at 30 and 60 min after reperfusion (P less than 0.05), and there was lower bile output (P less than 0.05) at 90 and 120 min as compared to the controls. In livers stored cold for 24 hr in UW solution and perfused hypothermically, as compared to ones perfused normothermically, statistically higher hepatic-artery and portal-vein resistances (P less than 0.05) were observed throughout the perfusion period and 60 min after reperfusion, respectively. In addition, bile output and oxygen consumption of these livers were statistically lower than those of ones perfused normothermically (P less than 0.05). In contrast, chemistries of the perfusate of livers perfused hypothermically were comparable to ones perfused normothermically. Histologic examination of the liver perfused hypothermically demonstrated hepatic arterial and/or portal venous congestion and mild-to-moderate hemorrhage in the portal triads. This study suggests that livers preserved for a prolonged period of time demonstrate a high hepatic arterial resistance shortly after revascularization, and that recipient hypothermia after revascularization may be a risk factor for the development of hepatic arterial thrombosis following liver transplantation.     

Hypothermic Machine Preservation in Human Liver Transplantation: The First Clinical Series

Hypothermic machine perfusion (HMP) is widely used to preserve kidneys for transplantation with improved results over cold storage (CS). To date, successful transplantation of livers preserved with HMP has been reported only in animal models. In this, the first prospective liver HMP study, 20 adults received HMP‐preserved livers and were compared to a matched group transplanted with CS livers. HMP was performed for 3–7 h using centrifugal perfusion with Vasosol^® solution at 4–6°C. There were no cases of primary nonfunction in either group. Early allograft dysfunction rates were 5% in the HMP group versus 25% in controls (p = 0.08). At 12 months, there were two deaths in each group, all unrelated to preservation or graft function. There were no vascular complications in HMP livers. Two biliary complications were observed in HMP livers compared with four in the CS group. Serum injury markers were significantly lower in the HMP group. Mean hospital stay was shorter in the HMP group (10.9 ± 4.7 days vs. 15.3 ± 4.9 days in the CS group, p = 0.006). HMP of donor livers provided safe and reliable preservation in this pilot case‐controlled series. Further multicenter HMP trials are now warranted.     

Pronlonged hypothermic machine perfusion preserves hepatocellular function but potentiates endothelial cell dysfunction in rat livers

BACKGROUND: Although hypothermic machine perfusion (HMP) preservation has been shown to improve organ function and to expand the organ donor pool, problems still exist with the current HMP technology for liver preservation. The present study was conducted to investigate endothelial and hepatocellular functions following extended HMP (> r =24 hr) in rat liver model. METHODS: Following 24-hour hypothermic HMP with University of Wisconsin (UW) solution or 24-hour simple cold storage (SCS), livers were reperfused with Krebs-Henseleit buffer solution at 37 degree C for 30 minutes. Hepatocyte damage and function were assessed by measuring lactate dehydrogenase (LDH) activity, bile production, and indocyanine green (ICG) extraction. Sinusoidal endothelial cell (SEC) function and permeability were determined by hyaluronic acid (HA) uptake and multiple indicator dilution (MID) method, respectively. RESULTS: After 24-hour hypothermic preservation, HMP livers showed lower released LDH levels, higher bile flow rate, and greater hepatic ICG uptake compared with SCS livers. However, LDH levels became significantly higher in HMP than in SCS after 30 minutes of warm perfusion. The increased enzyme levels were accompanied by a significant increase in endothelial permeability to albumin and a decrease in hyaluronic acid uptake in HMP compared to SCS. Liver wet/dry weight ratio confirmed a greater edema in HMP livers than SCS livers. CONCLUSION: These results suggest that 24-hour hypothermic HMP may help preservation of hepatocyte function, but endothelial cell dysfunction during the cold preservation may play a key role in hepatocyte dysfunction and parenchymal cell death upon reperfusion.     

Ex vivo Normothermic Machine Perfusion and Viability Testing of Discarded Human Donor Livers

In contrast to traditional static cold preservation of donor livers, normothermic machine perfusion may reduce preservation injury, improve graft viability and potentially allows ex vivo assessment of graft viability before transplantation. We have studied the feasibility of normothermic machine perfusion in four discarded human donor livers. Normothermic machine perfusion consisted of pressure and temperature controlled pulsatile perfusion of the hepatic artery and continuous portal perfusion for 6 h. Two hollow fiber membrane oxygenators provided oxygenation of the perfusion fluid. Biochemical markers in the perfusion fluid reflected minimal hepatic injury and improving function. Lactate levels decreased to normal values, reflecting active metabolism by the liver (mean lactate 10.0 ± 2.3 mmol/L at 30 min to 2.3 ± 1.2 mmol/L at 6 h). Bile production was observed throughout the 6 h perfusion period (mean rate 8.16 ± 0.65 g/h after the first hour). Histological examination before and after 6 h of perfusion showed well‐preserved liver morphology without signs of additional hepatocellular ischemia, biliary injury or sinusoidal damage. In conclusion, this study shows that normothermic machine perfusion of human donor livers is technically feasible. It allows assessment of graft viability before transplantation, which opens new avenues for organ selection, therapeutic interventions and preconditioning.     

Reduced oxidative stress during acellular reperfusion of the rat liver after hypothermic oscillating perfusion.

BACKGROUND: ATP resynthesis during reperfusion after liver preservation has been shown to be well correlated with the function of transplanted grafts. Nevertheless, the advantages of a cellular energy charge loading during the preservation period are yet not fully understood. This study evaluates the effects of different nucleotide levels at the end of preservation on metabolic changes and oxidative stress during reperfusion. METHODS: Two experimental groups were chosen reflecting different energy charge states after preservation: static cold storage for 10 hr and hypothermic oxygenated oscillating perfusion for 10 hr. In both experimental groups, normothermic ex vivo acellular reperfusion over 40 min was performed. A third group consisted of nonpreserved livers similarly reperfused for 40 min. Superoxide formation was detected by the superoxide dismutase inhibitable reduction of ferricytochrome c added to the normothermic perfusate. RESULTS: Superoxide formation and lipid peroxidation malondialdehyde were significantly lower during reperfusion after the energy charge loading before reperfusion by the hypothermic oscillating perfusion technique. However, oxygen radical formation, liver cell injury (lactate dehydrogenase [LDH] release), and TNFalpha release were significantly higher in energy charge-depleted groups (nonpreserved and cold stored livers). CONCLUSIONS: Hypothermic oscillating oxygenated perfusion led to the elevated energy charge during preservation and led to reduced oxygen radical formation as well as less lipid peroxidation during reperfusion, in contrast to cold stored livers and nonpreserved livers. This suggests a correlation between the energy charge before reperfusion and oxygen radical formation as well as liver injury at reperfusion.     

Functional recovery of donation after cardiac death liver graft by continuous machine perfusion preservation in pigs

Introduction

Grafts from donation after cardiac death (DCD) will greatly contribute to the expand the donor pool. However, these grafts may require the development of the preservation methods because of primary nonfunction and severe ischemic bile duct injury.

Methods

Porcine livers were perfused with a newly developed machine perfusion (MP) system. Each system for the portal vein or the hepatic artery had a roller pump, a flow meter, and a pressure sensor. The livers were perfused with University of Wisconsin (UW)-gluconate at 4°C-6°C for 3 hours after 2 hours simple cold storage (CS). The portal vein flow rate was 0.5 mL/min/g liver (pressure, 10 mm Hg) and the hepatic artery flow rate was 0.2 mL/min/g liver (pressure, 30 mm Hg). Orthotopic liver transplantation was performed in pigs comparing Group 1 (n = 4) procured after acute hemorrhagic shock preserved by MP, Group 2 (n = 3) procured after warm ischemia time (WIT) of 30 minutes with CS preservation, and Group 3 (n = 4) procured with 30 minutes of WIT and MP preservation.

Results

Collected effluent aspartate aminotransferase (AST) and lactate dehydrogenase (LDH) levels in the perfusion solution and serum AST and LDH were significantly lower in Group 1. AST and LDH results were lower in Group 3 than Group 2. Survival rates in Groups 1 and 3 were 3/4, but 0/3 in Group 2.

Conclusion

MP preservation was a useful promising preservation mode for DCD liver grafts.     

常温机械灌注修复边缘性供肝的临床疗效：单中心报道

目的探讨常温机械灌注（normothermic machine perfusion, NMP）修复边缘性供肝的安全性和有效性。 方法2018年9月至2019年9月,使用NMP进行6例边缘性供肝体外评估和修复,供肝来自4例心死亡和2例高胆红素血症患者。记录灌注过程的灌注参数、灌注液血气分析及生化检验指标,结合供肝外观等评估边缘性供肝是否适合移植。术后随访至少3个月,记录移植后7 d内肝功能指标、生化指标、并发症发生情况等。 结果NMP期间灌注参数稳定,肝动脉灌注流量为110~334 ml/min,门静脉灌注流量为540~1 180 ml/min。灌注液pH、PO₂、PCO₂在灌注0.5 h后基本恢复正常,乳酸水平迅速下降;肝酶水平无明显升高,胆汁pH>7.5。所有供肝灌注均匀,质地柔软,均被用于移植。受者肝移植后恢复情况良好,无一例发生原发性移植肝无功能或缺血性胆道病变,至末次随访所有患者及移植物均存活。 结论本科室在临床肝移植中应用NMP技术的初步经验提示,NMP用于边缘性供肝的保存是安全、有效的。