Preliminary Experience With Hypothermic Oxygenated Machine Perfusion in an Italian Liver Transplant Center

Background

Machine perfusion is increasingly utilized in liver transplantation to face the detrimental consequences of the use of extended-criteria donors. Hypothermic oxygenated machine perfusion (HOPE) appears to be more protective relative to static cold storage. Conversely, normothermic machine perfusion (NMP) allows a better graft evaluation. We describe a pilot prospective study on machine perfusion in selected grafts.

Comparing Normothermic Machine Perfusion Preservation With Different Perfusates on Porcine Livers From Donors After Circulatory Death

The utilization of normothermic machine perfusion (NMP) may be an effective strategy to resuscitate livers from donation after circulatory death (DCD). There is no consensus regarding the efficacy of different perfusates on graft and bile duct viability. The aim of this study was to compare, in an NMP porcine DCD model, the preservation potential of three different perfusates. Twenty porcine livers with 60 min of warm ischemia were separated into four preservation groups: cold storage (CS), NMP with Steen solution (Steen; XVIVO Perfusion Inc., Denver, CO), Steen plus red blood cells (RBCs), or whole blood (WB). All livers were preserved for 10 h and reperfused to simulate transplantation for 24 h. During preservation, the NMP with Steen group presented the highest hepatocellular injury. At reperfusion, the CS group had the lowest bile production and the worst hepatocellular injury compared with all other groups, followed by NMP with Steen; the Steen plus RBC and WB groups presented the best functional and hepatocellular injury outcomes, with WB livers showing lower aspartate aminotransferase release and a trend toward better results for most parameters. Based on our results, a perfusate that contains an oxygen carrier is most effective in a model of NMP porcine DCD livers compared with Steen solution. Specifically, WB‐perfused livers showed a trend toward better outcomes compared with Steen plus RBCs.     

Impact of Temperature on Porcine Liver Machine Perfusion From Donors After Cardiac Death

Normothermic machine perfusion (NMP) has been introduced as a promising technology to preserve and possibly repair marginal liver grafts. The aim of this study was to compare the effect of temperature on the preservation of donation after cardiac death (DCD) liver grafts in an ex vivo perfusion model after NMP (38.5°C) and subnormothermic machine perfusion (SNMP, 21°C) with a control group preserved by cold storage (CS, 4°C). Fifteen porcine livers with 60 min of warm ischemia were preserved for 10 h by NMP, SNMP or CS (n = 5/group). After the preservation phase all livers were reperfused for 24 h in an isolated perfusion system with whole blood at 38.5°C to simulate transplantation. At the end of transplant simulation, the NMP group showed significantly lower hepatocellular enzyme level (AST: 277 ± 69 U/L; ALT: 22 ± 2 U/L; P < 0.03) compared to both SNMP (AST: 3243 ± 1048 U/L; ALT: 127 ± 70 U/L) and CS (AST: 3150 ± 1546 U/L; ALT: 185 ± 97 U/L). There was no significant difference between SNMP and CS. Bile production was significantly higher in the NMP group (219 ± 43 mL; P < 0.01) compared to both SNMP (49 ± 84 mL) and CS (12 ± 16 mL) with no significant difference between the latter two groups. Histologically, the NMP livers showed preserved cellular architecture compared to the SNMP and CS groups. NMP was able to recover DCD livers showing superior hepatocellular integrity, biliary function, and microcirculation compared to SNMP and CS. SNMP showed some significant benefit over CS, yet has not shown any advantage over NMP.     

Hypothermic,oxygenated perfusion (HOPE) provides cardioprotection via succinate oxidation prior to normothermic perfusion in a rat model of donation after circulatory death (DCD)

In donation after circulatory death (DCD), cardiac grafts are subjected to warm ischemia in situ, prior to a brief period of cold, static storage (CSS) at procurement, and ex situ, normothermic, machine perfusion (NMP) for transport and graft evaluation. Cold ischemia and normothermic reoxygenation during NMP could aggravate graft injury through continued accumulation and oxidation, respectively, of mitochondrial succinate, and the resultant oxidative stress. We hypothesized that replacing CSS with hypothermic, oxygenated perfusion (HOPE) could provide cardioprotection by reducing cardiac succinate levels before NMP. DCD was simulated in male Wistar rats. Following 21 minutes in situ ischemia, explanted hearts underwent 30 minutes hypothermic storage with 1 of the following: (1) CSS, (2) HOPE, (3) hypothermic deoxygenated perfusion (HNPE), or (4) HOPE + AA5 (succinate dehydrogenase inhibitor) followed by normothermic reperfusion to measure cardiac and metabolic recovery. After hypothermic storage, tissue ATP/ADP levels were higher and succinate concentration was lower in HOPE vs CSS, HNPE, and HOPE + AA5 hearts. After 60 minutes reperfusion, cardiac function was increased and cellular injury was decreased in HOPE compared with CSS, HNPE, and HOPE + AA5 hearts. HOPE provides improved cardioprotection via succinate oxidation prior to normothermic reperfusion compared with CSS, and therefore is a promising strategy for preservation of cardiac grafts obtained with DCD.     

Predictor parameters of liver viability during porcine normothermic ex situ liver perfusion in a model of liver transplantation with marginal grafts

Normothermic ex situ liver perfusion (NEsLP) offers the opportunity to assess biomarkers of graft function and injury. We investigated NEsLP parameters (biomarkers and markers) for the assessment of liver viability in a porcine transplantation model. Grafts from heart‐beating donors (HBD), and from donors with 30 minutes (donation after cardiac death [DCD]30′), 70 minutes (DCD70′), and 120 minutes (DCD120′) of warm ischemia were studied. The HBD, DCD30′, and DCD70′‐groups had 100% survival. In contrast, 70% developed primary nonfunction (PNF) and died in the DCD120′‐group. Hepatocellular function during NEsLP showed low lactate (≤1.1 mmol/L) in all the groups except the DCD120′‐group (>2 mmol/L) at 4 hours of perfusion (P = .04). The fold‐urea increase was significantly lower in the DCD120′‐group (≤0.4) compared to the other groups (≥0.65) (P = .01). As for cholangiocyte function, bile/perfusate glucose ratio was significantly lower (<0.6) in all the groups except the DCD120′‐group (≥0.9) after 3 hours of perfusion (<0.01). Bile/perfusate Na+ ratio was significantly higher (≥1.2) after 3 hours of perfusion in all the groups except for the DCD120′‐group (≤1) (P < .01). Three hours after transplantation, the DCD120′‐group had a significantly higher international normalized ratio (>5) compared to the rest of the groups (≤1.9) (P = .02). Rocuronium levels were higher at all the time‐points in the animals that developed PNF during NEsLP and after transplantation. This study demonstrates that biomarkers and markers of hepatocellular and cholangiocyte function during NEsLP correlate with the degree of ischemic injury and posttransplant function.     

Liver Transplantation Using Donation After Cardiac Death Donors: Long-Term Follow-Up from a Single Center

There is a lack of universally accepted clinical parameters to guide the utilization of donation after cardiac death (DCD) donor livers and it is unclear as to which patients would benefit most from these organs. We reviewed our experience in 141 patients who underwent liver transplantation using DCD allografts from 1993 to 2007. Patient outcomes were analyzed in comparison to a matched cohort of 282 patients who received livers from donation after brain death (DBD) donors. Patient survival was similar, but 1-, 5- and 10-year graft survival was significantly lower in DCD (69%, 56%, 44%) versus DBD (82%, 73%, 63%) subjects (p < 0.0001). Primary nonfunction and biliary complications were more common in DCD patients, accounting for 67% of early graft failures. A donor warm ischemia time >20 min, cold ischemia time >8 h and donor age >60 were associated with poorer DCD outcomes. There was a lack of survival benefit in DCD livers utilized in patients with model for end-stage liver disease (MELD) ≤30 or those not on organ-perfusion support, as graft survival was significantly lower compared to DBD patients. However, DCD and DBD subjects transplanted with MELD >30 or on organ-perfusion support had similar graft survival, suggesting a potentially greater benefit of DCD livers in critically ill patients.     

In situ normothermic perfusion of livers in controlled circulatory death donation may prevent ischemic cholangiopathy and improve graft survival

Livers from controlled donation after circulatory death (DCD) donors suffer a higher incidence of nonfunction, poor function, and ischemic cholangiopathy. In situ normothermic regional perfusion (NRP) restores a blood supply to the abdominal organs after death using an extracorporeal circulation for a limited period before organ recovery. We undertook a retrospective analysis to evaluate whether NRP was associated with improved outcomes of livers from DCD donors. NRP was performed on 70 DCD donors from whom 43 livers were transplanted. These were compared with 187 non‐NRP DCD donor livers transplanted at the same two UK centers in the same period. The use of NRP was associated with a reduction in early allograft dysfunction (12% for NRP vs. 32% for non‐NRP livers, P = .0076), 30‐day graft loss (2% NRP livers vs. 12% non‐NRP livers, P = .0559), freedom from ischemic cholangiopathy (0% vs. 27% for non‐NRP livers, P < .0001), and fewer anastomotic strictures (7% vs. 27% non‐NRP, P = .0041). After adjusting for other factors in a multivariable analysis, NRP remained significantly associated with freedom from ischemic cholangiopathy (P < .0001). These data suggest that NRP during organ recovery from DCD donors leads to superior liver outcomes compared to conventional organ recovery.     

Incidence of Ischemia Reperfusion Injury Related Biliary Complications in Liver Transplantation: Effect of Different Types of Donors

Liver transplantation from donors after circulatory death (DCD) is associated with considerable rates of primary nonfunction and ischemic-type biliary lesions. Compared with donation was after brain death (DBD), the biggest disadvantage of DCD is warm ischemia injury in the procurement stage. Donation after brain death followed by circulatory death (DBCD) is a unique practice in China. Such donors should donate according to the DCD procedure, that is, remove life support and donate after cardiac arrest. We retrospectively analyzed donor and recipient characteristics with preoperative and postoperative parameters according to 3 donation types to comprehensively describe incidence of ischemia reperfusion injury (IRI) related biliary complications among different donor type adult liver transplantation recipients. A total of 50 patients were included in this study (DBD group n = 17, DCD group n = 26, DBCD group n = 7). Only 1 patient, whose donor type was DBCD was diagnosed with ischemic-type biliary lesions demonstrated cast and retrograde ascending cholangitis. Rates of primary graft non-function (DBD n = 1, 5.9%; DCD n = 2, 7.7%; DBCD, 0%; P = .546) were similar and total biliary complications (DBD n = 1, 5.9%; DCD n = 1, 3.8%; DBCD N = 2, 28.6%; P = .042) were different. No differences were found regarding development of postreperfusion syndrome or coagulopathy in 3 groups. Compared with standard DBD donor, the clinical outcome of DCD donor liver transplantation was satisfactory, with no increase in the incidence of IRI, and, no difference in the incidence of ischemic bile duct complications. This work was carried out in compliance with the Helsinki Congress and the Declaration of Istanbul.     

Model for early allograft function is predictive of early graft loss in donation after circulatory death liver transplantation

Donation after circulatory death (DCD) liver transplantation is associated with higher rates of graft loss. In this paper, we explored whether the Model for Early Allograft Function (MEAF) predicted outcome in DCD liver transplantation. We performed a retrospective analysis of prospectively collected data from all adult DCD (Maastricht 3) livers transplanted in Cambridge and Edinburgh between 1 January 2011 and 30 June 2017, excluding those undergoing any form of machine perfusion. 187 DCD liver transplants were performed during the study period. DCD liver transplants with a lower MEAF score had a significantly better survival compared to those with a high MEAF score (Mantel-Cox P < .0001); this was largely due to early graft loss. Beyond 28 days post-transplant, there were no significant long-term graft or patient survival differences irrespective of the grade of MEAF (Mantel-Cox P = .64 and P = .43, respectively). The MEAF score correlated with the length of ICU (P = .0011) and hospital stay (P = .0007), but did not predict the requirement for retransplantation for ischemic cholangiopathy (P = .37) or readmission (P = .74). In this study, a high MEAF score predicted early graft loss, but not the subsequent need for re-transplantation or late graft failure as a result of intrahepatic ischemic bile duct pathology.     

Ultrasound-Targeted Microbubble Cavitation During Machine Perfusion Reduces Microvascular Thrombi and Graft Injury in a Rat Liver Model of Donation After Circulatory Death

BackgroundIschemic cholangiopathy is a process of bile duct injury that might result from peribiliary vascular plexus (PBP) thrombosis and remains a dreaded complication in liver transplantation from donors after circulatory death (DCD). The aim of this study was to propose a mechanical method of clot destruction to clear microvascular thrombi in DCD livers before transplantation.MethodsSonothrombolysis (STL) is a process by which inertial cavitation of circulating microbubbles entering an ultrasound field create a high-energy shockwave at a microbubble-thrombus interface, causing mechanical clot destruction. The effectiveness of STL in DCD liver treatment remains unclear. We carried out STL treatment during normothermic, oxygenated, ex vivo machine perfusion (NMP), introducing microbubbles into the perfusate with the liver enveloped in an ultrasound field.ResultsThe STL livers showed reduction in hepatic arterial and PBP thrombus and decreases in hepatic arterial and portal venous flow resistance, reduced parenchymal injury as measured by aspartate transaminase release and oxygen consumption, and improved cholangiocyte function. Light and electron microscopy showed reduction of hepatic arterial and PBP thrombus in STL livers compared with controls and preserved hepatocyte structure, sinusoid endothelial morphology, and biliary epithelial microvilli.ConclusionIn this model, STL improved flow and functional measures in DCD livers undergoing NMP. These data suggest a novel therapeutic approach to treat PBP injury in DCD livers, which may ultimately increase the pool of grafts available to patients awaiting liver transplantation.     

An experimental study of the recovery of injured porcine lungs with prolonged normothermic cellular ex vivo lung perfusion following donation after circulatory death

Donation after circulatory death (DCD) is an underused source of donor lungs. Normothermic cellular ex vivo lung perfusion (EVLP) is effective in preserving standard donor lungs but may also be useful in the preservation and assessment of DCD lungs. Using a model of DCD and prolonged EVLP, the effects of donor warm ischemia and postmortem ventilation on graft recovery were evaluated. Adult male swine underwent general anesthesia and heparinization. In the control group (n = 4), cardioplegic arrest was induced and the lungs were procured immediately. In the four treatment groups, a period of agonal hypoxia was followed by either 1 h of warm ischemia with (n = 4) or without (n = 4) ventilation or 2 h of warm ischemia with (n = 4) or without (n = 4) ventilation. All lungs were studied on an EVLP platform for 24 h. Hemodynamic measures, compliance, and oxygenation on EVLP were worse in all DCD lungs compared with controls. Hemodynamics and compliance normalized in all lungs after 24 h of EVLP, but DCD lungs demonstrated impaired oxygenation. Normothermic cellular EVLP is effective in preserving and monitoring of DCD lungs. Early donor postmortem ventilation and timely procurement lead to improved graft function.     

常温机械灌注对大鼠心脏死亡器官捐献供肝微循环保护作用的研究

目的探讨常温机械灌注(normothermic machine perfusion,NMP)对大鼠心脏死亡器官捐献(donation after circulatory death,DCD)供肝微循环发挥的作用。方法采用夹闭SD大鼠胸主动脉热缺血30 min获取DCD供肝;在体外建立大鼠NMP系统。根据不同的保存供肝方式,将实验分为:正常(Normal)组(n=6),留取血清及肝脏待用;NMP组(n=30),保存4、6和8 h后收集肝脏标本,灌注后0、2、4、6 h和8 h收集流入道、流出道灌注液待检;静态冷保存(SCS)组(n=6),肝脏以20 ml 4℃UW液冲出肝内血液,并于UW液中4℃SCS 6 h后收集肝脏标本。采用生物化学方法检测流出道灌注液的肝功能;HE染色观察肝组织病理学改变;透射电镜观察肝细胞超微结构;TUNEL检测肝脏细胞凋亡情况;Western blot检测肝脏组织内皮素-1(endothelin-1,ET-1)、内皮型一氧化氮合酶(endothelial nitric oxide synthase,eNOS)、诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)、血管性血友病因子(von Willebrand factor,vWF)、细胞间黏附分子(intercellular adhesion molecule-1,ICAM-1)和血管间黏附分子-1(intervascular adhesion molecule-1,VCAM-1)的表达情况。结果与SCS比较,NMP能显著改善DCD供肝组织学损伤,Suzuki评分NMP组(3.40±0.55)显著低于SCS组(7.00±0.71,F=229.75,P<0.05);减轻肝细胞凋亡,NMP组凋亡细胞数(9.80±1.48)显著低于SCS组(33.40±4.39,F=166.58,P<0.05);同时能修复肝细胞线粒体损伤,NMP组不可逆损伤线粒体数量(1.60±0.55)显著低于SCS组(2.80±0.45,F=36.29,P<0.05)。进一步发现,NMP可改善DCD肝脏微循环:①抑制细胞间黏附,改善内皮细胞损伤,与SCS比较,NMP显著抑制肝内ICAM-1(F=1728.45,P<0.05)、VCAM-1(F=254.72,P<0.05)和vWF(F=595.30,P<0.05)的表达;②改善肝脏ET-1/NOS平衡和微循环灌注,与SCS相比,NMP显著抑制肝内ET-1(F=1372.51,P<0.05)、iNOS(F=1102.20,P<0.05)的表达,促进eNOS(F=271.66,P<0.05)的表达。结论NMP能够改善大鼠DCD供肝质量,其机制可能通过抑制细胞间黏附,改善窦内皮损伤及微循环灌注发挥保护作用的。     

NRF2 assessment in discarded liver allografts: A role in allograft function and salvage

Antioxidant defence mechanisms, such as the nuclear factor-erythroid 2-related-factor-2 (NRF2) axis, are integral to oxidative stress responses and ischemic injury. Hepatic antioxidant capacity is contingent on parenchymal quality, and there is a need to develop new insights into key molecular mechanisms in marginal liver allografts that might provide therapeutic targets. This study examines the clinical relevance of NRF2 in donor livers and its response to normothermic machine perfusion (NMP). Discarded donor livers (n = 40) were stratified into a high NRF2 and low NRF2 group by quantifying NRF2 expression. High NRF2 livers had significantly lower transaminase levels, hepatic vascular inflammation and peri-portal CD3⁺ T cell infiltration. Human liver allografts (n = 8) were then exposed to 6-h of NMP and high NRF2 livers had significantly reduced liver enzyme alterations and improved lactate clearance. To investigate these findings further, we used a rat fatty-liver model, treating livers with an NRF2 agonist during NMP. Treated livers had increased NRF2 expression and reduced transaminase derangements following NMP compared to vehicle control. These results support the association of elevated NRF2 expression with improved liver function. Targeting this axis could have a rationale in future studies and NRF2 agonists may represent a supplemental treatment strategy for rescuing marginal donor livers.     

Improved Preservation of Warm Ischemic Livers by Hypothermic Machine Perfusion with Supplemented University of Wisconsin Solution

Hypothermic machine perfusion (HMP) has the potential to improve recovery and preservation of Donation after Cardiac Death (DCD) livers, including uncontrolled DCD livers. However, current perfusion solutions lack the needed substrates to improve energy recovery and minimize hepatic injury, if warm ischemic time (WIT) is extended. This proof-of-concept study tested the hypothesis that the University of Wisconsin (UW) solution supplemented with anaplerotic substrates, calcium chloride, thromboxane A₂ inhibitor, and antioxidants could improve HMP preservation and minimize reperfusion injury of warm ischemic livers. Preflushed rat livers subjected to 60 min WIT were preserved for 5 h with standard UW or supplemented UW (SUW) solution. Post preservation hepatic functions and viability were assessed during isolated perfusion with Krebs–Henseleit solution. Livers preserved with SUW showed significantly (p <. 001) improved recovery of tissue ATP levels (μ mol/g liver), 2.06 ± 0.10 (mean ± SE), as compared to the UW group, 0.70 ± 0.10, and the level was 80% of that of fresh control livers (2.60 ± 0.13). At the end of 1 h of rewarming, lactate dehydrogenase (U/L) in the perfusate was significantly (p <. 05) lower in the SUW group (429 ± 58) as compared to ischemia–reperfusion (IR) (781 ± 12) and the UW group (1151 ± 83). Bile production (μ g/min/g liver) was significantly (p <. 05) higher in the SUW group (280 ± 13) as compared to the IR (224 ± 24) and the UW group (114 ± 14). The tissue edema formation assessed by tissue wet–dry ratio was significantly (p <. 05) higher in UW group. Histology showed well-preserved hepatic structure in the SUW group. In conclusion, this study suggests that HMP with SUW solution has the potential to restore and preserve livers with extended WIT.     

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.     

Pretransplant sequential hypo‐ and normothermic machine perfusion of suboptimal livers donated after circulatory death using a hemoglobin‐based oxygen carrier perfusion solution

Ex situ dual hypothermic oxygenated machine perfusion (DHOPE) and normothermic machine perfusion (NMP) of donor livers may have a complementary effect when applied sequentially. While DHOPE resuscitates the mitochondria and increases hepatic adenosine triphosphate (ATP) content, NMP enables hepatobiliary viability assessment prior to transplantation. In contrast to DHOPE, NMP requires a perfusion solution with an oxygen carrier, for which red blood cells (RBC) have been used in most series. RBC, however, have limitations and cannot be used cold. We, therefore, established a protocol of sequential DHOPE, controlled oxygenated rewarming (COR), and NMP using a new hemoglobin‐based oxygen carrier (HBOC)‐based perfusion fluid (DHOPE‐COR‐NMP trial, NTR5972). Seven livers from donation after circulatory death (DCD) donors, which were initially declined for transplantation nationwide, underwent DHOPE‐COR‐NMP. Livers were considered transplantable if perfusate pH and lactate normalized, bile production was ≥10 mL and biliary pH > 7.45 within 150 minutes of NMP. Based on these criteria five livers were transplanted. The primary endpoint, 3‐month graft survival, was a 100%. In conclusion, sequential DHOPE‐COR‐NMP using an HBOC‐based perfusion fluid offers a novel method of liver machine perfusion for combined resuscitation and viability testing of suboptimal livers prior to transplantation.     

Reduction of hepatic ischemia/reperfusion injury by a soluble P-selectin glycoprotein ligand-1.

OBJECTIVE: The authors' goal was to determine the effects of specific binding and blockade of P- and E-selectins by a soluble P-selectin glycoprotein ligand-1 (PSGL-1) in rat models of hepatic in vivo warm ischemia and ex vivo cold ischemia. The authors also sought to determine the effect of selectin blockade on isograft survival in a syngeneic rat orthotopic liver transplant model. SUMMARY BACKGROUND DATA: Ischemia/reperfusion (I/R) injury is a major factor in poor graft function after liver transplantation, which may profoundly influence early graft function and late changes. It is hypothesized that I/R injury leads to the upregulation of P-selectin, which is then rapidly translocated to endothelial cell surfaces within 5 minutes of reperfusion of the liver, initiating steps leading to tethering of polymorphonuclear neutrophil leukocytes to the vascular intima. Local production by leukocytes of interleukin-1, tumor necrosis factor-alpha, or both induces P-selectin expression on the endothelium and continues the cascade of events, which increases cell adherence and infiltration of the organ. METHODS: To examine directly the effects of selectins in a warm hepatic I/R injury model, 100 microg of PSGL-1 or saline was given through the portal vein at the time of total hepatic inflow occlusion. The effects of PSGL-1 in cold ischemia were assessed using an isolated perfused rat liver after 6 hours of 4 degrees C storage in University of Wisconsin (UW) solution, with or without the instillation of PSGL-1 before the storage. To evaluate the effect of selectin blockade on liver transplant survival, syngeneic orthotopic liver transplants were performed between inbred male Sprague-Dawley rats after 24 hours of cold ischemic storage in UW solution. A separate group of animals received two doses of 100 microg of PSGL-1 through the portal vein before storage and before reperfusion of the transplanted liver. Recipient survival was assessed at 7 days, and the Kaplan-Meier product limit estimate method was used for univariate calculations of time-dependent recipient survival events. RESULTS: In an in vivo warm rat liver ischemia model, perfusion with PSGL-1 afforded considerable protection from I/R injury, as demonstrated by decreased transaminase release, reduced histologic hepatocyte damage, and suppressed neutrophil infiltration, versus controls (p < 0.05). When cold stored livers were reperfused, PSGL-1 reduced the degree of hepatocyte transaminase release, reduced neutrophil infiltration, and decreased histologic hepatocyte damage (p < 0.05 vs. UW-only controls). On reperfusion, livers treated with PSGL-1 demonstrated increased portal vein blood flow and bile production (p < 0.05 vs. UW-only controls). In addition, 90% of the rats receiving liver isografts stored in UW solution supplemented with PSGL-1 survived 7 days versus 50% of those whose transplanted syngeneic livers had been stored in UW alone (p < 0.05). CONCLUSIONS: Selectins play an important role in I/R injury of the liver. Early modulation of the interaction between P-selectin and its ligand decreases hepatocyte injury, neutrophil adhesion, and subsequent migration in both warm and cold rat liver ischemia models. In addition, the use of PSGL-1 before ischemic storage and before transplantation prevents hepatic injury, as documented by a significant increase in liver isograft survival. These findings have important clinical ramifications: early inhibition of alloantigen-independent mechanisms during the I/R damage may influence both short- and long-term survival of liver allografts.     

N-acetylcysteine in pig liver transplantation from non-heart-beating donors.

Lipid peroxidation due to oxygen free radicals (OFR) seems to play a major role in loss of liver graft viability after warm ischemia, preservation, and transplantation. N-acetylcysteine (NAC) is an antioxidant that has a direct effect on OFR, and is also a glutathione precursor, another antioxidant. This study was designed to evaluate the efficacy of NAC in preventing ischemia-reperfusion damage of liver grafts harvested from non-heart-beating donors. Liver transplantation was performed on pigs divided into five groups: group 1 (control group; n=5) received livers from heart-beating donors; livers were subjected to 30 min of warm ischemia in groups 2 (n=3, no NAC) and group 3 (n=3; NAC treatment); warm ischemia time lasted 60 min in groups 4 (n=4; no NAC) and 5 (n=5; NAC treatment). Studied parameters included graft survival for more than 3 days, aspartate aminotransferase plasma levels, liver histology, and hepatic total glutathione concentrations. Graft survival was 100% in groups 1, 2, and 3, 0% in group 4, and 20% in group 5. NAC treatment did not influence initial mean aspartate aminotransferase release which was greater in warm ischemic livers than in controls. NAC treatment had no effect on liver hepatic total glutathione after reperfusion of animals receiving warm ischemic grants. Finally, no effect on liver histology was observed with NAC treatment. Our study suggests that in liver transplantation from non-heart-beating donors, NAC has no effect in both graft viability and lipid peroxidation. The role of OFR in primary dysfunction of transplanted warm ischemic livers remains controversial.     

Assessing warm ischemic injury of pig livers at hypothermic machine perfusion

Background

Livers originating from donation after circulatory death (DCD) donors are exposed to warm ischemia (WI) before liver transplantation (LTx). Currently, there are no objective tests to evaluate the damage sustained before LTx. This study aims to identify surrogate markers for liver injury that can be assessed during hypothermic machine perfusion (HMP) preservation. In addition, we want to use mathematical equation modeling combining these markers to improve our assessment of DCD livers for transplantation.

Materials and methods

Porcine livers were exposed to incremental periods of WI (0–120 min) and subsequently HMP preserved for 4 h. Biochemical and hemodynamic parameters were repeatedly measured in the perfusate during HMP. Subsequently, to mimic LTx, normothermic isolated-liver perfusion was applied for 2 h and the injury assessed using a morphological score.

Results

With increasing WI periods, the perfusate became more acidotic, and levels of aspartate aminotransferase (AST), liver fatty acid binding protein, redox-active iron, and arterial vascular resistance increased. A damage index, combining AST and pH (damage index = 2 − 37 × β_AST − 257 × β_pH) based on multifactorial analysis of the changing pattern of these markers, had increased sensitivity and specificity to reflect WI and reperfusion injury.

Conclusions

This proof of concept study demonstrated the potential role for objective evaluation of DCD porcine livers during HMP and the advantage to use multifactorial analysis on the markers' changing pattern.     

The price of donation after cardiac death in liver transplantation: a prospective cost‐effectiveness study

This study aims to perform a detailed prospective observational multicenter cost‐effectiveness study by comparing liver transplantations with Donation after Brain Death (DBD) and Donation after Cardiac Death (DCD) grafts. All liver transplantations in the three Dutch liver transplant centers between 2004 and 2009 were included with 1‐year follow‐up. Primary outcome parameter was cost per life year after transplantation. Secondary outcome parameters were 1‐year patient and graft survival, complications, and patient‐level costs. From 382 recipients that underwent 423 liver transplantations, 293 were primarily transplanted with DBD and 89 with DCD organs. Baseline characteristics were not different between both groups. The Donor Risk Index was significantly different as were cold and warm ischemic time. Ward stay was significantly longer in DCD transplantations. Patient and graft survival were not significantly different. Patients receiving DCD organs had more and more severe complications. The cost per life year for DBD was € 88 913 compared to € 112 376 for DCD. This difference was statistically significant. DCD livers have more and more severe complications, more reinterventions and consequently higher costs than DBD livers. However, patient and graft survival was not different in this study. Reimbursement should be differentiated to better accommodate DCD transplantations.