Hypothermic machine perfusion in liver transplantation.

Dynamic preservation strategies are a promising option to improve graft quality before transplantation, and to extend preservation time for either logistic or treatment reasons. In contrast to normothermic oxygenated perfusion, which intends to mimic physiological conditions in the human body, with subsequent clinical application for up to 24 hrs, hypothermic perfusion is mainly used for a relatively short period with protection of mitochondria and subsequent reduction of oxidative injury upon implantation. The results from two randomized controlled trials, where recruitment has finished are expected this year. Both ex situ perfusion techniques are increasingly applied in clinical transplantation including recent reports on viability assessment, which could open the door for an increased liver utilization in the future.

['Will I receive a liver transplant in time?'; chance of survival of patients on the liver transplant waiting list]. / 'Komt mijn nieuwe lever wel op tijd?'.

OBJECTIVE: To calculate the chance of receiving a liver transplant for patients on the liver transplant waiting list in the Netherlands. DESIGN: Retrospective cohort research. METHOD: Data of all patients in the Netherlands on the waiting list for liver transplantation, from the introduction of the model of end-stage liver disease score on 16th December 2006 through to 31st December 2013 were collected. Survival analysis was computed with competing risk analyses. RESULTS: A total of 851 patients were listed, of whom 236 patients with hepatocellular carcinoma, 147 patients with primary sclerosing cholangitis, 142 patients with post-alcoholic liver disease, 93 patients with metabolic liver disease, 78 with viral hepatitis and 155 patients listed for other indications. The median waiting time till transplantation was 196 days. The chance to be transplanted at two years from listing was 65% and the risk of death was 17%. Patients with metabolic liver disease had the highest chance of undergoing liver transplantation. Patients with viral hepatitis were at highest risk of death while on the list, as well as having the lowest chance of undergoing liver transplantation. CONCLUSION: Our study shows a 65% chance of getting transplanted in time after a median waiting time of 6 months in the Netherlands. Sadly, 1 in 6 patients die before liver transplantation can be performed, with the highest risk of death occurring in patients with viral hepatitis.

Dual hypothermic oxygenated machine perfusion in liver transplants donated after circulatory death.

BACKGROUND: Experimental studies have suggested that end-ischaemic dual hypothermic oxygenated machine perfusion (DHOPE) may restore hepatocellular energy status and reduce reperfusion injury in donation after circulatory death (DCD) liver grafts. The aim of this prospective case-control study was to assess the safety and feasibility of DHOPE in DCD liver transplantation. METHODS: In consecutive DCD liver transplantations, liver grafts were treated with end-ischaemic DHOPE. Outcome was compared with that in a control group of DCD liver transplantations without DHOPE, matched for donor age, donor warm ischaemia time, and recipient Model for End-stage Liver Disease (MELD) score. All patients were followed for 1 year. RESULTS: Ten transplantations involving liver grafts treated with DHOPE were compared with 20 control procedures. There were no technical problems. All 6-month and 1-year graft and patient survival rates were 100 per cent in the DHOPE group. Six-month graft survival and 1-year graft and patient survival rates in the control group were 80, 67 and 85 per cent respectively. During DHOPE, median (i.q.r.) hepatic adenosine 5'-triphosphate (ATP) content increased 11-fold, from 6 (3-10) to 66 (42-87) µmol per g protein (P = 0·005). All DHOPE-preserved livers showed excellent early function. At 1 week after transplantation peak serum alanine aminotransferase (ALT) and bilirubin levels were twofold lower in the DHOPE group than in the control group (ALT: median 966 versus 1858 units/l respectively, P = 0·006; bilirubin: median 1·0 (i.q.r. 0·7-1·4) versus 2·6 (0·9-5·1) mg/dl, P = 0·044). None of the ten DHOPE-preserved livers required retransplantation for non-anastomotic biliary stricture, compared with five of 20 in the control group (P = 0·140). CONCLUSION: This clinical study of end-ischaemic DHOPE in DCD liver transplantation suggests that the technique restores hepatic ATP, reduces reperfusion injury, and is safe and feasible. RCTs with larger numbers of patients are warranted to assess the efficacy in reducing post-transplant biliary complications.

[Liver transplantation in paediatric patients in the Netherlands; evolution over the past two decades]. / Levertransplantaties bij kinderen in Nederland.

OBJECTIVE: To evaluate the results of the national paediatric liver transplantation programme in the University Medical Centre (UMC) Groningen in the Netherlands during the past two decades. DESIGN: Retrospective cohort study. METHOD: We analysed data from paediatric patients who underwent liver transplantation at UMC Groningen in the period 1995-2016. We compared outcomes from children who had undergone a liver transplantation in the period 1995-2005 (cohort A; n = 126) and in the period 2006-2016 (cohort B; n = 169). We performed a subanalysis in cohort B between liver transplantations with deceased donor livers (n = 132) and living donor liver transplantations (LDLT; n = 37). RESULTS: In cohort A, almost all livers came from deceased donors (99%), whereas in cohort B, 37 LDLTs (22%) were performed. The median age of recipients was significantly higher in cohort A (4.4 vs. 2.5 years; p = 0.015). Postoperative complications were comparable for both cohorts. Re-transplantations within a year after transplantation were more often performed in cohort A than in cohort B (25% vs. 12%; p = 0.004). Following LDLT, there was 2 times (5.4%) an indication for re-transplantation. In cohort B the 5-year survival rate was better than in cohort A (83 vs. 71%; p = 0.014). In cohort B, 5-year survival was higher after LDLT than after transplantation with a deceased donor liver (95 vs. 81%; p = 0.025). CONCLUSION: Outcomes after paediatric liver transplantation in the Netherlands have further improved during the past two decades. With an actuarial 5-year survival of 83% in the most recent cohort, and as high as 95% following LDLT, we can say that the UMC Groningen has a successful national paediatric liver transplant programme.

Machine Perfusion of Donor Livers for Transplantation: A Proposal for Standardized Nomenclature and Reporting Guidelines.

With increasing demand for donor organs for transplantation, machine perfusion (MP) promises to be a beneficial alternative preservation method for donor livers, particularly those considered to be of suboptimal quality, also known as extended criteria donor livers. Over the last decade, numerous studies researching MP of donor livers have been published and incredible advances have been made in both experimental and clinical research in this area. With numerous research groups working on MP, various techniques are being explored, often applying different nomenclature. The objective of this review is to catalog the differences observed in the nomenclature used in the current literature to denote various MP techniques and the manner in which methodology is reported. From this analysis, we propose a standardization of nomenclature on liver MP to maximize consistency and to enable reliable comparison and meta-analyses of studies. In addition, we propose a standardized set of guidelines for reporting the methodology of future studies on liver MP that will facilitate comparison as well as clinical implementation of liver MP procedures.

Balance between von Willebrand factor and ADAMTS13 following major partial hepatectomy.

BACKGROUND: Conventional coagulation tests are frequently prolonged after liver surgery, suggesting a postoperative bleeding tendency. At the same time, thrombotic complications following partial hepatectomy (PH) are not uncommon. Little is known about changes in the platelet adhesive protein von Willebrand factor (VWF) and its cleaving protease a disintegrin and metalloprotease with a thrombospondin type 1 motif, member 13 (ADAMTS13) following a PH. METHODS: Plasma samples were collected before and after PH and pylorus-preserving pancreaticoduodenectomy (PPPD), and from 24 healthy individuals. Plasma levels of VWF and ADAMTS13, VWF activity and VWF-dependent platelet adhesion were measured, and compared between the groups. RESULTS: Median (i.q.r.) VWF levels increased more after PH (17 patients) than following PPPD (10), reaching the highest level on postoperative day (POD) 3 (570 (473-656) versus 354 (305-476) per cent respectively; P = 0·009). VWF levels remained raised on POD 30. A decrease in median (i.q.r.) ADAMTS13 activity was observed for both patient groups, reaching the lowest level on POD 7 (24 (16-32) versus 38 (23-66) per cent for PH and PPPD respectively; P = 0·049), and levels remained significantly reduced at POD 30. VWF activity was significantly higher on day 7 following PH compared with PPPD (median (i.q.r.) 517 (440-742) versus 385 (322-484) per cent respectively; P = 0·009), and remained increased at POD 30. VWF-dependent platelet adhesion under conditions of flow was increased until POD 30 in patients after PH and PPPD, but was more pronounced in the PH group. CONCLUSION: There are changes in the balance between VWF and ADAMTS13 levels and activity in patients after both PH and PPPD. Changes in the VWF-ADAMTS13 axis were more pronounced and of longer duration after PH than following PPPD.

Procoagulant changes in fibrin clot structure in patients with cirrhosis are associated with oxidative modifications of fibrinogen.

UNLABELLED: Essentials Patients with cirrhosis have hemostatic changes, which may contribute to a risk of thrombosis. This in vitro study compares clot formation and structure between patients and healthy subjects. Clot formation is delayed in patients; ultimately, however, clot permeability is decreased. The thrombogenic structure of fibrin clots may contribute to the thrombotic risk in cirrhosis. ABSTRACT: Background and Objectives Patients with cirrhosis can be at risk of thrombotic complications due to an imbalance between hemostatic components. However, little is known on how the disease affects clot generation or how alterations in the structure of fibrin clots may affect the hemostatic function of these patients. Methods We investigated the formation and structure of clots generated with plasma and purified fibrinogen of 42 patients with cirrhosis. Clots generated with plasma and fibrinogen of 29 healthy volunteers were studied for comparison. Clot formation and structure were assessed by turbidity, permeation studies, confocal laser and scanning electron microscopy (SEM). The extent of fibrinogen oxidation was assessed by measuring the carbonyl content of purified fibrinogen samples. Results Tissue factor and thrombin-induced clotting of plasma was delayed in patients. The clotting rate was also decreased, but change in turbidity, fibrin density and fiber thickness were largely comparable to healthy volunteers. Conversely, clot permeability was significantly decreased in patients. When clots were generated with purified fibrinogen, differences in clot formation and structure similar to those in plasma were found. The carbonyl content was increased in patient fibrinogen and correlated with disease severity and clot permeability. Conclusions Delayed clot formation in cirrhosis ultimately results in decreased clot permeability. Similar alterations in clots generated with purified fibrinogen suggest that modifications of the molecule are (partly) responsible. Taken together, these findings are indicative of hypercoagulable features of clots of patients with cirrhosis, which may explain the increased risk of thrombosis associated with this condition.

Ex vivo addition of fibrinogen concentrate improves the fibrin network structure in plasma samples taken during liver transplantation.

BACKGROUND: Optimal hemostatic management during orthotopic liver transplantation (OLT) remains a challenge. The cause of bleeding during OLT is multifactorial, and may include hemostatic imbalance. Fibrinogen concentrates are increasingly being used to control perioperative bleeding during OLT. However, administration is based on arbitrary thresholds of fibrinogen levels. Importantly, studies on fibrin clot structure during OLT are lacking. OBJECTIVE: We determined the hemostatic efficacy of fibrinogen concentrate in correcting the fibrin structure. METHODS: Plasma samples taken at various times during OLT from 15 patients and 15 healthy controls were spiked with 1 g L(-1) fibrinogen concentrate or saline. Turbidity, fibrin fiber density and permeability of the fibrin clots were assessed. RESULTS: Clotting rate and turbidity were significantly decreased at the start of surgery, and decreased even further during surgery. Addition of fibrinogen significantly increased the clotting rate and turbidity at all time points, but did not normalize it. Fibrin density was significantly reduced after reperfusion as compared with the density at the start of surgery and in healthy controls. Fibrin density improved significantly after addition of fibrinogen in samples taken at the start of surgery and after reperfusion. The severely impaired polymerization and decreased density after reperfusion were accompanied by significantly increased permeability of the clot as compared with the start of surgery and in controls, which was completely restored after addition of fibrinogen. CONCLUSIONS: Ex vivo addition of fibrinogen concentrate during OLT substantially improves the structural properties of the fibrin clot, which, particularly after reperfusion, shows hypocoagulable features. These data support the use of fibrinogen concentrate to control bleeding complications during OLT.

Development of a Hyperactive Primary Hemostatic System During Off-Pump Lung Transplantation Resulting From an Unbalance Between von Willebrand Factor and Its Cleaving Protease ADAMTS13.

An unbalance between the platelet-adhesive protein von Willebrand factor (VWF) and its cleaving protease ADAMTS13 is a risk factor for thrombosis. Here, we assessed levels and functionality of VWF and ADAMTS13 in patients undergoing off-pump lung transplantation. We analyzed plasma of 10 patients and distinguished lung transplantation-specific effects from those generally accompanying open-chest surgeries by comparing results with 11 patients undergoing off-pump coronary bypass graft (CABG) surgery. Forty healthy volunteers were included for reference values. VWF antigen levels as well as the VWF ristocetin cofactor activity/VWF antigen ratio increased during lung transplantation and after CABG surgery. An increase in VWF propeptide levels was paralleled by a decrease in ADAMTS13 activity. This was more pronounced during lung transplantation. Similarly, the capacity of plasma to support platelet aggregation under shear flow conditions in vitro was more increased during lung transplantation. The proportion of high molecular weight VWF multimers was elevated in both groups without evidence for ultra-large VWF. VWF's collagen binding activity remained unchanged. In conclusion, a hyperactive primary hemostatic system develops during lung transplantation resulting both from a pronounced (functional) increase of the VWF molecule and decrease of ADAMTS13. This may increase the risk of platelet thrombosis within the allograft.

Hypercoagulability following major partial liver resection - detected by thrombomodulin-modified thrombin generation testing.

BACKGROUND: Conventional coagulation tests are frequently prolonged after liver surgery, suggesting a post-operative hypocoagulability. However, these tests are unreliable for assessment of the haemostatic status in these patients. In contrast, thrombin generation testing measures the true balance between pro- and anti-coagulant factors. AIM: To study the perioperative coagulation status in patients undergoing hemi-hepatectomy using thrombin generation assays. METHODS: We examined thrombin generation profiles in serial plasma samples taken from seventeen patients undergoing right hemi-hepatectomy. Results were compared to ten patients undergoing pancreatic resection and twenty-four healthy volunteers. In addition, we measured conventional coagulation tests and plasma levels of several haemostatic proteins. RESULTS: Following liver resection, the endogenous thrombin potential (ETP) slightly decreased until post-operative day 7. However, in the presence of thrombomodulin, the ETP increased [from 542 nM*min (417-694) at baseline to 845 nM*min (789-1050) on post-operative day 3] to values higher than that in healthy subjects (558 nM*min (390-680); P < 0.001), which contrasts with substantially prolonged PT levels. Normal to decreased thrombin generation was observed following pancreatic resection. Thrombin generation was only slightly affected by thrombomodulin after hemi-hepatectomy, while thrombin generation in healthy subjects decreased profoundly upon addition of thrombomodulin. This hypercoagulability following liver resection may be explained by decreased levels of protein C, S, and antithrombin and by elevated levels of factor VIII. CONCLUSIONS: Thrombin generation in the presence of thrombomodulin revealed hypercoagulability in patients following liver resection. These results support the recently advocated restrictive use of plasma during liver resection and the exploration of more extensive use of post-operative thrombosis prophylaxis.

Subnormothermic machine perfusion for ex vivo preservation and recovery of the human liver for transplantation.

To reduce widespread shortages, attempts are made to use more marginal livers for transplantation. Many of these grafts are discarded for fear of inferior survival rates or biliary complications. Recent advances in organ preservation have shown that ex vivo subnormothermic machine perfusion has the potential to improve preservation and recover marginal livers pretransplantation. To determine the feasibility in human livers, we assessed the effect of 3 h of oxygenated subnormothermic machine perfusion (21°C) on seven livers discarded for transplantation. Biochemical and microscopic assessment revealed minimal injury sustained during perfusion. Improved oxygen uptake (1.30 [1.11-1.94] to 6.74 [4.15-8.16] mL O2 /min kg liver), lactate levels (4.04 [3.70-5.99] to 2.29 [1.20-3.43] mmol/L) and adenosine triphosphate content (45.0 [70.6-87.5] pmol/mg preperfusion to 167.5 [151.5-237.2] pmol/mg after perfusion) were observed. Liver function, reflected by urea, albumin and bile production, was seen during perfusion. Bile production increased and the composition of bile (bile salts/phospholipid ratio, pH and bicarbonate concentration) became more favorable. In conclusion, ex vivo subnormothermic machine perfusion effectively maintains liver function with minimal injury and sustains or improves various hepatobiliary parameters postischemia.

Staging laparoscopy in patients scheduled for pancreaticoduodenectomy minimizes hospitalization in the remaining life time when metastatic carcinoma is found.

OBJECTIVE: To compare the burden of total hospitalization as a ratio of survival of staging laparoscopy versus prophylactic bypass surgery in patients with unresectable periampullary adenocarcinoma. BACKGROUND: Periampullary adenocarcinoma is an aggressive cancer with up to 35% of the patients at surgery found to be unresectable. Palliative prophylactic surgical bypass versus endoscopic stenting has been addressed by randomized controlled trials, but none reported on the burden of hospitalization. METHODS: From a prospective database all patients with periampullary adenocarcinomas with a preoperative patent biliary stent and absent gastric outlet obstruction, but found unresectable during surgery, were analysed. They underwent a staging laparoscopy only versus prophylactic palliative bypass surgery. In-hospital days of the initial admission as well as all consecutive admission days during the remaining life span were compared both in absolute numbers and as relative impact. RESULTS: The inclusion criteria were met by 205 patients. Of these 131 patients underwent a staging laparoscopy detecting metastases in 21 patients. In 184 laparotomies 54 patients underwent prophylactic palliative bypass surgery for unresectable disease. Median total in-hospital-stay in the Laparoscopy Group was 3 days versus 11 days in the Palliative Bypass Group (p = 0.0003). Patients with metastatic disease found during laparoscopy stayed 3.5% of the remaining life time in hospital vs. 10.0% (p = 0.029) in patients with metastatic disease who underwent bypass surgery. CONCLUSIONS: Staging laparoscopy and early discharge in patients with metastatic peri-ampullary carcinoma resulted in reduced hospitalization, both in absolute number of days and as a rate of survival time.

In vitro effects of proteases in human pancreatic juice on stability of liquid and carrier-bound fibrin sealants.

BACKGROUND: Fibrin sealants are used in pancreatic surgery to prevent leakage of pancreatic fluid and reduce associated complications. The efficacy of this approach is unclear. METHODS: Fibrin clots were generated in vitro from two commercially available liquid fibrin sealants (Tissucol Duo® and Evicel®) and the carrier-bound fibrin sealant Tachosil®, and exposed to normal saline or human pancreatic fluid. Stability of the sealants was assessed by release of the fibrin and collagen degradation products, D-dimer and hydroxyproline. The effect of protease inhibitors on sealant breakdown was assessed. RESULTS: Clots generated from liquid fibrin sealants degraded rapidly in pancreatic fluid, but not in normal saline. D-dimer release from fibrin clots by pancreatic fluid was approximately 1700 µg/ml after 24 h and less than 20 µg/ml by saline. Pancreatic fluid, but not normal saline, degraded both the fibrin and collagen component of Tachosil®. After 6 h, mean(s.e.m.) D-dimer levels in pancreatic fluid exposed to Tachosil® were 850(183) ng/ml, compared with 60(6) ng/ml in normal saline. The mean(s.e.m.) hydroxyproline concentration in pancreatic fluid was 497(17) µg/ml after a 24-h exposure to Tachosil®, compared with 26(12) µg/ml in normal saline. Protease inhibitors significantly inhibited breakdown of liquid sealants (D-dimer levels less than 50 µg/ml after 24 h) and Tachosil® (D-dimer release 179(12) ng/ml at 6 h; hydroxyproline release 181(29) µg/ml at 24 h). CONCLUSION: Proteases in pancreatic juice effectively degrade both liquid and carrier-bound fibrin sealants in vitro. The use of these products in pancreatic surgery with the aim of preventing leakage of pancreatic fluid is not supported by this experimental study.

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.

TAFI deficiency promotes liver damage in murine models of liver failure through defective down-regulation of hepatic inflammation.

Emerging evidence indicates that various haemostatic components can regulate the progression of liver disease. Thrombin-activatable fibrinolysis inhibitor (TAFI) possesses anti-inflammatory properties besides its anti-fibrinolytic function. Here, we investigated the contribution of TAFI to the progression of disease in murine models of chronic and acute liver failure. Chronic carbon tetrachloride (CCL4) administration induced liver damage and fibrosis both in TAFI knockout (TAFI-/-) mice and wild-type controls. Smooth muscle actin-α (α-SMA) content of liver tissue was significantly increased after 1 and 3 weeks, and pro-collagen α1 expression was significantly increased after 3 and 6 weeks in TAFI-/- mice. TAFI-/- mice showed significantly elevated levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) after 3 weeks of CCL4. Neutrophil influx was significantly increased in TAFI-/- mice after 6 weeks of CCL4. No difference in hepatic fibrin deposition between TAFI-/- and wild-types was observed. After acetaminophen intoxication, necrosis was significantly increased in TAFI-/- mice at 24 hours (h) after injection. AST and ALT levels were decreased at 2 and 6 h after acetaminophen injection in TAFI-/- mice, but were significantly higher in the TAFI-/- mice at 24 h. Similarly, hepatic fibrin deposition was decreased at 6 h in TAFI-/- mice, but was comparable to wild-types at 24 h after injection. In conclusion, TAFI deficiency results in accelerated fibrogenesis and increased liver damage in murine models of chronic and acute liver disease, which may be related to increased inflammation.