Machine perfusion in liver transplantation: recent advances and coming challenges.

PURPOSE OF REVIEW: Machine perfusion has been adopted into clinical practice in Europe since the mid-2010s and, more recently, in the United States (US) following approval of normothermic machine perfusion (NMP). We aim to review recent advances, provide discussion of potential future directions, and summarize challenges currently facing the field. RECENT FINDINGS: Both NMP and hypothermic-oxygenated perfusion (HOPE) improve overall outcomes after liver transplantation versus traditional static cold storage (SCS) and offer improved logistical flexibility. HOPE offers additional protection to the biliary system stemming from its' protection of mitochondria and lessening of ischemia-reperfusion injury. Normothermic regional perfusion (NRP) is touted to offer similar protective effects on the biliary system, though this has not been studied prospectively.The most critical question remaining is the optimal use cases for each of the three techniques (NMP, HOPE, and NRP), particularly as HOPE and NRP become more available in the US. There are additional questions regarding the most effective criteria for viability assessment and the true economic impact of these techniques. Finally, with each technique purported to allow well tolerated use of riskier grafts, there is an urgent need to define terminology for graft risk, as baseline population differences make comparison of current data challenging. SUMMARY: Machine perfusion is now widely available in all western countries and has become an essential tool in liver transplantation. Identification of the ideal technique for each graft, optimization of viability assessment, cost-effectiveness analyses, and proper definition of graft risk are the next steps to maximizing the utility of these powerful tools.

Viability assessment of the liver during ex-situ machine perfusion prior to transplantation.

PURPOSE OF REVIEW: In an attempt to reduce waiting list mortality in liver transplantation, less-than-ideal quality donor livers from extended criteria donors are increasingly accepted. Predicting the outcome of these organs remains a challenge. Machine perfusion provides the unique possibility to assess donor liver viability pretransplantation and predict postreperfusion organ function. RECENT FINDINGS: Assessing liver viability during hypothermic machine perfusion remains challenging, as the liver is not metabolically active. Nevertheless, the levels of flavin mononucleotide, transaminases, lactate dehydrogenase, glucose and pH in the perfusate have proven to be predictors of liver viability. During normothermic machine perfusion, the liver is metabolically active and in addition to the perfusate levels of pH, transaminases, glucose and lactate, the production of bile is a crucial criterion for hepatocyte viability. Cholangiocyte viability can be determined by analyzing bile composition. The differences between perfusate and bile levels of pH, bicarbonate and glucose are good predictors of freedom from ischemic cholangiopathy. SUMMARY: Although consensus is lacking regarding precise cut-off values during machine perfusion, there is general consensus on the importance of evaluating both hepatocyte and cholangiocyte compartments. The challenge is to reach consensus for increased organ utilization, while at the same time pushing the boundaries by expanding the possibilities for viability testing.

Regulations and Procurement Surgery in DCD Liver Transplantation: Expert Consensus Guidance From the International Liver Transplantation Society.

Donation after circulatory death (DCD) donors are an increasingly more common source of livers for transplantation in many parts of the world. Events that occur during DCD liver recovery have a significant impact on the success of subsequent transplantation. This working group of the International Liver Transplantation Society evaluated current evidence as well as combined experience and created this guidance on DCD liver procurement. Best practices for the recovery and transplantation of livers arising through DCD after euthanasia and organ procurement with super-rapid cold preservation and recovery as well as postmortem normothermic regional perfusion are described, as are the use of adjuncts during DCD liver procurement.

Comprehensive assessment of deceased donor kidneys with clinical characteristics, pre-implant biopsy histopathology and hypothermic mechanical perfusion parameters is highly predictive of delayed graft function.

Background: Due to the current high demand for transplant tissue, an increasing proportion of kidney donors are considered extended criteria donors, which results in a higher incidence of delayed graft function (DGF) in organ recipients. Therefore, it is important to fully investigate the risk factors of DGF, and establish a prediction system to assess donor kidney quality before transplantation.Methods: A total of 333 donation after cardiac death kidney transplant recipients were included in this retrospective study. Both univariate and multivariate analyses were used to analyze the risk factors of DGF occurrence. Receiver operating characteristic (ROC) curves were used to analyze the predictive value of variables on DGF posttransplant.Results: The donor clinical scores, kidney histopathologic Remuzzi scores and hypothermic mechanical perfusion (HMP) parameters (flow and resistance index) were all correlated. 46 recipients developed DGF postoperatively, with an incidence of 13.8% (46/333). Multivariate logistic regression analysis of the kidney transplants revealed that the independent risk factors of DGF occurrence post-transplantation included donor score (OR = 1.12, 95% CI 1.06-1.19, p < 0.001), Remuzzi score (OR = 1.21, 95% CI 1.02-1.43, p = 0.029) and acute tubular injury (ATI) score (OR = 4.72, 95% CI 2.32-9.60, p < 0.001). Prediction of DGF with ROC curve showed that the area under the curve was increased to 0.89 when all variables (donor score, Remuzzi score, ATI score and HMP resistance index) were considered together.Conclusions: Combination of donor clinical information, kidney pre-implant histopathology and HMP parameters provide a more accurate prediction of DGF occurrence post-transplantation than any of the measures alone.

The Effect of Histidine-tryptophan-ketoglutarate Solution and University of Wisconsin Solution: An Analysis of the Eurotransplant Registry.

BACKGROUND: Both University of Wisconsin (UW) and histidine-tryptophan-ketoglutarate (HTK) solutions are currently used in the Eurotransplant region for preservation of liver allografts. Previous studies on their effect have led to a lot of discussion. This study aims to compare the effect of HTK and UW on graft survival. METHODS: First liver transplantations in recipients 18 years or older from January 1, 2007, until December 31, 2016, were included. Graft survival was compared for livers preserved with HTK and UW at 30 days, 1, 3, and 5 years. Multivariable analysis of risk factors was performed and outcome was adjusted for important confounders. RESULTS: Of all 10 628 first liver transplantations, 8176 (77%) and 2452 (23%) were performed with livers preserved with HTK and UW, respectively. Kaplan-Meier curves showed significant differences in graft survival between HTK and UW at 30 days (89% vs 93%, P=<0.001), 1 year (75% vs 82%, P=<0.001), 3 years (67% vs 72%, P<0.001), and at 5 years (60% vs 67%, P<0.001). No significant differences in outcome were observed in separate analyses of Germany or non-German countries. In multivariable analysis, UW was associated with a decreased risk of graft loss at 30 days (HR 0.772, P=0.002) and at 1 year (0.847 (0.757-0.947). When adjusted for risk factors, no differences in long term outcome could be detected. CONCLUSIONS: Because the use of preservation fluids is clustered geographically, differences in outcome by preservation fluids are strongly affected by regional differences in donor and recipient characteristics. When adjusted for risk factors, no differences in graft survival exist between transplantations performed with livers preserved with either HTK or UW.

The role of corneal endothelial morphology in graft assessment and prediction of endothelial cell loss during organ culture of human donor corneas.

PURPOSE: Endothelial assessment is crucial in the release of corneas for grafting. We retrospectively analysed the role of endothelial morphology parameters in predicting endothelial cell loss during organ culture. METHODS: Human donor corneas were cultured in minimal essential medium with 2% fetal calf serum and antibiotics. Initial endothelial morphology was assessed microscopically using score parameters polymegethism (POL), pleomorphism (PLE), granulation (GRA), vacuolization (VAC), segmentation of cell membranes (SEG), Descemet's folds (DF), trypan blue-positive cells (TBPC) and endothelial cell-free areas (ECFA). Some corneas were primarily rejected based on endothelial assessment. Endothelial cell density (ECD) was assessed at the beginning (I-ECD) and end of culture. Corneas were then placed in dehydration medium (as above + 5% dextran 500). In a subgroup, ECD was reassessed after dehydration. Endothelial cell loss during culture (ECL@Culture) and culture+dehydration (ECL-Culture&Dehydration) were calculated. Data were given as mean ± SD and analysed using multiple linear and logistic regression. Odds ratios (OR) and 95% confidence intervals (CI) were calculated. RESULT: I-ECD was 2812 ± 360/mm2 (n = 2356). The decision to reject a cornea due to endothelial assessment was associated negatively with I-ECD (OR = 0.77/100 cells, CI 0.7-0.82) and positively with ECFA (OR = 2.7, CI 1.69-4.35), SEG (OR =1.3, CI 1.01-1.68) and donor age (OR = 1.26/decade, CI 1.33-1.41). ECL@Culture was 153 ± 201/mm2 (n = 1277), ECL@Culture&Dehydration was 169 ± 183/mm2 (n = 918). ECL@Culture was associated positively with donor age, I-ECD, GRA and TBPC, and negatively with PLE, and DF. ECL@Culture&Dehydration was associated positively with age, sex, initial ECD, POL, PLE, VAC and TBPC. CONCLUSION: Morphological parameters displayed associations with the exclusion of corneas from culture and with endothelial cell loss. Appropriate parameter selection for screening purposes may help improve graft quality.

Eight-Hour Continuous Normothermic Ex Vivo Kidney Perfusion Is a Safe Preservation Technique for Kidney Transplantation: A New Opportunity for the Storage, Assessment, and Repair of Kidney Grafts.

BACKGROUND: Hypothermic kidney storage causes preservation injury and is poorly tolerated by renal grafts. We investigated whether static cold storage (SCS) can be safely replaced with a novel technique of pressure-controlled normothermic ex vivo kidney perfusion (NEVKP) in heart-beating donor kidney transplantation. METHODS: Right kidneys were removed from 30 kg Yorkshire pigs in a model of heart-beating donation and either preserved in cold histidine-tryptophan-ketoglutarate solution for 8 hours (n = 5), or subjected to 8 hours of pressure-controlled NEVKP (n = 5) followed by renal heterotopic autotransplantation. RESULTS: During NEVKP, physiologic perfusion conditions were maintained with low intrarenal resistance and normal electrolyte and pH parameters. Aspartate aminotransferase and lactate dehydrogenase as injury markers were below the detectable analyzer range (<4 and <100 U/L, respectively). Perfusate lactate concentration decreased from baseline until the end of perfusion (10.38 ± 0.76 mmol/L vs 1.22 ± 0.26 mmol/L; P < 0.001). Posttransplantation, animals transplanted with NEVKP versus SCS grafts demonstrated similar serum creatinine peak levels (NEVKP, 2.0 ± 0.5 vs SCS 2.7 ± 0.7 mg/dL; P = 0.11) and creatinine clearance on day 10 (NEVKP, 65.9 ± 18.8 mL/min vs SCS 61.2 ± 15.6 mL/min; P = 0.74). After 10 days of follow-up, animals transplanted with NEVKP grafts had serum creatinine and blood urea nitrogen values comparable to their basal levels (P = 0.49 and P = 0.59), whereas animals transplanted with SCS grafts had persistently elevated serum creatinine and blood urea nitrogen when compared with basal levels (P = 0.01 and P = 0.03). CONCLUSIONS: Continuous pressure-controlled NEVKP is feasible and safe in good quality heart-beating donor kidney grafts. It maintains a physiologic environment and excellent graft function ex vivo during preservation without causing graft injury.

Efficacy assessment of cryostorants of donor hearts by ImageJ based image analysis.

AIM: Donor organ injury during cold preservation before transplantation negatively impacts graft survival. The current study was to examine available evidences for the efficacy of different cold storage solutions that are used to preserve donor hearts in vitro prior to orthotopic transplantation. METHODS: A systematic search of full-length articles published from 1980 to August 2012 was performed in PubMed and Google Scholar. Detailed searches were also made for availability of any sourceware for histopathology images of endomyocardial biopsies of stored hearts. RESULTS: Not even a single controlled trial has been published relating to this topic. However, we assessed all available literature pertaining to this topic, and performed original, simple yet innovative analyses using ImageJ, a Java based image analyses program, to show the tremendous power to objectively examine the efficacy of the storage solution. Our analysis suggest that ImageJ may be conveniently used to obtain evidences (or lack of it) of ischemic injury of donor hearts during cold storage. CONCLUSION: Even the UNOS database does not provide histopathological evidences of cardiac biopsies of orthotopically transplanted hearts. We, however, make the case of the need for image analyses and making availability of images to allow establishing evidence of the usefulness of these storage solutions. We recommend obtaining endomyocardial biopsy prior to orthotopic transplantation and create a registry of H&E stained slides. This is the only step that will direct us towards evidence based care of such highly critical patients who need the equally challenging surgical intervention of cardiac transplantation.

The value of machine perfusion biomarker concentration in DCD kidney transplantations.

BACKGROUND: Donation after cardiac death (DCD) increases the number of donor kidneys but is associated with more primary nonfunction (PNF) and delayed graft function (DGF). It has been suggested that biomarkers in the preservation solution of machine perfused kidneys may predict PNF, although evidence is lacking. METHODS: We analyzed the diagnostic accuracy of the perfusate biomarkers glutathione S-transferase, lactate dehydrogenase (LDH), heart-type fatty acid binding protein, redox-active iron, interleukin (IL)-18, and neutrophil gelatinase-associated lipocalin to predict PNF and DGF in 335 DCD kidneys preserved by hypothermic machine perfusion at our center between 1 January 1997 and 1 January 2008. The diagnostic accuracy of these biomarkers to predict PNF was evaluated with the area under the receiver operating characteristics curves. Additionally, the risk of DGF and graft failure was assessed. RESULTS: LDH and IL-18 concentrations were associated with PNF (odds ratio [95% confidence interval], 1.001 [1.000-1.002]; P=0.005 and 1.001 [1.000-1.002]; P=0.003, respectively) in a multivariate analysis; the diagnostic accuracy for PNF was "poor" for all biomarkers but increased to "fair" for redox-active iron and IL-18 in a multivariate analysis (area under the receiver operating characteristics curves, 0.701 and 0.700, respectively). LDH and IL-18 concentrations were associated with DGF; biomarker concentration was not associated with 1-year graft survival. CONCLUSIONS: The diagnostic accuracy of the perfusate biomarkers glutathione S-transferase, LDH, heart-type fatty acid binding protein, redox-active iron, IL-18, and neutrophil gelatinase-associated lipocalin to predict viability of DCD kidneys varies from "poor" to "fair". Therefore, DCD kidneys should not be discarded because of high biomarker perfusate concentration.

Graft quality assessment in kidney transplantation: not an exact science yet!

PURPOSE OF REVIEW: The enduring donor shortage necessitates the development of tools capable of objectively assessing kidney graft quality and thereby allowing the safer and wider use of expanded criteria donors and kidneys donated after cardiac death. We summarize current assessment tools available prior to procurement and during preservation. RECENT FINDINGS: Several donor risk scores, combining donor and recipient risk factors of inferior graft outcome, exist but all lack predictive power. Histological scoring of glomerulosclerosis, tubular atrophy, interstitial fibrosis, and vascular damage in pretransplantation kidney biopsies can supply reliable, reproducible data on the actual kidney state but prospective data on their use in graft assessment are lacking. Renal resistance and certain perfusate biomarker concentrations during machine perfusion are independent risk factors of delayed graft function, but neither method has sufficient predictive power to allow kidney discard. SUMMARY: Available tools for graft quality assessment have their intrinsic value but none offer the necessary power to predict graft outcome for a specific donor-recipient pair. This is probably due to the multitude of donor, preservation, and recipient factors at stake. Only combining these factors might improve prediction of graft outcome and allow safer use of expanded criteria donors and kidneys donated after cardiac death.

Assessment of preservation induced reperfusion injury via intraoperative renal transplant blood flow and endothelin concentration studies.

PURPOSE: We investigated a possible relationship between levels of the vasoconstrictive peptide endothelin and renal transplant reperfusion injury, and modified a technique for measuring renal blood flow with an ultrasonic perivascular transit time flow probe. MATERIALS AND METHODS: Renal grafts in a swine transplant model were cold flushed with either Collins-2 or University of Wisconsin solution. Renal blood flow and renal vein endothelin levels after reperfusion of transplanted grafts, as well as histological parameters within the transplanted kidney were measured. The 5-minute post-reperfusion renal blood flow was used as the baseline allograft flow. The definition of reperfusion injury was a decrease in flow from baseline with no recovery within 1 hour of reperfusion. In 9 human recipients reperfusion injury was further verified by monitoring subsequent serum creatinine, urine output, graft survival and rejection episodes. RESULTS: In the swine model and human transplant recipients no evidence of post-reperfusion ischemia was noted by histological examination, supporting that moderate to mild reperfusion injury or ischemic injury cannot be clinically determined with this method. In the swine model the decrease flow from baseline in allograft post-reperfusion renal blood flow was significantly greater in kidneys preserved in Collins'-2 than in University of Wisconsin solution (41.75 +/- 5.69 versus 11.18 +/- 13.99 ml. per minute, p = 0.005), supporting that this technique can assess mild to moderate reperfusion injury. The increase in serum endothelin in the allografts from the swine model and in humans was not significantly different from baseline. Clinically, post-reperfusion renal blood flow changes correlated well with subsequent function. The 4 patients with renal transplant reperfusion injury had significantly higher serum creatinine values and lower urine output 1 week postoperatively than 5 patients with no evidence of injury (serum creatinine: 6.75 +/- 3.03 versus 2.08 +/- 1.28 mg./dl., p = 0.015). Reperfusion injury patients had more rejections (2 versus 1) and less graft survival (75% versus 100%) at 1-year followup compared to the nonreperfusion injury patients. CONCLUSIONS: Vasoactive factors other than endothelin most likely contribute to reperfusion injury. Furthermore, the ultrasonic transit time flow probe accurately measures post-reperfusion renal blood flow and offers a practical method for assessing acute reperfusion injury, which may help to optimize immunosuppressive strategies to decrease allograft loss associated with delayed graft function.

Organ preservation.

Our clinical transplantation results have been very satisfying, with about 90% graft survival after 1 year. Currently, preservation of the liver, pancreas, and kidney, although not ideal, appears to meet all our clinical needs. Improvements in heart and lung preservation are needed and will result in increasing the number of cadaveric organs available for patients with end-stage intrathoracic organ diseases. In the future, machine perfusion may become the standard method for organ preservation for most organs because of the excellent preservation results and long-term preservation achieved.

Assessment of hepatic phagocytic activity by in vivo microscopy after liver transplantation in the rat.

Phagocytic activity of sinusoidal lining cells was studied in 32 livers of male Lewis rats by in vivo fluorescence microscopy with epiillumination. Normal livers (group 1, n = 8) were compared with orthotopic syngeneic liver grafts 90 min after reperfusion after a period of cold storage in University of Wisconsin solution for 17 hr (group 2, n = 10) or 24 hr (group 3, n = 14). After bolus injection of fluorescence-labeled latex particles (3 x 10(8)/kg; diameter = 1.1 micron), zonal distribution and kinetics of adherence of latex beads were quantified by off-line video analysis. Hepatocellular function was estimated by the rate of bile production and biliary concentrations of bile acids. In normal livers 50%, 37% and 13% of injected latex beads adhered in zones 1, 2 and 3, respectively, whereas in transplanted livers a more homogeneous distribution was found (group 2: 37%, 41%, 22%; group 3: 37%, 39%, 24%; p less than 0.01 vs. controls by analysis of variance). Kinetic analysis of phagocytic activity showed no significant difference between group 1 (88% adherence of visible latex beads 3 min after injection) and group 2 (90% adherence). However, after long-term preservation in group 3, sinusoidal adherence was significantly faster (96.4% adherence; p less than 0.001). Bile secretion in group 2 was lower than in controls and severely depressed in group 3 (group 1: 1.1 +/- 0.07 microliter/min/gm liver [mean +/- S.E.M.]; group 2: 0.8 +/- 0.07; group 3: 0.1 +/- 0.04; p less than 0.001) without significant changes in bile acid concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluation of preservation damage after porcine liver transplantation by assessment of hepatic microcirculation.

The ischemic damage following liver transplantation (LTX) is predominantly located at the endothelial cell level and is a major cause for a disturbance of microcirculation. The present study was designed to test the hypothesis that changes in the quality of organ preservation are correlated with changes in microcirculation: 16 pigs underwent LTX, preservation by Bretschneider's HTK-solution (Histidin, Tryptophan, alpha-Ketoglutarat) complemented by indomethacin (50 mumol/L). Cold ischemia times were 9 hr (n = 8) and 18 hr (n = 8), respectively. Using the H2-clearance technique, hepatic microcirculation was measured before, 30 min, and 20 hr after LTX. Normal tissue perfusion was 107 +/- 16 ml/100 g/min, at 30 min posttransplantation 91 +/- 13 ml/100 g/min in the short-term and 48 +/- 7 ml/100 g/min in the long-term preservation group. Whereas no animal of the long-term preservation group survived longer than 8 hr, all animals of the short-term preservation group survived, and tissue perfusion could be measured 20 hr postoperatively (101 +/- 19 ml/100 g/min). At 30 min postoperatively, all surviving animals had tissue perfusion rates greater than 70, and all nonsurvivors had values below 60 ml/100 g/min. We conclude therefore that the extent of decrease of microcirculation after LTX may be a useful predictor of organ function and survival.