Machine Perfusion Research Articles

Perfusate Biomarkers of DCD Cardiac Graft Quality Identified With Proteomics: Studies in an Isolated Rat Heart Model.

Heart transplantation with donation after circulatory death (DCD) enhances cardiac graft availability, but exposes hearts to potentially damaging conditions, such as warm ischemia. Normothermic machine perfusion (NMP), used for graft transportation, allows biomarker determination in perfusate. Using our isolated, rat heart model of DCD, we evaluated potent. Isolated, perfused adult male Wistar rat hearts (n = 5/group) underwent different warm ischemic durations to simulate DCD, followed by reperfusion to simulate NMP. Perfusate samples were collected after 10 min reperfusion, and proteins were analyzed using mass spectrometry. Cardiac recovery was evaluated after 60 min reperfusion. The relationship between perfusate proteins and cardiac recovery was investigated. Cardiac recovery decreased with increasing ischemic duration. Principal component analysis of perfusate proteins demonstrated segregation by ischemic group. Several proteins demonstrated an On-Off pattern, and correlated with key outcome measurements. Other proteins were released by all hearts and were confirmed as predictors of cardiac recovery, for example, heat shock protein 70 and valosin-containing protein (area under the curve [AUC] = 0.962-0.968, respectively; P < 0.05 for all). Additionally, proteins such as glycogen phosphorylase, muscle associated (AUC = 0.9632; P < 0.05) showed potential as novel biomarkers for evaluating cardiac graft quality, unlike lactate release after 10 min of reperfusion (AUC = 0.60). Multiple perfusate proteins, such as heat shock protein 70, valosin-containing protein, or glycogen phosphorylase, muscle associated, released during early reperfusion are promising as biomarkers for assessing graft quality during NMP. Perfusate proteins, as biomarkers, offer the possibility of both rapid immune detection and out-of-hospital implementation, and may provide valuable information about graft quality, especially when profiled with serial sampling during NMP.

Comment on “Impact of Back-to-Base Normothermic Machine Perfusion on Complications and Costs: A Multi-Center, Real-World Risk-Matched Analysis”

Comment on “Impact of Back-to-Base Normothermic Machine Perfusion on Complications and Costs: A Multi-Center, Real-World Risk-Matched Analysis”

Liver Transplant Using Normothermic Machine Perfusion in Patients With High-Acuity Illness

This case-control study compares outcomes of normothermic machine perfusion vs static cold storage used in liver transplant for complex patients.

Current basic research in normothermic machine perfusion.

Background Normothermic machine perfusion (NMP) is gradually being introduced into clinical transplantation to improve the quality of organs and increase utilisation. This review details current understanding of the underlying mechanistic effects of NMP in the heart, lung, liver and kidney. It also considers recent advancements to extend the perfusion interval in these organs and the use of NMP to introduce novel therapeutic interventions, with a focus on organ modulation. Summary The re-establishment of circulation during NMP leads to the upregulation of inflammatory and immune mediators, similar to an ischaemia reperfusion injury (IRI) response. The level of injury is determined by the condition of the organ, but inflammation may also be exacerbated by the passenger leucocytes that emerge from the organ during perfusion. There is evidence that damaged organs can recover and that prolonged NMP may be advantageous. In the liver, successful 7 day NMP has been achieved. The delivery of therapeutic agents to an organ can aid repair and be used to modify the organ to reduce immunogenicity or change the structure of the blood group antigens to create a universal donor blood group organ. Key messages The application of NMP in organ transplantation is a growing area of research and is increasingly being used in the clinic. In the future NMP may offer the opportunity to change practice. If organs can be preserved for days on an NMP system, transplantation may become an elective rather than an emergency procedure. The ability to introduce therapies during NMP is an effective way to treat an organ and avoid the complexity of treating the recipient.

Machine Perfusion or Straight to Transplant? Predictive Value of Flavin Mononucleotide Levels in Flush Solution of Human Liver Allograft.

This study examined the predictive value of Flavin Mononucleotide (FMN) levels in the flush solution used during cold storage of donor livers on outcomes post-transplantation. Static cold storage for liver grafts induces hypoxia with subsequent impaired mitochondrial function and Flavin Mononucleotide (FMN) release upon reperfusion. This study enrolled 62 recipients who received whole liver grafts from donation after brain death (n=50) and circulatory death donors (n=12) between June 2022 and July 2023. FMN concentrations were measured in flush solutions on the back-table. ROC-curve analysis identified an FMN level cut-off for graft survival. Post-transplant outcomes were examined according to FMN levels. FMN level was significantly associated with graft survival, with an area-under-the-curve (AUC) of 0.858 (95%CI: 0.754-0.963, P<0.001), outperforming the donor risk index (AUC 0.571, 95%CI: 0.227-0.915, P=0.686). The study cohort was divided into low-FMN (<37.5ng/mL, n=40) and high-FMN groups (≥37.5ng/mL, n=22). The low-FMN group had superior one-year graft survival compared with the high-FMN group (100% vs. 77%, P=0.003). Levels of transaminases within 7 days post-transplant were significantly higher in the high-FMN group (P=0.003). The high-FMN group developed acute rejections (41% vs. 15%, P=0.023) and early allograft dysfunction (50% vs. 20%, P=0.014) more frequently. Median comprehensive complication index in the high-FMN group was significantly higher (54 [interquartile range, 40-78] vs. 42 [interquartile range, 28-52], P=0.017). The FMN level measured in donor livers' cold storage flush solution is a valid biomarker to predict post-transplant outcomes. Liver grafts with high FMN levels may benefit from machine perfusion to improve outcomes.

A reproducible extended ex-vivo normothermic machine liver perfusion protocol utilising improved nutrition and targeted vascular flows

BackgroundNormothermic machine perfusion of donor livers has become standard practice in the field of transplantation, allowing the assessment of organs and safe extension of preservation times. Alongside its clinical uses, there has been expanding interest in extended normothermic machine perfusion (eNMP) of livers as a potential vehicle for medical research. Reproducible extended normothermic machine perfusion has remained elusive due to its increased complexity and monitoring requirements. We set out to develop a reproducible protocol for the extended normothermic machine perfusion of whole human livers.MethodsHuman livers declined for transplantation were perfused using a blood-based perfusate at 36 °C using the Liver Assist device (XVIVO, Sweden), with continuous veno-venous haemofiltration in-parallel. We developed the protocol in a stepwise fashion.ResultsPerfusion techniques utilised included: targeted physiological vascular flows, phosphate replacement (to prevent hypophosphataemia), N-acetylcysteine (to prevent methaemoglobin accumulation), and the utilisation of sodium lactate as both a nutritional source and real-time measure of hepatocyte function. All five human livers perfused with the developed protocol showed preserved function with a median perfusion time of 168 h (range 120–184 h), with preserved viability throughout.ConclusionsLivers can be reproducibly perfused in excess of 120 (range 121–184) hours with evidence of preserved hepatocyte and cholangiocyte function.

Normothermic Ex Vivo Perfusion Before Transplantation of the Kidney (NEXT-Kidney): A Single-center, Nonrandomized Feasibility Study.

There is conflicting evidence regarding the efficacy of normothermic machine perfusion (NMP) in suboptimal deceased donor kidneys. We aimed to assess the feasibility and short-term efficacy of brief preimplantation NMP in circulatory death (DCD) kidneys. In this nonrandomized, single-center, prospective clinical trial, DCD kidneys underwent 1 to 3 h of NMP before implantation, aiming to achieve short ischemic times off NMP. The primary outcomes included feasibility and safety. Secondary outcomes included efficacy outcomes (delayed graft function and estimated glomerular filtration rate (eGFR) at 1, 6, and 12 mo), which were compared with the contralateral kidney that did not receive NMP. Eighteen DCD kidneys underwent NMP between 2020 and 2022, with at least 1 h completed in 16 (88.9%) of these kidneys (median 1 h); one kidney was removed after 5 min because of cannula failure and another at 54 min because of a sudden drop in blood flows. There was no episode of graft loss on the machine or postoperative vascular thromboses. All 18 kidneys were transplanted, with no cases of PNF or graft loss at 12 mo. Seventeen of the contralateral CS kidneys were transplanted. Compared with the contralateral kidneys, a lower incidence of delayed graft function (23.5% versus 64.7%; P = 0.046) was observed. There were no differences in the eGFR slopes between the two groups over time (P = 0.254). NMP is safe, feasible and efficacious in the Australian setting, with this relatively small cohort demonstrating good early outcomes compared to CS alone in our study of DCD kidneys.

Expression of sPD-L1 levels in an ex vivo liver perfusion model.

The programmed cell death protein 1 (PD-1) acts as a central inhibitory immune checkpoint receptor. The soluble form of its primary ligand, sPD-L1, was found to be elevated in serum of patients with cancer, infectious diseases and chronic inflammation. So far, the hepatic origin of sPD-L1 has received relatively little attention and is therefore subject of this study in the context of normothermic machine perfusion (NMP) of liver grafts. sPD-L1 concentrations as well as several well-established clinically relevant laboratory parameters were determined in the perfusate of 16 donor liver grafts undergoing normothermic machine perfusion (NMP) up to 30 hours (h). sPD-L1 levels continuously increased during NMP and significantly correlated with markers of hepatic synthesis (cholinesterase), acute-phase proteins (von Willebrand factor, procalcitonin, antithrombin, interleukin-6, fibrinogen) and liver decay markers (gamma-glutamyltransferase, alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase). Perfusate leukocytes were in the lower reference range, and decreased after 12h. Mean sPD-L1 levels in the perfusate correlated with donor levels of gamma-glutamyltransferase, alanine aminotransferase, creatinine and blood urea nitrogen. Our study reveals a significant increase of concentration of sPD-L1 following ischemia-reperfusion injury in a hepatic ex vivo model. sPD-L1 concentrations during NMP correlate with established acute-phase proteins and liver cell decay markers suggesting that hepatic sPD-L1 synthesis or shedding increases during the acute phase and cell decay. Furthermore, sPD-L1 correlates with established liver function and synthesis parameters as well as with donor laboratory values and might therefore be a potential biomarker for hepatic function of liver grafts.

The Interconnection Between the Liver and Other Organs: Insights for Hepatic Long–Term Normothermic Machine Perfusion

ABSTRACTNormothermic machine perfusion (NMP) is emerging as a promising technique for organ preservation, evaluation, and scientific research, offering the potential to enhance the viability and function of donor organs. To fully harness the benefits of NMP, sustained long–term NMP (LNMP) at 36°C for periods exceeding 24 h is required. However, the development of stable LNMP devices is hindered by our limited understanding of ex vivo liver physiology. This review synthesizes insights from clinical and experimental studies to highlight the knowledge gaps associated with the impact of the absence of various organs or systems on isolated livers during ex vivo perfusion. Additionally, it discusses liver injuries and adverse events documented in published liver LNMP studies, elucidating their underlying mechanisms. By analyzing these findings, the paper proposes a series of recommendations to establish a more stable LNMP platform, aiming to optimize the outcomes of liver preservation and expand the applications of NMP in clinical practice and research.

Incremental impact of organ allocation changes and machine perfusion technology on liver transplant waitlist and volumes

Incremental impact of organ allocation changes and machine perfusion technology on liver transplant waitlist and volumes

Dual hypothermic oxygenated machine perfusion of the liver reduces post transplant biliary complications - a retrospective cohort study.

Corroborating evidence for use of hypothermic oxygenated machine perfusion (HOPE) prior to orthotopic liver transplantation (OLT) suggests a beneficial effect in regards to biliary complications. Here, we aim to evaluate whether perfusion via portal vein alone (sHOPE) or via additional perfusion of the hepatic artery (dHOPE) have diverging impact on outcomes after OLT when compared to use of static cold storage (SCS). Consecutive patients undergoing OLT at Medical University of Vienna (2018 to 2023) were retrospectively analyzed. Donor organs were procured using SCS, or subjected to end-ischemic sHOPE or dHOPE. Severity of biliary complications was classified according to degree of therapeutic intervention (endoscopic retrograde cholangiopancreatography or surgical revision). 247 patients were included (69 SCS, 76 sHOPE, 102 dHOPE). Hospitalization was shorter for patients after HOPE (median in days: SCS=25 vs HOPE=20, P=0.019). Biliary complications were less frequent in patients after HOPE (SCS=37.7% vs HOPE=22.5%, P=0.015). A significantly lower incidence of surgical revisions for biliary complications was observed in the HOPE cohort (24.6% vs 11.8%, P=0.012). When evaluating outcome according to HOPE-modality, a significant reduction in biliary complications (P=0.006) and surgical revisions (P=0.002) was only observed in dHOPE patients in comparison to SCS. Further, only dHOPE was significantly associated with reduced need for surgical revision for biliary complications upon uni- and multivariable logistic regression (odds ratio=0.336, P=0.011). HOPE leads to a reduction of biliary complications and associated surgical revisions. This effect seems to be primarily associated with use of dHOPE, while both methods appear as feasible options for preconditioning of donor grafts prior to OLT.

Letter to the Editor: Does the utilization of machine perfusion require a boundary for livers donated after circulatory death?

Letter to the Editor: Does the utilization of machine perfusion require a boundary for livers donated after circulatory death?

Reply: Does the utilization of machine perfusion require a boundary for livers donated after circulatory death?

Reply: Does the utilization of machine perfusion require a boundary for livers donated after circulatory death?

Preservation of coagulation function by normothermic machine perfusion in liver transplant as evidenced by thromboelastography parameters

The use of normothermic machine perfusion over static cold storage in liver transplantation has been shown to reduce post-transplant risks of early allograft dysfunction, primary nonfunction, and ischemic cholangiopathy, and its increasing use has played a role in the expanded utilization of marginal livers. While studies have demonstrated improved clinical outcomes utilizing normothermic machine perfusion over static cold storage preservation, real-time intraoperative data reflecting the quality and viability of normothermic machine perfusion livers is limited. This retrospective, single-center study compared normothermic machine perfusion vs static cold storage livers in first-time liver transplant recipients through the evaluation of synthetic coagulation function as measured by thromboelastography and conventional coagulation testing. Secondarily, transfusion utilization between the two cohorts were reviewed. 186 liver transplant recipients receiving allografts from donors after circulatory death were included in the study, of which 99 (53%) allografts were preserved in static cold storage, and 87 (47%) allografts were placed on the TransMedics Organ Care System. Study findings showed normothermic machine perfusion livers supported with the TransMedics Organ Care System were associated with increased synthetic coagulation function and less excess fibrinolysis in the post-reperfusion period compared to static cold storage livers, and that these findings were better reflected in real-time with thromboelastography monitoring vs conventional coagulation testing. Following reperfusion, there was a significant decrease in the transfusion of blood products in the normothermic machine perfusion group compared with that in the static cold storage group. Overall, we determined that the use of intraoperative thromboelastography can provide real-time data to assess one aspect of reperfusion liver quality and viability.

Hyperspectral imaging of human liver allografts for prediction of initial graft function

PurposeIschemia reperfusion injury represents a significant yet difficult to assess risk factor for short- and long-term graft impairment in human liver transplantation (LT). As a non-invasive, non-ionizing tool, hyperspectral imaging (HSI) is capable of correlating optical properties with organ microperfusion. Hence, we here performed a study of human liver allografts assessed by HSI for microperfusion and prediction of initial graft function.MethodsImages of liver parenchyma of 37 human liver allografts were acquired at bench preparation, during normothermic machine perfusion (NMP), if applicable, and after reperfusion in the recipient. A specialized HSI acquisition software computed oxygen saturation (StO2), tissue hemoglobin indices (THI), near infrared perfusion indices (NIR), and tissue water indices (TWI). HSI parameters were analyzed for differences with regard to preservation technique, reperfusion sequence and presence of early allograft dysfunction (EAD).ResultsOrgan preservation was performed by means of NMP (n = 31) or static cold storage (SCS; n = 6). Patients’ demographics, donor characteristics, presence of EAD (NMP 36.7% vs. SCS 50%, p = 0.6582), and HSI parameters were comparable between both groups of preservation method. In organs developing EAD, NIR at 1, 2, and 4 h NMP and after reperfusion in the recipient was significantly lower (1 h NMP: 18.6 [8.6–27.6] vs. 28.3 [22.5–39.4], p = 0.0468; 2 h NMP: 19.4 [8.7–30.4] vs. 37.1 [27.5–44.6], p = 0.0011; 4 h NMP: 26.0 [6.8–37.1] vs. 40.3 [32.3–49.9], p = 0.0080; reperfusion: 13.0 [11.5–34.3] vs. 30.6 [19.3–44.0], p = 0.0212).ConclusionHSI assessment of human liver allografts is feasible during organ preservation and in the recipient. NIR during NMP and after reperfusion might predict the onset of EAD. Larger trials are warranted for assessment of this novel technique in human LT.

Ischemia‐Free Kidney Transplantation From Deceased Donor Kidneys: The First Retrospective Cohort Study

ABSTRACTBackgroundThe study aimed to evaluate the safety and effectiveness of ischemia‐free kidney transplantation (IFKT) in preventing ischemia‐reperfusion injury (IRI), a common complication associated with organ transplantation.MethodsTwelve kidneys were obtained from six donors and divided into two groups: the IFKT and the conventional kidney transplantation (CKT) group. The left kidneys underwent conventional cold perfusion and storage, while the right kidneys were procured using normothermic machine perfusion (NMP) to maintain uninterrupted blood supply. Perfusion parameters, metabolic and electrolyte parameters in the perfusate and urine, along with post‐transplant clinical data of the recipients, were collected for subsequent analysis.ResultsPerfusion flow rates varied from 100 to 300 mL/min under a pressure of 65 mmHg across different donor kidneys. Biochemical parameters such as the pH value, oxygen partial pressure, and electrolyte levels, including Na+ and K+ concentrations in the perfusate, remained stable within physiological ranges during NMP. Urine samples demonstrated significantly lower pH values, reduced Na+ concentrations, and elevated K+ concentrations compared to the perfusate, indicating effective kidney function in acid excretion, sodium retention, and potassium excretion during NMP. All patients survived, and the kidney grafts showed satisfactory function during the 12‐month follow‐up period. Clinical outcomes for recipients in the IFKT group were comparable to those in the CKT group, including similar rates of patient and graft survival at 1 year, as well as equivalent serum creatinine levels and eGFR at 1, 3, 6, and 12 post‐transplantation. Delayed graft function (DGF) occurred in one case (16.67%) from the same donor in both groups. One patient in the IFKT group experienced acute rejection, while no instances of primary graft nonfunction or other complications were observed in either group.ConclusionsOur findings indicate that the kidneys maintained in a physiological environment through NMP exhibit stable function, and post‐transplant allograft function is comparable between IFKT and CKT. The IFKT procedure demonstrates both feasibility and safety, presenting a promising approach to effectively mitigating IRI in kidney transplantation.

The Rapidly Evolving Landscape of DCD Heart Transplantation.

To summarise current international clinical outcomes from donation after circulatory death heart transplantation (DCD-HT); discuss procurement strategies, their impact on outcomes and overall organ procurement; and identify novel approaches and future areas for research in DCD-HT. Globally, DCD-HT survival outcomes (regardless of procurement strategy) are comparable to heart transplantation from brain dead donors (BDD). Experience with normothermic machine perfusion sees improvement in rates of primary graft dysfunction. Techniques have evolved to reduce cold ischaemic exposure to directly procured DCD hearts, though controlled periods of cold ischaemia can likely be tolerated. There is interest in hypothermic machine perfusion (HMP) for directly procured DCD hearts, with promising early results. Survival outcomes are firmly established to be equivalent between BDD and DCD-HT. Procurement strategy (direct procurement vs. regional perfusion) remains a source of debate. Methods to improve allograft warm ischaemic tolerance are of interest and will be key to the uptake of HMP for directly procured DCD hearts.

Cost-effectiveness of Dual Hypothermic Oxygenated Machine Perfusion Versus Static Cold Storage in DCD Liver Transplantation.

Ex situ machine perfusion of the donor liver, such as dual hypothermic oxygenated machine perfusion (DHOPE), is increasingly used in liver transplantation. Although DHOPE reduces ischemia/reperfusion-related complications after liver transplantation, data on cost-effectiveness are lacking. Our objective was to evaluate the cost-effectiveness of DHOPE in donation after circulatory death (DCD) liver transplantation. We performed an economic evaluation of DHOPE versus static cold storage (SCS) based on a multicenter randomized controlled trial in DCD liver transplantation (DHOPE-DCD trial; ClinicalTrials.gov number, NCT02584283). All patients enrolled in the 3 participating centers in the Netherlands were included. Costs related to the transplant procedure, hospital stay, readmissions, and outpatients treatments up to 1 y posttransplant were calculated. The cost for machine perfusion was calculated using 3 scenarios: (1) costs for machine perfusion, (2) machine perfusion costs plus costs for personnel, and (3) scenario 2 plus depreciation expenses for a dedicated organ perfusion room. Of 119 patients, 60 received a liver after DHOPE and 59 received a liver after SCS alone. The mean total cost per patient up to 1 y posttransplant was €126 221 for the SCS group and €110 794 for the DHOPE group. The most significant reduction occurred in intensive care costs (28.4%), followed by nonsurgical interventions (24.3%). In cost scenario 1, DHOPE was cost-effective after 1 procedure. In scenarios 2 and 3, cost-effectiveness was achieved after 25 and 30 procedures per year, respectively. Compared with conventional SCS, machine perfusion using DHOPE is cost-effective in DCD liver transplantation, reducing the total medical costs up to 1 y posttransplant.

A compact machine perfusion device for whole blood perfusion in isolated rat liver.

We established a compact machine perfusion system for whole blood perfusion of rat liver by making use of oxygenation filters as an artificial lung. Livers removed from rats were divided into Krebs-Henseleit (control), 50% blood (hemoglobin: 7g/dL), and whole blood (hemoglobin: 14g/dL) groups, then perfused (total perfusate volume: 25ml) with a small oxygenation filter at 37°C for 120min. Blood or perfusate was collected over time, and blood gas and blood cell were measured. In addition, bile volume and portal venous pressure measurements were taken. In all groups, the partial pressure of oxygen was controlled to approximately 400mmHg. Flow rates were maintained at approximately about 20-30ml/min according to liver size. Portal venous pressure was normal in the 50% blood and whole blood groups, while lower than the reference value in the Krebs-Henseleit group. Twice as much bile was produced in the 50% blood and whole blood groups relative with the Krebs-Henseleit group. We observed no differences in hemoglobin and red blood cell levels. Lactate levels were normal in the 50% blood and whole blood groups, but were elevated in the Krebs-Henseleit group. Our compact perfusion system using oxygenation filters was able to maintain rat liver function by perfusing a small amount of extracorporeal blood. This system is simple and stable, and may contribute to the future development of machine perfusion systems.

A-154 Non-standard body fluid validation of d-dimer analysis of organox liver perfusate to support clinical evaluations of donor livers prior to transplantation

Abstract Background Non-standard body fluid analysis is considered a challenge to laboratories since very few have been approved by the FDA for routine testing. As medical procedures evolve, the laboratory must balance the requirement to uphold regulatory standards while also supporting procedures that improve patient care. The LUMC Liver Transplant Team recently implemented the use of the FDA-approved OrganOx Metra normothermic machine perfusion instrument for maintaining donor livers under physiologic conditions prior to transplantation. The team mitigates risk for biliary complications associated with fibrin microthrombi by perfusing a combination of alteplase rTPA with fresh frozen plasma and monitoring the prevalence of fibrin through the release of D-dimers over the course of ex situ normothermic preservation. Liver perfusate is not an FDA-approved specimen type for the analysis of D-dimer and requires laboratory verification as a compatible specimen for analysis. Methods The D-dimer HS 500 assay was performed on an ACL TOP 750 analyzer. Carry-over studies evaluated activated partial thromboplastin time (aPTT) of a normal control (citrated plasma) specimen initially, and repeatedly in-between sets of D-dimer measurements of heparinized perfusate specimen. Recovery studies were performed by mixing liver perfusate with citrated plasma spiked with D-dimer in known ratios, allowing for the calculation of an expected value of D-dimer. Results Carry-over studies measuring aPTTs of normal plasma specimens provided %CVs of 1 and 3%. Recovery of D-dimer in seven sets of independent add-mixture series (three mixed specimen per set) was on average 105% with a range from 95 to 113% recovery. Conclusions Concern for cross-contamination of perfusate specimen containing heparin anticoagulant on analyzers used to evaluate routine coagulation testing was mitigated by the results of the carry-over study, which provided %CV within the manufacturer limit for intraday precision. Recovery studies evaluating matrix effect for the analysis of D-dimer on liver perfusate generated under the LUMC liver transplant protocol indicated that the matrix of the perfusate did not impact D-dimer analysis, as compared to the FDA-approved specimen type.