Prolonged dialysis during ex vivo lung perfusion promotes inflammatory responses.

Ex-vivo lung perfusion (EVLP) has extended the number of transplantable lungs by reconditioning marginal organs. However, EVLP is performed at 37°C without homeostatic regulation leading to metabolic wastes' accumulation in the perfusate and, as a corrective measure, the costly perfusate is repeatedly replaced during the standard of care procedure. As an interesting alternative, a hemodialyzer could be placed on the EVLP circuit, which was previously shown to rebalance the perfusate composition and to maintain lung function and viability without appearing to impact the global gene expression in the lung. Here, we assessed the biological effects of a hemodialyzer during EVLP by performing biochemical and refined functional genomic analyses over a 12h procedure in a pig model. We found that dialysis stabilized electrolytic and metabolic parameters of the perfusate but enhanced the gene expression and protein accumulation of several inflammatory cytokines and promoted a genomic profile predicting higher endothelial activation already at 6h and higher immune cytokine signaling at 12h. Therefore, epuration of EVLP with a dialyzer, while correcting features of the perfusate composition and maintaining the respiratory function, promotes inflammatory responses in the tissue. This finding suggests that modifying the metabolite composition of the perfusate by dialysis during EVLP can have detrimental effects on the tissue response and that this strategy should not be transferred as such to the clinic.

Hypothermic Oxygenated Perfusion Improves Vascular and Contractile Function by Preserving Endothelial Nitric Oxide Production in Cardiac Grafts Obtained With Donation After Circulatory Death.

BACKGROUND: Cardiac donation after circulatory death is a promising option to increase graft availability. Graft preservation with 30 minutes of hypothermic oxygenated perfusion (HOPE) before normothermic machine perfusion may improve cardiac recovery as compared with cold static storage, the current clinical standard. We investigated the role of preserved nitric oxide synthase activity during HOPE on its beneficial effects. METHODS AND RESULTS: Using a rat model of donation after circulatory death, hearts underwent in situ ischemia (21 minutes), were explanted for a cold storage period (30 minutes), and then reperfused under normothermic conditions (60 minutes) with left ventricular loading. Three cold storage conditions were compared: cold static storage, HOPE, and HOPE with Nω-nitro-L-arginine methyl ester (nitric oxide synthase inhibitor). To evaluate potential confounding effects of high coronary flow during early reperfusion in HOPE hearts, bradykinin was administered to normalize coronary flow to HOPE levels in 2 additional groups (cold static storage and HOPE with Nω-nitro-L-arginine methyl ester). Cardiac recovery was significantly improved in HOPE versus cold static storage hearts, as determined by cardiac output, left ventricular work, contraction and relaxation rates, and coronary flow (P<0.05). Furthermore, HOPE attenuated postreperfusion calcium overload. Strikingly, the addition of Nω-nitro-L-arginine methyl ester during HOPE largely abolished its beneficial effects, even when early reperfusion coronary flow was normalized to HOPE levels. CONCLUSIONS: HOPE provides superior preservation of ventricular and vascular function compared with the current clinical standard. Importantly, HOPE's beneficial effects require preservation of nitric oxide synthase activity during the cold storage. Therefore, the application of HOPE before normothermic machine perfusion is a promising approach to optimize graft recovery in donation after circulatory death cardiac grafts.

Clinical outcome of a branch-first approach with a novel continuous whole-brain perfusion strategy for total arch surgery.

BACKGROUND: Cerebral protection strategies have been investigated since the introduction of aortic arch surgery and have been modified over the centuries. However, the cerebral protective effects of unilateral and bilateral antegrade cerebral perfusion are similar, with opportunities for further improvement. METHODS: A total of 30 patients who underwent total arch surgery were enrolled in this study. Patients were assigned to the novel continuous whole-brain or unilateral antegrade cerebral perfusion group according to the cerebral perfusion technique used. Preoperative clinical data and 1-year postoperative follow-up data were collected and analyzed. RESULTS: There were no significant differences between the two groups in terms of the incidence of permanent neurological deficit, mortality, or therapeutic efficacy. However, the incidence of temporary neurological dysfunction in the novel whole-brain perfusion group was significantly lower than that in the unilateral antegrade cerebral perfusion group. CONCLUSIONS: In this study, the branch-first approach with a novel whole-brain perfusion strategy had no obvious disadvantages compared with unilateral antegrade cerebral perfusion in terms of cerebral protection and surgical safety. These findings suggest that this new technique is feasible and has application value for total arch surgery.

The impact and relevance of techniques and fluids on lung injury in machine perfusion of lungs.

Primary graft dysfunction (PGD) is a common complication after lung transplantation. A plethora of contributing factors are known and assessment of donor lung function prior to organ retrieval is mandatory for determination of lung quality. Specialized centers increasingly perform ex vivo lung perfusion (EVLP) to further assess lung functionality and improve and extend lung preservation with the aim to increase lung utilization. EVLP can be performed following different protocols. The impact of the individual EVLP parameters on PGD development, organ function and postoperative outcome remains to be fully investigated. The variables relate to the engineering and function of the respective perfusion devices, such as the type of pump used, functional, like ventilation modes or physiological (e.g. perfusion solutions). This review reflects on the individual technical and fluid components relevant to EVLP and their respective impact on inflammatory response and outcome. We discuss key components of EVLP protocols and options for further improvement of EVLP in regard to PGD. This review offers an overview of available options for centers establishing an EVLP program and for researchers looking for ways to adapt existing protocols.

Negligible In Vitro Recovery of Macromolecules from Microdialysis Using 100 kDa Probes and Dextran in Perfusion Fluid.

Microdialysis is applied in neurointensive care to monitor cerebral glucose metabolism. If recoverable, macromolecules may also serve as biomarkers in brain disease and provide clues to their passage across the blood-brain barrier. Our study aimed to investigate the in vitro recovery of human micro- and macromolecules using microdialysis catheters and perfusion fluids approved for clinical use. In vitro microdialysis of a bulk solution containing physiological or supraphysiological concentrations of glucose, lactate, pyruvate, human IgG, serum albumin, and hemoglobin was performed using two different catheters and perfusion fluids. One had a membrane cut-off of 20 kDa and was used with a standard CNS perfusion fluid, and the other had a membrane cut-off of 100 kDa and was perfused with the same solution supplemented with dextran. The flow rate was 0.3 µl/min. We used both push and push-pull methods. Dialysate samples were collected at 2-h intervals for 6 h and analyzed for relative recovery of each substance. The mean relative recovery of glucose, pyruvate, and lactate was > 90% in all but two sets of experiments. In contrast, the relative recovery of human IgG, serum albumin, and hemoglobin from both bulk solutions was below the lower limit of quantification (LLOQ). Using a push-pull method, recovery of human IgG, serum albumin, and hemoglobin from a bulk solution with supraphysiological concentrations were above LLOQ but with low relative recovery (range 0.9%-1.6%). In summary, exchanging the microdialysis setup from a 20 kDa catheter with a standard perfusion fluid for a 100 kDa catheter with a perfusion solution containing dextran did not affect the relative recovery of glucose and its metabolites. However, it did not result in any useful recovery of the investigated macromolecules at physiological levels, either with or without a push-pull pump system.

A Split-Lung Ex Vivo Perfusion Model for Time- and Cost-Effective Evaluation of Therapeutic Interventions to the Human Donor Lung.

With the ongoing shortage of donor lungs, ex vivo lung perfusion (EVLP) offers the opportunity for objective assessment and potential therapeutic repair of marginal organs. There is a need for robust research on EVLP interventions to increase the number of transplantable organs. The use of human lungs, which have been declined for transplant, for these studies is preferable to animal organs and is indeed essential if clinical translation is to be achieved. However, experimental human EVLP is time-consuming and expensive, limiting the rate at which promising interventions can be assessed. A split-lung EVLP model, which allows stable perfusion and ventilation of two single lungs from the same donor, offers advantages scientifically, financially and in time to yield results. Identical parallel circuits allow one to receive an intervention and the other to act as a control, removing inter-donor variation between study groups. Continuous hemodynamic and airway parameters are recorded and blood gas, perfusate, and tissue sampling are facilitated. Pulmonary edema is assessed directly using ultrasound, and indirectly using the lung tissue wet:dry ratio. Evans blue dye leaks into the tissue and can quantify vascular endothelial permeability. The split-lung ex vivo perfusion model offers a cost-effective, reliable platform for testing therapeutic interventions with relatively small sample sizes.

Twelve-hour normothermic liver perfusion in a rat model: characterization of the changes in the ex-situ bio-molecular phenotype and metabolism.

The partial understanding of the biological events that occur during normothermic machine perfusion (NMP) and particularly during prolonged perfusion might hinder its deployment in clinical transplantation. The aim of our study was to implement a rat model of prolonged NMP to characterize the bio-molecular phenotype and metabolism of the perfused organs. Livers (n = 5/group) were procured and underwent 4 h (NMP4h) or 12 h (NMP12h) NMP, respectively, using a perfusion fluid supplemented with an acellular oxygen carrier. Organs that were not exposed to any procedure served as controls (Native). All perfused organs met clinically derived viability criteria at the end of NMP. Factors related to stress-response and survival were increased after prolonged perfusion. No signs of oxidative damage were detected in both NMP groups. Evaluation of metabolite profiles showed preserved mitochondrial function, activation of Cori cycle, induction of lipolysis, acetogenesis and ketogenesis in livers exposed to 12 h-NMP. Increased concentrations of metabolites involved in glycogen synthesis, glucuronidation, bile acid conjugation, and antioxidant response were likewise observed. In conclusion, our NMP12h model was able to sustain liver viability and function, thereby deeply changing cell homeostasis to maintain a newly developed equilibrium. Our findings provide valuable information for the implementation of optimized protocols for prolonged NMP.

Stepwise development and expansion of an abdominal normothermic regional perfusion program for donation after circulatory determination of death organ procurement.

INTRODUCTION: Normothermic regional perfusion (NRP) represents an innovative technology that improves the outcomes for liver and kidney recipients of donation after circulatory determination of death (DCD) organs but protocols for abdominal-only NRP (A-NRP) DCD are lacking in the US. METHODS: We describe the implementation and expansion strategies of a transplant-center-based A-NRP DCD program that has grown in volume, geographical reach, and donor acceptance parameters, presented as four eras. RESULTS: In the implementation era, two donors were attempted, and one liver graft was transplanted. In the local expansion era, 33% of attempted donors resulted in transplantation and 42% of liver grafts from donors who died within the functional warm ischemic time (fWIT) limit were transplanted. In the Regional Expansion era, 25% of attempted donors resulted in transplantation and 50% of liver grafts from donors who died within the fWIT limit were transplanted. In the Donor Acceptance Expansion era, 46% of attempted donors resulted in transplantation and 72% of liver grafts from donors who died within the fWIT limit were transplanted. Eight discarded grafts demonstrated a potential opportunity for utilization. CONCLUSION: The stepwise approach to building an A-NRP program described here can serve as a model for other transplant centers.

Financial impact of donation after circulatory death heart transplantation: A single-center analysis.

INTRODUCTION: Clinical success of donation after circulatory death (DCD) heart transplantation is leading to growing adoption of this technique. In comparison to procurement from a brain-dead donor, DCD requires additional resources. The economic impact of DCD heart transplantation from the hospital perspective is not well known. METHODS: We compared the financial data of patients who received DCD allografts to those who received a DBD organ at our institution from January 1, 2021 to December 31, 2022. We also compared the cost of ex-situ machine perfusion to in-situ organ perfusion employed during DCD recovery. RESULTS: We performed 58 DBD and 22 DCD heart-alone transplantations during the study period. Out of 22 DCD grafts, 16 were recovered with thoracoabdominal normothermic regional perfusion (TA-NRP) and six with direct procurement followed by normothermic machine perfusion (DP-NMP). The contribution margin per case for DBD versus DCD was $234,362 and $235,440 (P = .72). The direct costs did not significantly differ between the two groups ($171,949 and 186,250; P = .49). In comparing the two methods of procuring hearts from DCD donors, the direct cost of TA-NRP was $155,955 in comparison to $223,399 for DP-NMP (P = .21). This difference translated into a clinically meaningful but not statistically significant greater contribution margin for TA-NRP ($242, 657 vs. $175,768; P = .34). CONCLUSIONS: Our data showed that the adoption of DCD procurement did not have a negative financial impact on the contribution margin in our institution. Programs considering starting DCD heart transplantation, and those who are currently performing DCD procurement should evaluate their own financial situation.

Real-time monitoring of mitochondrial oxygenation during machine perfusion using resonance Raman spectroscopy predicts organ function.

Organ transplantation is a life-saving procedure affecting over 100,000 people on the transplant waitlist. Ischemia reperfusion injury (IRI) is a major challenge in the field as it can cause post-transplantation complications and limit the use of organs from extended criteria donors. Machine perfusion technology has the potential to mitigate IRI; however, it currently fails to achieve its full potential due to a lack of highly sensitive and specific assays to assess organ quality during perfusion. We developed a real-time and non-invasive method of assessing organs during perfusion based on mitochondrial function and injury using resonance Raman spectroscopy. It uses a 441 nm laser and a high-resolution spectrometer to quantify the oxidation state of mitochondrial cytochromes during perfusion. This index of mitochondrial oxidation, or 3RMR, was used to understand differences in mitochondrial recovery of cold ischemic rodent livers during machine perfusion at normothermic temperatures with an acellular versus cellular perfusate. Measurement of the mitochondrial oxidation revealed that there was no difference in 3RMR of fresh livers as a function of normothermic perfusion when comparing acellular versus cellular-based perfusates. However, following 24 h of static cold storage, 3RMR returned to baseline faster with a cellular-based perfusate, yet 3RMR progressively increased during perfusion, indicating injury may develop over time. Thus, this study emphasizes the need for further refinement of a reoxygenation strategy during normothermic machine perfusion that considers cold ischemia durations, gradual recovery/rewarming, and risk of hemolysis.

Improved outcomes after hypothermic oxygenated machine perfusion in liver transplantation-Long-term follow-up of a multicenter randomized controlled trial.

BACKGROUND: While 4 randomized controlled clinical trials confirmed the early benefits of hypothermic oxygenated machine perfusion (HOPE), high-level evidence regarding long-term clinical outcomes is lacking. The aim of this follow-up study from the HOPE-ECD-DBD trial was to compare long-term outcomes in patients who underwent liver transplantation using extended criteria donor allografts from donation after brain death (ECD-DBD), randomized to either HOPE or static cold storage (SCS). METHODS: Between September 2017 and September 2020, recipients of liver transplantation from 4 European centers receiving extended criteria donor-donation after brain death allografts were randomly assigned to HOPE or SCS (1:1). Follow-up data were available for all patients. Analyzed endpoints included the incidence of late-onset complications (occurring later than 6 months and graded according to the Clavien-Dindo Classification and the Comprehensive Complication Index) and long-term graft survival and patient survival. RESULTS: A total of 46 patients were randomized, 23 in both arms. The median follow-up was 48 months (95% CI: 41-55). After excluding early perioperative morbidity, a significant reduction in late-onset morbidity was observed in the HOPE group (median reduction of 23 Comprehensive Complication Index-points [p=0.003] and lower incidence of major complications [Clavien-Dindo ≥3, 43% vs. 85%, p=0.009]). Primary graft loss occurred in 13 patients (HOPE n=3 vs. SCS n=10), resulting in a significantly lower overall graft survival (p=0.029) and adverse 1-, 3-, and 5-year survival probabilities in the SCS group, which did not reach the level of significance (HOPE 0.913, 0.869, 0.869 vs. SCS 0.783, 0.606, 0.519, respectively). CONCLUSIONS: Our exploratory findings indicate that HOPE reduces late-onset morbidity and improves long-term graft survival providing clinical evidence to further support the broad implementation of HOPE in human liver transplantation.

The Advent of Semi-Elective Lung Transplantation-Prolonged Static Cold Storage at 10°C.

Since the early days of clinical lung transplantation the preservation of donor organs has become a fairly standardized procedure and most centers do follow similar processes. This includes the use of low-potassium high dextran flush solutions and static cold storage (SCS) in a cooler filled with ice. Depending on the length of SCS, organs usually arrive at the recipient hospital at a temperature of 0°C-4°C. The question of the optimal storage temperature for donor lung preservation has been revisited as data from large animal experiments demonstrated that organs stored at 10°C experience less mitochondrial damage. Thus, prolonged cold ischemic times can be better tolerated at 10°C-even in pre-damaged organs. The clinical applicability of these findings was demonstrated in an international multi-center observational study including three high-volume lung transplant centers. Total clinical preservation times of up to 24 hrs have been successfully achieved in organs stored at 10°C without hampering primary organ function and short-term outcomes. Currently, a randomized-controlled trial (RCT) is recruiting patients with the aim to compare standard SCS on ice with prolonged SCS protocol at 10°C. If, as anticipated, this RCT confirms data from previous studies, lung transplantation could indeed become a semi-elective procedure.

Cannula Placement for Cerebral Protection Without Circulatory Arrest in Patients Undergoing Hemiarch Aortic Aneurysm Repair.

BACKGROUND: Aortic aneurysms involving the proximal aortic arch, which require hemiarch-type repair, typically require circulatory arrest with antegrade cerebral perfusion. Left carotid antegrade cerebral perfusion (LCP) via distal arch cannulation without circulatory arrest was used in this study's patient population. The goal was to assess the operative efficiency and clinical outcomes of using a distal arch cannulation technique that would not require any hypothermic circulatory arrest (HCA) time compared with more traditional brachiocephalic artery cannulation with right-sided unilateral antegrade cerebral perfusion (RCP) and HCA. METHODS: A single-center retrospective review of patients with replacement of the distal ascending aorta involving the proximal arch was performed. Patients with an intramural hematoma or dissection were excluded. Between January 2015 and December 2019, 68 adult patients had undergone a hemiarch repair because of aneurysmal disease. Analysis of baseline demographics, operative data, and clinical outcomes was performed. RESULTS: Comparing the 68 patients: 21 patients were treated with RCP (via brachiocephalic artery graft with HCA), and 47 patients were treated with LCP (via distal aortic arch cannulation with cross-clamp between the brachiocephalic and left common carotid arteries without HCA). Baseline characteristics and outcomes were evaluated for both groups. The LCP group was younger (LCP median [IQR] age, 60 [53-65] years vs RCP median [IQR] age, 67 [59-71] years]. Sex, race, body mass index, comorbidities, and ejection fraction were similar between the groups. Cardiopulmonary bypass time (LCP, 123 minutes vs RCP, 149 minutes) and unilateral cerebral perfusion time (LCP, 17 minutes vs RCP, 22 minutes) were longer in the RCP group. Bleeding, prolonged ventilatory support, kidney failure, and length of stay were similar. In-hospital mortality was 2% in the LCP group vs 0% in the RCP group. Stroke occurred in 2 patients (4.2%) in the LCP group and in 0% of the RCP group. Mortality at 6 months in the LCP and RCP groups was 3% and 10%, respectively. CONCLUSION: Distal arch cannulation with LCP without HCA is a reasonable and safe alternative strategy for patients requiring hemiarch replacement for aneurysmal disease. This technique may provide additional benefits by avoiding circulatory arrest in these complex cases.

Machine perfusion of the liver and in vivo animal models: A systematic review of the preclinical research landscape.

Machine perfusion (MP) is often referred to as one of the most promising advancements in liver transplantation research of the last few decades, with various techniques and modalities being evaluated in preclinical studies using animal models. However, low scientific rigor and subpar reporting standards lead to limited reproducibility and translational potential, hindering progress. This pre-registered systematic review (PROSPERO: CRD42021234667) aimed to provide a thematic overview of the preclinical research landscape on MP in liver transplantation using in vivo transplantation models and to explore methodological and reporting standards, using the ARRIVE (Animal Research: Reporting of In Vivo Experiments) score. In total 56 articles were included. Studies were evenly distributed across Asia, Europe, and the Americas. Porcine models were used in 57.1% of the studies, followed by rats (39.3%) and dogs (3.6%). In terms of graft type, 55.4% of the studies used donation after cardiac death grafts, while donation after brain death grafts accounted for 37.5%. Regarding MP modalities, the distribution was as follows: 41.5% of articles utilized hypothermic MP, 21.5% normothermic MP, 13.8% subnormothermic MP, and 16.9% utilized hypothermic oxygenated MP. The stringent documentation of ARRIVE elements concerning precise experimental execution, group size and selection, the choice of statistical methods, as well as adherence to the principles of the 3Rs, was notably lacking in the majority of publications, with less than 30% providing comprehensive details. Postoperative analgesia and antibiotics treatment were not documented in 82.1% of all included studies. None of the analyzed studies fully adhered to the ARRIVE Guidelines. In conclusion, the present study emphasizes the importance of adhering to reporting standards to promote reproducibility and adequate animal welfare in preclinical studies in machine perfusion. At the same time, it highlights a clear deficiency in this field, underscoring the need for further investigations into animal welfare-related topics.

Assessing Liver Viability: Insights From Mitochondrial Bioenergetics in Ischemia-Reperfusion Injury.

Orthotopic liver transplantation remains the definitive treatment for patients with end-stage liver disease. Unfortunately, the increasing demand for donor livers and the limited supply of viable organs have both led to a critical need for innovative strategies to expand the pool of transplantable organs. The mitochondrion, central to hepatic cellular function, plays a pivotal role in hepatic ischemic injury, with impaired mitochondrial function and oxidative stress leading to cell death. Mitochondrial protection strategies have shown promise in mitigating IRI and resuscitating marginal organs for transplant. Machine perfusion (MP) has been proven a valuable tool for reviving marginal organs with very promising results. Evaluation of liver viability during perfusion traditionally relies on parameters including lactate clearance, bile production, and transaminase levels. Nevertheless, the quest for more comprehensive and universally applicable viability markers persists. Normothermic regional perfusion has gained robust attention, offering extended recovery time for organs from donation after cardiac death donors. This approach has shown remarkable success in improving organ quality and reducing ischemic injury using the body's physiological conditions. The current challenge lies in the absence of a reliable assessment tool for predicting graft viability and post-transplant outcomes. To address this, exploring insights from mitochondrial function in the context of ischemia-reperfusion injury could offer a promising path toward better patient outcomes and graft longevity. Indeed, hypoxia-induced mitochondrial injury may serve as a surrogate marker of organ viability following oxygenated resuscitation techniques in the future.

A cost-effective and efficient ex vivo, ex situ human whole brain perfusion protocol for immunohistochemistry.

BACKGROUND: Chemical fixation of the brain can be executed through either the immersion method or the perfusion method. Perfusion fixation allows for better preservation of the brain tissue's ultrastructure, as it provides rapid and uniform delivery of the fixative to the tissue. Still, not all facilities have the expertise to perform perfusion fixation, with initial high cost and complexity of perfusion systems as the main factors limiting its widespread usage. NEW METHOD: Here we present our low-cost approach of whole brain ex situ perfusion fixation to overcome the aforementioned limitations. Our self-made perfusion system, constructed utilising commercially accessible and affordable medical resources alongside laboratory and everyday items, demonstrates the capability to generate superior histological stainings of brain tissue. The perfused tissue can be stored prior to proceeding with IHC for at least one year. RESULTS: Our method yielded high-quality results in histological stainings using both free-floating cryosections and paraffin-embedded tissue sections. The system is fully reusable and complies with the principles of sustainable management. COMPARISON WITH EXISTING METHODS: Our whole brain perfusion system has been assembled from simple components and is able to achieve a linear flow with a pressure of 70 mmHg corresponding to the perfusion pressure of the brain. CONCLUSIONS: Our ex situ method can be especially useful in research settings where expensive perfusion systems are not affordable or in any field with high time pressure, making it suitable for the field of forensic medicine or pathology in general.

Transcriptomic Changes in the Myocardium and Coronary Artery of Donation after Circulatory Death Hearts following Ex Vivo Machine Perfusion.

Donation after circulatory death (DCD) hearts are predominantly maintained by normothermic blood perfusion (NBP). Nevertheless, it was shown that hypothermic crystalloid perfusion (HCP) is superior to blood perfusion to recondition left ventricular (LV) contractility. However, transcriptomic changes in the myocardium and coronary artery in DCD hearts after HCP and NBP have not been investigated yet. In a pig model, DCD hearts were harvested and maintained for 4 h by NBP (DCD-BP group, N = 8) or HCP with oxygenated histidine-tryptophane-ketoglutarate (HTK) solution (DCD-HTK, N = 8) followed by reperfusion with fresh blood for 2 h. In the DCD group (N = 8), hearts underwent reperfusion immediately after procurement. In the control group (N = 7), no circulatory death was induced. We performed transcriptomics from LV myocardial and left anterior descending (LAD) samples using microarrays (25,470 genes). We applied the Boruta algorithm for variable selection to identify relevant genes. In the DCD-BP group, compared to DCD, six genes were regulated in the myocardium and 1915 genes were regulated in the LAD. In the DCD-HTK group, 259 genes were downregulated in the myocardium and 27 in the LAD; and 52 genes were upregulated in the myocardium and 765 in the LAD, compared to the DCD group. We identified seven genes of relevance for group identification: ITPRIP, G3BP1, ARRDC3, XPO6, NOP2, SPTSSA, and IL-6. NBP resulted in the upregulation of genes involved in mitochondrial calcium accumulation and ROS production, the reduction in microvascular endothelial sprouting, and inflammation. HCP resulted in the downregulation of genes involved in NF-κB-, STAT3-, and SASP-activation and inflammation.

Comprehensive bile acid pool analysis during ex-vivo liver perfusion in a porcine model of ischemia-reperfusion injury.

Bile acids (BA) are key for liver regeneration and injury. This study aims at analyzing the changes in the BA pool induced by ischemia-reperfusion (IRI) and investigates the impact of hypothermic oxygenated perfusion (HOPE) on the BA pool compared to static cold storage (SCS). In a porcine model of IRI, liver grafts underwent 30 min of asystolic warm ischemia followed by 6 h of SCS (n = 6) ± 2 h of HOPE (n = 6) and 2 h of ex-situ warm reperfusion. The BA pool in bile samples was analyzed with liquid chromatography coupled with tandem mass spectrometry. We identified 16 BA and observed significant changes in response to ischemia-reperfusion, which were associated with both protective and injury mechanisms. Second, HOPE-treated liver grafts exhibited a more protective BA phenotype, characterized by a more hydrophilic BA pool compared to SCS. Key BA, such as GlycoCholic Acid, were identified and were associated with a decreased transaminase release and improved lactate clearance during reperfusion. Partial Least Square-Discriminant Analysis revealed a distinct injury profile for the HOPE group. In conclusion, the BA pool changes with liver graft IRI, and preservation with HOPE results in a protective BA phenotype compared to SCS.