Tryptophan Metabolism via the Kynurenine Pathway: Implications for Graft Optimization during Machine Perfusion

Access to liver transplantation continues to be hindered by the severe organ shortage. Extended-criteria donor livers could be used to expand the donor pool but are prone to ischemia-reperfusion injury (IRI) and post-transplant graft dysfunction. Ex situ machine perfusion may be used as a platform to rehabilitate discarded or extended-criteria livers prior to transplantation, though there is a lack of data guiding the utilization of different perfusion modalities and therapeutics. Since amino acid derivatives involved in inflammatory and antioxidant pathways are critical in IRI, we analyzed differences in amino acid metabolism in seven discarded non-steatotic human livers during normothermic- (NMP) and subnormothermic-machine perfusion (SNMP) using data from untargeted metabolomic profiling. We found notable differences in tryptophan, histamine, and glutathione metabolism. Greater tryptophan metabolism via the kynurenine pathway during NMP was indicated by significantly higher kynurenine and kynurenate tissue concentrations compared to pre-perfusion levels. Livers undergoing SNMP demonstrated impaired glutathione synthesis indicated by depletion of reduced and oxidized glutathione tissue concentrations. Notably, ATP and energy charge ratios were greater in livers during SNMP compared to NMP. Given these findings, several targeted therapeutic interventions are proposed to mitigate IRI during liver machine perfusion and optimize marginal liver grafts during SNMP and NMP.

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Ex situ Perfusion of Pancreas for Whole-Organ Transplantation: Is it Safe and Feasible? A Systematic Review

Journal of Diabetes Science and Technology ◽

10.1177/1932296819869312 ◽

2019 ◽

Vol 14 (1) ◽

pp. 120-134 ◽

Cited By ~ 1

Author(s):

Thomas Prudhomme ◽

Delphine Kervella ◽

Stéphanie Le Bas-Bernardet ◽

Diego Cantarovich ◽

Georges Karam ◽

...

Keyword(s):

Organ Transplantation ◽

Pancreas Transplantation ◽

Ex Vivo ◽

English Literature ◽

Ischemia Reperfusion ◽

Graft Loss ◽

Ex Situ ◽

Machine Perfusion ◽

Pancreas Perfusion ◽

Ischemic Reperfusion

Introduction: Pancreas transplantation is currently one of the best treatments proposed in highly selected patients with unstable and brittle type 1 diabetes. The objective of pancreas transplantation is to restore normoglycemia and avoid the occurrence of complications associated with diabetes. Graft pancreatitis and thrombosis, arising from ischemia reperfusion injuries, are major causes of graft loss in the postoperative period. Ex situ perfusion, in hypothermic or normothermic settings, allowed to improve ischemic reperfusion injury in other organ transplantations (kidney, liver, or lung). The development of pancreatic graft perfusion techniques would limit these ischemic reperfusion injuries. Objective: Evaluation of the safety and feasibility of ex situ perfusion of pancreas for whole-organ transplantation. Methods: English literature about pancreas perfusion was analyzed using electronic database Medline via PubMed (1950-2018). Exclusion criteria were studies that did not specify the technical aspects of machine perfusion and studies focused only on pancreas perfusion for islet isolation. Results: Hypothermic machine perfusion for pancreas preservation has been evaluated in nine studies and normothermic machine perfusion in ten studies. We evaluated machine perfusion model, types of experimental model, anatomy, perfusion parameters, flushing and perfusion solution, length of perfusion, and comparison between static cold storage and perfusion. Conclusions: This review compared ex vivo machine perfusion of experimental pancreas for whole-organ transplantation. Pancreas perfusion is feasible and could be a helpful tool to evaluate pancreas prior to transplantation. Pancreas perfusion (in hypothermic or normothermic settings) could reduce ischemic reperfusion injuries, and maybe could avoid pancreas thrombosis and reduce morbidity of pancreas transplantation.

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Split-Liver Ex Situ Machine Perfusion: A Novel Technique for Studying Organ Preservation and Therapeutic Interventions

Journal of Clinical Medicine ◽

10.3390/jcm9010269 ◽

2020 ◽

Vol 9 (1) ◽

pp. 269 ◽

Cited By ~ 2

Author(s):

Viola Huang ◽

Negin Karimian ◽

Danielle Detelich ◽

Siavash Raigani ◽

Sharon Geerts ◽

...

Keyword(s):

Liver Perfusion ◽

Energy Charge ◽

Therapeutic Interventions ◽

Ex Situ ◽

Preclinical Studies ◽

Machine Perfusion ◽

Large Numbers ◽

Split Liver ◽

Human Livers ◽

Lactate Levels

Ex situ machine perfusion is a promising technology to help improve organ viability prior to transplantation. However, preclinical studies using discarded human livers to evaluate therapeutic interventions and optimize perfusion conditions are limited by significant graft heterogeneity. In order to improve the efficacy and reproducibility of future studies, a split-liver perfusion model was developed to allow simultaneous perfusion of left and right lobes, allowing one lobe to serve as a control for the other. Eleven discarded livers were surgically split, and both lobes perfused simultaneously on separate perfusion devices for 3 h at subnormothermic temperatures. Lobar perfusion parameters were also compared with whole livers undergoing perfusion. Similar to whole-liver perfusions, each lobe in the split-liver model exhibited a progressive decrease in arterial resistance and lactate levels throughout perfusion, which were not significantly different between right and left lobes. Split liver lobes also demonstrated comparable energy charge ratios. Ex situ split-liver perfusion is a novel experimental model that allows each graft to act as its own control. This model is particularly well suited for preclinical studies by avoiding the need for large numbers of enrolled livers necessary due to the heterogenous nature of discarded human liver research.

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Ischemia-Reperfusion Injury in Marginal Liver Grafts and the Role of Hypothermic Machine Perfusion: Molecular Mechanisms and Clinical Implications

Journal of Clinical Medicine ◽

10.3390/jcm9030846 ◽

2020 ◽

Vol 9 (3) ◽

pp. 846 ◽

Cited By ~ 4

Author(s):

Zoltan Czigany ◽

Isabella Lurje ◽

Moritz Schmelzle ◽

Wenzel Schöning ◽

Robert Öllinger ◽

...

Keyword(s):

Reperfusion Injury ◽

Molecular Mechanisms ◽

Ischemia Reperfusion Injury ◽

Ex Vivo ◽

Ischemia Reperfusion ◽

Graft Function ◽

Organ Shortage ◽

Machine Perfusion ◽

Hypothermic Machine Perfusion ◽

Critical Organ

Ischemia-reperfusion injury (IRI) constitutes a significant source of morbidity and mortality after orthotopic liver transplantation (OLT). The allograft is metabolically impaired during warm and cold ischemia and is further damaged by a paradox reperfusion injury after revascularization and reoxygenation. Short-term and long-term complications including post-reperfusion syndrome, delayed graft function, and immune activation have been associated with IRI. Due to the current critical organ shortage, extended criteria grafts are increasingly considered for transplantation, however, with an elevated risk to develop significant features of IRI. In recent years, ex vivo machine perfusion (MP) of the donor liver has witnessed significant advancements. Here, we describe the concept of hypothermic (oxygenated) machine perfusion (HMP/HOPE) approaches and highlight which allografts may benefit from this technology. This review also summarizes clinical applications and the main aspects of ongoing randomized controlled trials on hypothermic perfusion. The mechanistic aspects of IRI and hypothermic MP—which include tissue energy replenishment, optimization of mitochondrial function, and the reduction of oxidative and inflammatory damage following reperfusion—will be comprehensively discussed within the context of current preclinical and clinical evidence. Finally, we highlight novel trends and future perspectives in the field of hypothermic MP in the context of recent findings of basic and translational research.

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Hypothermic Oxygenated Machine Perfusion of Extended Criteria Kidney Allografts from Brain Dead Donors: Protocol for a Prospective Pilot Study

JMIR Research Protocols ◽

10.2196/14622 ◽

2019 ◽

Vol 8 (10) ◽

pp. e14622 ◽

Cited By ~ 3

Author(s):

Franziska Alexandra Meister ◽

Zoltan Czigany ◽

Jan Bednarsch ◽

Jörg Böcker ◽

Iakovos Amygdalos ◽

...

Keyword(s):

Kidney Transplantation ◽

Pilot Study ◽

Brain Death ◽

Ischemia Reperfusion ◽

Kidney Tissue ◽

Organ Shortage ◽

Machine Perfusion ◽

Extended Criteria Donor ◽

Prospective Pilot Study ◽

Clinical Series

Background Kidney transplantation is the only curative treatment option for end-stage renal disease. The unavailability of adequate organs for transplantation has resulted in a substantial organ shortage. As such, kidney donor allografts that would have previously been deemed unsuitable for transplantation have become an essential organ pool of extended criteria donor allografts that are now routinely being transplanted on a global scale. However, these extended criteria donor allografts are associated with significant graft-related complications. As a result, hypothermic oxygenated machine perfusion (HOPE) has emerged as a powerful, novel technique in organ preservation, and it has recently been tested in preclinical trials in kidney transplantation. In addition, HOPE has already provided promising results in a few clinical series of liver transplantations where the liver was donated after cardiac death. Objective The present trial is an investigator-initiated prospective pilot study on the effects of HOPE on extended criteria donor allografts donated after brain death and used in kidney transplantation. Methods A total of 15 kidney allografts with defined inclusion/exclusion criteria will be submitted to two hours of HOPE via the renal artery before implantation, and are going to be compared to a case-matched group of 30 patients (1:2 matching) who had kidneys transplanted after conventional cold storage. Primary (posttransplant dialysis within 7 days) and secondary (postoperative complications, early graft function, duration of hospital and intensive care unit stay, and six-month graft survival) endpoints will be analyzed within a six-month follow-up period. The extent of ischemia-reperfusion injury will be assessed using kidney tissue, perfusate, and serum samples taken during the perioperative phase of kidney transplantation Results The results of this trial are expected in the first quarter of 2020 and will be presented at national and international scientific meetings and published in international peer-reviewed medical journals. The trial was funded in the third quarter of 2017 and patient enrollment is currently ongoing. Conclusions This prospective study is designed to explore the effects of HOPE on extended criteria donor kidney allografts donated after brain death. The present report represents the preresults phase. Trial Registration Clinicaltrials.gov NCT03378817; https://clinicaltrials.gov/ct2/show/NCT03378817

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Subnormothermic Machine Perfusion of Steatotic Livers Results in Increased Energy Charge at the Cost of Anti-Oxidant Capacity Compared to Normothermic Perfusion

Metabolites ◽

10.3390/metabo9110246 ◽

2019 ◽

Vol 9 (11) ◽

pp. 246 ◽

Cited By ~ 2

Author(s):

Negin Karimian ◽

Siavash Raigani ◽

Viola Huang ◽

Sonal Nagpal ◽

Ehab O. A. Hafiz ◽

...

Keyword(s):

Energy Charge ◽

Redox Status ◽

High Energy ◽

Ex Situ ◽

Machine Perfusion ◽

Normothermic Perfusion ◽

Normothermic Machine Perfusion ◽

Anti Oxidant ◽

Human Livers ◽

The Cost

There continues to be significant debate regarding the most effective mode of ex situ machine perfusion of livers for transplantation. Subnormothermic (SNMP) and normothermic machine perfusion (NMP) are two methods with different benefits. We examined the metabolomic profiles of discarded steatotic human livers during three hours of subnormothermic or normothermic machine perfusion. Steatotic livers regenerate higher stores of ATP during SNMP than NMP. However, there is a significant depletion of available glutathione during SNMP, likely due to an inability to overcome the high energy threshold needed to synthesize glutathione. This highlights the increased oxidative stress apparent in steatotic livers. Rescue of discarded steatotic livers with machine perfusion may require the optimization of redox status through repletion or supplementation of reducing agents.

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Ex-Vivo Pharmacological Defatting of the Liver: A Review

Journal of Clinical Medicine ◽

10.3390/jcm10061253 ◽

2021 ◽

Vol 10 (6) ◽

pp. 1253

Author(s):

Claire Goumard ◽

Célia Turco ◽

Mehdi Sakka ◽

Lynda Aoudjehane ◽

Philippe Lesnik ◽

...

Keyword(s):

Ex Vivo ◽

Ex Situ ◽

Special Focus ◽

Organ Shortage ◽

Machine Perfusion ◽

Pharmacological Agents ◽

Promising Tool ◽

Normothermic Machine Perfusion ◽

C Metabolism ◽

Liver Machine Perfusion

The ongoing organ shortage has forced transplant teams to develop alternate sources of liver grafts. In this setting, ex-situ machine perfusion has rapidly developed as a promising tool to assess viability and improve the function of organs from extended criteria donors, including fatty liver grafts. In particular, normothermic machine perfusion represents a powerful tool to test a liver in full 37 °C metabolism and add pharmacological corrections whenever needed. In this context, many pharmacological agents and therapeutics have been tested to induce liver defatting on normothermic machine perfusion with promising results even on human organs. This systematic review makes a comprehensive synthesis on existing pharmacological therapies for liver defatting, with special focus on normothermic liver machine perfusion as an experimental ex-vivo translational model.

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The role of antifreeze glycopeptides (AFGP) and polyvinyl alcohol/polyglycerol (X/Z-1000) cocktails as ice modulators during partial freezing of rat livers.

10.1101/2021.08.04.455092 ◽

2021 ◽

Author(s):

Shannon N. Tessier ◽

Omar Haque ◽

Casie A. Pendexter ◽

Stephanie E.J. Cronin ◽

Lindong Weng ◽

...

Keyword(s):

Polyvinyl Alcohol ◽

Structural Integrity ◽

Cell Damage ◽

Energy Charge ◽

Endothelial Damage ◽

Terminal Deoxynucleotidyl Transferase ◽

Tunel Staining ◽

Organ Shortage ◽

Machine Perfusion ◽

Rat Livers

The current liver organ shortage has pushed the field of liver transplantation to develop new methods to prolong the preservation time of livers from the current clinical standard of static cold storage. Our approach, termed partial freezing, aims to induce a thermodynamically stable frozen state at deeper storage temperatures (–10°C to –15°C) than can be achieved with supercooling, while simultaneously maintaining a sufficient unfrozen fraction to limit dehydration and ice damage. This research first demonstrated that partially frozen glycerol treated rat livers were functionally similar after thawing from either –10 or –15°C with respect to subnormothermic machine perfusion metrics and histology. Next, we assessed the effect of adding either of two ice modulators, antifreeze glycoprotein (AFGP) and a polyvinyl alcohol/polyglycerol combination (X/Z-1000), on the viability and structural integrity of partially frozen rat livers compared to glycerol-only control livers. Results showed that AFGP livers had high levels of ATP and the least edema but suffered from significant endothelial cell damage. X/Z-1000 livers had the highest levels of ATP and energy charge (EC) but also demonstrated endothelial damage and post-thaw edema. Glycerol-only control livers exhibited the least DNA damage on Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining but also had the lowest levels of ATP and EC. Further research is necessary to optimize the ideal ice modulator cocktail for our partial-freezing protocol. Modifications to cryoprotective agent (CPA) combinations, as well as improvements to machine perfusion CPA loading and unloading, can help improve the viability of these partially frozen organs.

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Hyperthermia-induced changes in liver physiology and metabolism: a rationale for hyperthermic machine perfusion

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00101.2020 ◽

2020 ◽

Vol 319 (1) ◽

pp. G43-G50 ◽

Cited By ~ 1

Author(s):

Adam M. Thorne ◽

Rinse Ubbink ◽

Isabel M. A. Brüggenwirth ◽

Maarten W. Nijsten ◽

Robert J. Porte ◽

...

Keyword(s):

Ischemia Reperfusion Injury ◽

Core Body Temperature ◽

Ischemia Reperfusion ◽

Supply And Demand ◽

Metabolic Effects ◽

Ex Situ ◽

Machine Perfusion ◽

Extended Criteria Donor ◽

Different Temperatures ◽

Donor Organs

Liver transplantation is the standard treatment for end-stage liver disease. However, due to the ongoing disparity between supply and demand for optimal donor organs, there is increasing usage of extended criteria donor organs, including steatotic liver grafts. To mitigate the increased risks associated with extended criteria donor livers, ex situ oxygenated machine perfusion (MP) has received increasing attention in recent years as an emerging platform for dynamic preservation, reconditioning, and viability assessment to increase organ utilization. MP can be applied at different temperatures. During hypothermic MP (4–12°C), liver metabolism is reduced, while oxygenation restores the intracellular levels of adenosine triphosphate. The liver is quickly “recharged” to support metabolism when at normothermia (35–37°C) and to ameliorate the detrimental effects of ischemia/reperfusion injury during transplantation. During normothermia, MP can be applied to assess hepatocellular and cholangiocellular viability. MP at hyperthermic (>38°C) temperatures (HyMP), however, remains relatively understudied. The liver is an important component in the regulation of core body temperature and, as such, displays significant physiological and metabolic changes in response to different temperatures. Hyperthermia may promote vasodilation, increase aerobic metabolism and induce production of protective molecules such as heat shock proteins. Therefore, HyMP could provide an attractive reconditioning strategy for steatotic livers. In this review, we describe current literature on the physiological and metabolic effects of the liver at hyperthermia for human, rodents, and pigs and provide a rationale for using therapeutic HyMP during isolated liver machine perfusion to recondition extended criteria donor livers, including steatotic livers, before transplantation.

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Hypothermic Oxygenated Machine Perfusion of Extended Criteria Kidney Allografts from Brain Dead Donors: Protocol for a Prospective Pilot Study (Preprint)

10.2196/preprints.14622 ◽

2019 ◽

Author(s):

Franziska Alexandra Meister ◽

Zoltan Czigany ◽

Jan Bednarsch ◽

Jörg Böcker ◽

Iakovos Amygdalos ◽

...

Keyword(s):

Kidney Transplantation ◽

Pilot Study ◽

Brain Death ◽

Ischemia Reperfusion ◽

Kidney Tissue ◽

Organ Shortage ◽

Machine Perfusion ◽

Extended Criteria Donor ◽

Prospective Pilot Study ◽

Clinical Series

BACKGROUND Kidney transplantation is the only curative treatment option for end-stage renal disease. The unavailability of adequate organs for transplantation has resulted in a substantial organ shortage. As such, kidney donor allografts that would have previously been deemed unsuitable for transplantation have become an essential organ pool of extended criteria donor allografts that are now routinely being transplanted on a global scale. However, these extended criteria donor allografts are associated with significant graft-related complications. As a result, hypothermic oxygenated machine perfusion (HOPE) has emerged as a powerful, novel technique in organ preservation, and it has recently been tested in preclinical trials in kidney transplantation. In addition, HOPE has already provided promising results in a few clinical series of liver transplantations where the liver was donated after cardiac death. OBJECTIVE The present trial is an investigator-initiated prospective pilot study on the effects of HOPE on extended criteria donor allografts donated after brain death and used in kidney transplantation. METHODS A total of 15 kidney allografts with defined inclusion/exclusion criteria will be submitted to two hours of HOPE via the renal artery before implantation, and are going to be compared to a case-matched group of 30 patients (1:2 matching) who had kidneys transplanted after conventional cold storage. Primary (posttransplant dialysis within 7 days) and secondary (postoperative complications, early graft function, duration of hospital and intensive care unit stay, and six-month graft survival) endpoints will be analyzed within a six-month follow-up period. The extent of ischemia-reperfusion injury will be assessed using kidney tissue, perfusate, and serum samples taken during the perioperative phase of kidney transplantation RESULTS The results of this trial are expected in the first quarter of 2020 and will be presented at national and international scientific meetings and published in international peer-reviewed medical journals. The trial was funded in the third quarter of 2017 and patient enrollment is currently ongoing. CONCLUSIONS This prospective study is designed to explore the effects of HOPE on extended criteria donor kidney allografts donated after brain death. The present report represents the preresults phase. CLINICALTRIAL Clinicaltrials.gov NCT03378817; https://clinicaltrials.gov/ct2/show/NCT03378817

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Methionine and Arginine Supply Alters Abundance of Amino Acid, Insulin Signaling, and Glutathione Metabolism-Related Proteins in Bovine Subcutaneous Adipose Explants Challenged with N-Acetyl-d-sphingosine

Animals ◽

10.3390/ani11072114 ◽

2021 ◽

Vol 11 (7) ◽

pp. 2114

Author(s):

Yusheng Liang ◽

Nana Ma ◽

Danielle N. Coleman ◽

Fang Liu ◽

Yu Li ◽

...

Keyword(s):

Adipose Tissue ◽

Amino Acid ◽

Insulin Signaling ◽

Elongation Factor ◽

In Vivo Studies ◽

Glutathione Metabolism ◽

Protein Abundance ◽

Related Proteins ◽

Subcutaneous Adipose

The objective was to perform a proof-of-principle study to evaluate the effects of methionine (Met) and arginine (Arg) supply on protein abundance of amino acid, insulin signaling, and glutathione metabolism-related proteins in subcutaneous adipose tissue (SAT) explants under ceramide (Ce) challenge. SAT from four lactating Holstein cows was incubated with one of the following media: ideal profile of amino acid as the control (IPAA; Lys:Met 2.9:1, Lys:Arg 2:1), increased Met (incMet; Lys:Met 2.5:1), increased Arg (incArg; Lys:Arg 1:1), or incMet plus incArg (Lys:Met 2.5:1 Lys:Arg 1:1) with or without 100 μM exogenous cell-permeable Ce (N-Acetyl-d-sphingosine). Ceramide stimulation downregulated the overall abundance of phosphorylated (p) protein kinase B (AKT), p-mechanistic target of rapamycin (mTOR), and p-eukaryotic elongation factor 2 (eEF2). Without Ce stimulation, increased Met, Arg, or Met + Arg resulted in lower p-mTOR. Compared with control SAT stimulated with Ce, increased Met, Arg, or Met + Arg resulted in greater activation of mTOR (p-mTOR/total mTOR) and AKT (p-AKT/total AKT), with a more pronounced response due to Arg. The greatest protein abundance of glutathione S-transferase Mu 1 (GSTM1) was detected in response to increased Met supply during Ce stimulation. Ceramide stimulation decreased the overall protein abundance of the Na-coupled neutral amino acid transporter SLC38A1 and branched-chain alpha-ketoacid dehydrogenase kinase (BCKDK). However, compared with controls, increased Met or Arg supply attenuated the downregulation of BCKDK induced by Ce. Circulating ceramides might affect amino acid, insulin signaling, and glutathione metabolism in dairy cow adipose tissue. Further in vivo studies are needed to confirm the role of rumen-protected amino acids in regulating bovine adipose function.

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