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Surgery Apr 2021Long-term ex situ liver perfusion may rescue injured grafts. Little is known about bile flow during long-term perfusion. We report the development of a bile stimulation...
BACKGROUND
Long-term ex situ liver perfusion may rescue injured grafts. Little is known about bile flow during long-term perfusion. We report the development of a bile stimulation protocol and motivate bile flow as a viability marker during long-term ex situ liver perfusion.
METHODS
Porcine and human livers were perfused with blood at close to physiologic conditions. Our perfusion protocol was established during phase 1 with porcine livers (n = 23). Taurocholic acid was applied to stimulate bile flow. The addition of piperacillin-tazobactam (tazobac) and methylprednisolone was modified from daily bolus to controlled continuous application. We adapted the protocol to human livers (n = 12) during phase 2. Taurocholic acid was replaced with medical grade ursodeoxycholic acid.
RESULTS
Phase 2: Despite administering taurocholic acid, bile flow declined from 29.3 ± 6.5 to 9.3 ± 1.4 mL/h (P < .001). Shortly after bolus of tazobac/methylprednisolone, bile flow recovered to 39.0 ± 9.7 mL/h with a decrease of solid bile components. This implied bile salt independent bile flow stimulation by tazobac/methylprednisolone. Phase 2: Ursodeoxycholic acid was shown to stimulate bile flow ex situ in human livers. Eight livers were perfused successfully for 1 week with continuous bile flow. The other 4 livers demonstrated progressive cell death, of which only 1 exhibited bile flow.
CONCLUSION
A lack of bile flow stimulation leads to a decline in bile flow and is not necessarily a sign of deterioration in liver function. Proper administration of stimulators can induce constant bile flow during ex situ liver perfusion for up to 1 week. Medical grade ursodeoxycholic acid is a suitable replacement for nonmedical grade taurocholic acid. The presence of bile flow alone is not sufficient to assess liver viability.
Topics: Animals; Bile; Biomarkers; Biopsy; Female; Graft Survival; Humans; In Vitro Techniques; Liver; Liver Function Tests; Liver Transplantation; Models, Animal; Perfusion; Swine
PubMed: 33422346
DOI: 10.1016/j.surg.2020.11.042 -
Zentralblatt Fur Chirurgie Aug 2021Machine perfusion of donor livers is currently regarded as the most important innovation in transplant surgery to address the continuing shortage of organs in liver... (Review)
Review
Machine perfusion of donor livers is currently regarded as the most important innovation in transplant surgery to address the continuing shortage of organs in liver transplantation. Hypothermic machine perfusion (HMP) is safe to use and appears to reduce the risk of biliary complications and improve the long-term survival of transplanted organs following preservation by cold static storage - even in donors after cardiac death. A potential functional test of donor organs during HMP uses flavin mononucleotide and is still under clinical investigation. Normothermic machine perfusion (NMP) has a greater risk of technical problems, but functional testing using conventional laboratory parameters during NMP allows significant expansion of the donor pool, even though no prospective randomised study has been able to demonstrate a survival advantage for transplanted organs after NMP. In addition, the preservation time of the donor organs can be significantly extended with the help of NMP, which is particularly advantageous for complex recipient operations and/or logistics. Both methods could be applied for various scenarios in transplantation medicine - theoretically also in combination. The majority of German transplant centres regard machine perfusion as an important innovation and already actively perform perfusions or are in preparation for doing so. However, the overall practical experience in Germany is still relatively low, with only 2 centres having performed more than 20 perfusions. In the coming years, multi-centre efforts to conduct clinical trials and to develop national guidelines on machine perfusion will therefore be indispensable in order to define the potential of these technological developments objectively and to exploit it optimally for the field of transplantation medicine.
Topics: Humans; Liver; Liver Transplantation; Organ Preservation; Perfusion; Prospective Studies; Tissue Donors
PubMed: 33761573
DOI: 10.1055/a-1363-2520 -
Transplant International : Official... Jun 2015Hypothermic machine perfusion (HMP) of kidneys is a long-established alternative to static cold storage and has been suggested to be a better preservation method. Today,... (Review)
Review
Hypothermic machine perfusion (HMP) of kidneys is a long-established alternative to static cold storage and has been suggested to be a better preservation method. Today, as our deceased donor profile continues to change towards higher-risk kidneys of lower quality, we are confronted with the limits of cold storage. Interest in HMP as a preservation technique is on the rise. Furthermore, HMP also creates a window of opportunity during which to assess the viability and quality of the graft before transplantation. The technology might also provide a platform during which the graft could be actively repaired, making it particularly attractive for higher-risk kidneys. We review the current evidence on HMP in kidney transplantation and provide an outlook for the use of the technology in the years to come.
Topics: Cold Temperature; Donor Selection; Drug Delivery Systems; Genetic Therapy; Graft Survival; Humans; Kidney Transplantation; Organ Preservation; Organ Preservation Solutions; Oxygen; Perfusion; Randomized Controlled Trials as Topic; Stem Cell Transplantation; Tissue Donors; Treatment Outcome
PubMed: 25630347
DOI: 10.1111/tri.12530 -
Methods in Molecular Biology (Clifton,... 2018Metabolic reprogramming has been associated to a plethora of diseases, and there has been increased demand for methodologies able to determine the metabolic alterations...
Metabolic reprogramming has been associated to a plethora of diseases, and there has been increased demand for methodologies able to determine the metabolic alterations that characterize the pathological states and help developing metabolically centered therapies. In this chapter, methodologies for monitoring TCA cycle turnover and its interaction with pyruvate cycling and anaplerotic reactions will be presented. These methodologies are based in the application of stable C isotope "tracers"/substrates and C-NMR isotopomer analysis of metabolic intermediates. These methodologies can be applied at several organizational levels, ranging from isolated organelles and organs to whole organisms/humans. For the sake of simplicity, only very simple and well-defined models will be presented, including isolated heart mitochondria and isolated perfused hearts and livers.
Topics: Animals; Carbon Isotopes; Carbon-13 Magnetic Resonance Spectroscopy; Energy Metabolism; Heart; Isomerism; Liver; Mice; Mitochondria, Heart; Perfusion; Proton Magnetic Resonance Spectroscopy; Rats
PubMed: 29851003
DOI: 10.1007/978-1-4939-7831-1_13 -
Multimedia Manual of Cardiothoracic... Mar 2021Ex vivo lung perfusion is an indispensable tool in the armamentarium of any lung transplant center. It helps to increase an already shrinking donor pool by offering a...
Ex vivo lung perfusion is an indispensable tool in the armamentarium of any lung transplant center. It helps to increase an already shrinking donor pool by offering a chance to assess suboptimal donor lungs in a systematic manner and improve them by treating them with low-molecular-weight perfusate. We offer a stepwise guide to carry out ex vivo lung perfusion on the donor lungs and criteria to accept them for transplants.
Topics: Donor Selection; Extracorporeal Circulation; Humans; Lung; Lung Transplantation; Male; Middle Aged; Organ Preservation; Perfusion; Tissue Donors
PubMed: 33904268
DOI: 10.1510/mmcts.2021.018 -
Transplant International : Official... 2022normothermic perfusion (EVNP) is an emerging strategy in kidney preservation that enables resuscitation and viability assessment under pseudo-physiological conditions... (Review)
Review
normothermic perfusion (EVNP) is an emerging strategy in kidney preservation that enables resuscitation and viability assessment under pseudo-physiological conditions prior to transplantation. The optimal perfusate composition and duration, however, remain undefined. A systematic literature search (Embase; Medline; Scopus; and BIOSIS Previews) was conducted. We identified 1,811 unique articles dating from January 1956 to July 2021, from which 24 studies were deemed eligible for qualitative analysis. The perfusate commonly used in clinical practice consisted of leukocyte-depleted, packed red blood cells suspended in Ringer's lactate solution with Mannitol, dexamethasone, heparin, sodium bicarbonate and a specific nutrient solution supplemented with insulin, glucose, multivitamins and vasodilators. There is increasing support in preclinical studies for non-blood cell-based perfusates, including Steen solution, synthetic haem-based oxygen carriers and acellular perfusates with supraphysiological carbogen mixtures that support adequate oxygenation whilst also enabling gradual rewarming. Extended durations of perfusion (up to 24 h) were also feasible in animal models. Direct comparison between studies was not possible due to study heterogeneity. Current evidence demonstrates safety with the aforementioned widely used protocol, however, extracellular base solutions with adequate oxygenation, supplemented with nutrient and metabolic substrates, show promise by providing a suitable environment for prolonged preservation and resuscitation. https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42021231381, identifier PROSPERO 2021 CRD42021231381.
Topics: Animals; Extracorporeal Circulation; Humans; Kidney; Kidney Transplantation; Organ Preservation; Perfusion
PubMed: 35634582
DOI: 10.3389/ti.2022.10236 -
Journal of Visualized Experiments : JoVE May 2021In basic research using mouse heart, isolating viable individual cardiomyocytes is a crucial technical step to overcome. Traditionally, isolating cardiomyocytes from...
In basic research using mouse heart, isolating viable individual cardiomyocytes is a crucial technical step to overcome. Traditionally, isolating cardiomyocytes from rabbits, guinea pigs or rats has been performed via retrograde perfusion of the heart with enzymes using a Langendorff apparatus. However, a high degree of skill is required when this method is used with a small mouse heart. An antegrade perfusion method that does not use a Langendorff apparatus was recently reported for the isolation of mouse cardiomyocytes. We herein report a complete protocol for the improved antegrade perfusion of the excised heart to isolate individual heart cells from adult mice (8 - 108 weeks old). Antegrade perfusion is performed by injecting perfusate near the apex of the left ventricle of the excised heart, the aorta of which was clamped, using an infusion pump. All procedures are carried out on a pre-warmed heater mat under a microscope, which allows for the injection and perfusion processes to be monitored. The results suggest that ventricular and atrial myocytes, and fibroblasts can be well isolated from a single adult mouse simultaneously.
Topics: Animals; Cell Separation; Fibroblasts; Heart Ventricles; Mice; Myocytes, Cardiac; Perfusion
PubMed: 34096909
DOI: 10.3791/61866 -
Artificial Organs Jul 2020Pancreatic static cold storage (SCS) is the gold-standard method for pancreas preservation. Our main objective was to evaluate feasibility of hypothermic perfusion (HP)... (Comparative Study)
Comparative Study
Pancreatic static cold storage (SCS) is the gold-standard method for pancreas preservation. Our main objective was to evaluate feasibility of hypothermic perfusion (HP) of nonhuman primates' pancreases for potential organ transplantation. Seven baboon pancreases were tested. Animals were included in a study approved by the French Research Ministry of Health. Two groups were compared: the control group (n = 2) was preserved using conventional SCS for 24-h and the perfusion group (n = 5) used HP for 24-h, with three different perfusion pressures (PP): 15 (n = 3), 20 (n = 1), and 25 mm Hg (n = 1). In the control group, focal congestion of islets was observed after 6-h. At 24-h, ischemic necrosis and multifocal congestion also occurred. In the HP group, at 15 mm Hg PP, multifocal congestion of islets was present at 24-h. At 20 mm Hg PP, no ischemic necrosis was found after 6-h. At 12-h and 24-h, focal congestion of islets was seen. At 25 mm Hg PP, focal congestion of islets appeared after 12-h. Immunostaining for insulin, glucagon, and somatostatin was normal and similar in controls and perfused pancreas transplants even after 24-h. Apoptosis index represented by cleaved caspase 3 activity, was less than 1% in perfusion and control groups, even after 24-h. HP of nonhuman primate pancreas is feasible and not deleterious as far as 24-h compared to SCS. SCS for more than 12-h was harmful for the transplants. Systolic perfusion pressure between 15-20 mm Hg did not cause any pathological injury of the tested organs.
Topics: Animals; Feasibility Studies; Male; Models, Animal; Necrosis; Organ Preservation; Pancreas; Pancreas Transplantation; Papio; Perfusion; Pressure; Tissue and Organ Harvesting
PubMed: 31995645
DOI: 10.1111/aor.13655 -
Military Medicine Dec 2020Hypothermic ex-situ machine perfusion (MP) has been shown to be a promising alternative to static cold storage (SCS) for preservation of solid organs for transplantation... (Comparative Study)
Comparative Study
INTRODUCTION
Hypothermic ex-situ machine perfusion (MP) has been shown to be a promising alternative to static cold storage (SCS) for preservation of solid organs for transplantation and vascularized composite allotransplantation. Perfusion with blood-based perfusion solutions in austere environments is problematic due to their need for appropriate storage and short shelf life, making it impractical for military and emergency use. Acellular perfusion has been shown to be effective, but the ideal perfusate solution for MP of amputated limbs is yet to be determined. The purpose of this study is to evaluate the efficacy of alternative perfusate solutions, such as dextran-enriched Phoxilium, Steen, and Phoxilium in ex-vivo hypothermic MP of amputated limbs in a porcine model.
MATERIALS AND METHODS
Amputated forelimbs from Yorkshire pigs (n = 8) were preserved either in SCS (n = 2) at 4°C for 12 hours or machine-perfused at 10°C for 12 hours with oxygenated perfusion solutions (n = 6) at a constant flow rate. The perfusates used include modified Steen-solution, Phoxilium (PHOX), or Phoxilium enriched with dextran-40 (PHODEX). The perfusate was exchanged after 1 and 6 hours of perfusion. Machine data were recorded continuously. Perfusate samples for clinical chemistry, blood gas analysis, and muscle biopsies were procured at specific timepoints and subsequently analyzed. In this semi in-vivo study, limb replantation has not been performed.
RESULTS
After amputation, every limb was successfully transferred and connected to our perfusion device. The mean total ischemia time was 77.5 ± 5.24 minutes. The temperature of the perfusion solution was maintained at 10.18 ± 2.01°C, and perfusion pressure at 24.48 ± 10.72 mmHg. Limb weight increased by 3% in the SCS group, 36% in the PHODEX group, 25% in the Steen group, and 58% in the PHOX group after 12 hours. This increase was significant in the PHOX group compared with the SCS group. All perfusion groups showed a pressure increase of 10.99 mmHg over time due to edema. The levels of HIF-1a decreased over time in all groups except the Steen and the PHODEX group. The biomarkers of muscle injury in the perfusate samples, such as creatine kinase and lactate-dehydrogenase, showed a significant difference between groups, with highest values in the PHODEX group. No significant differences were found in the results of the blood gas analysis.
CONCLUSION
With the exception of significantly higher levels of creatine kinase and lactate dehydrogenase, MP with dextran-enriched Phoxilium provides similar results as that of the commercially available perfusates such as Steen, without the need for cold storage, and at circa 5% of the cost of the Steen solution. Further large-scale replantation studies are necessary to evaluate the efficacy of dextran-enriched Phoxilium as an alternate perfusate solution.
Topics: Amputation, Surgical; Animals; Extremities; Organ Preservation; Perfusion; Replantation; Swine
PubMed: 33377496
DOI: 10.1093/milmed/usaa160 -
Journal of Hepatology Jan 2019
Review
Topics: Equipment Design; Humans; Liver Transplantation; Organ Preservation; Perfusion; Practice Guidelines as Topic
PubMed: 30409464
DOI: 10.1016/j.jhep.2018.09.019