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Management of Ischemia and Brain Death-Associated Injuries in Porcine Kidney Grafts
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Surgical Sciences, Transplantation Surgery.
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Organs from deceased donors after brain death (BD) remain the major source of organs for transplantation. The catastrophic event of BD and the inevitable consequences of ischemia reperfusion injury (IRI) are linked to impaired graft quality and transplantation outcome. The aim of this thesis was to create a BD model in pigs to assess early effects on IRI in kidneys preserved with an oxygenated solution and to evaluate the protective effects of coating the renal vessel walls with a heparin conjugate during hypothermic machine perfusion (HMP).

Brain death was achieved by raising the intracranial pressure (ICP) through stepwise increasing the volume of an epidurally placed balloon to the point of exceeding the mean arterial pressure (MAP) creating a negative cerebral perfusion pressure (CPP). This reproducible, clinically relevant experimental model makes evaluation of potential targeted methods to protect the organs possible. Kidneys retrieved from brain-dead pigs were preserved either in an oxygenated emulsion composed of 75% histidine-tryptophan-ketoglutarate (HTK) and 25% perfluorohexyloctane F6H8 or HTK alone. After 18h of cold storage the kidneys were transplanted into allogeneic pigs. F6H8 was associated with replenishment of adenosine triphosphate and lower gene expression of hypoxia inducible factor (HIF)-1a, vascular endothelial growth factor (VEGF), interleukin (IL)-1α and tumour necrosis factor (TNF)-α. F6H8 reduced early IRI at both the cellular and molecular level.

Kidneys from BD pigs were evaluated for the feasibility of coating the vessel walls with the heparin conjugate CHC (Corline Systems AB, Uppsala, Sweden) to restore glycocalyx. Porcine kidneys were preserved by HMP for 20h with 50 mg biotinylated CHC added to the perfusion solution. CHC was detected on the inner surface of the kidney vessels by immunofluorescence, and its uptake in kidneys was confirmed by reduced content in the perfusate. An ex vivo normothermic perfusion circuit was developed to assess kidney function. Perfusion with CHC during HMP was associated with lower creatinine levels, increased urine volume and reduced tubular injury. Modifying renal vessels walls using CHC during HMP improved early graft function. Preservation with the oxygenated F6H8 solution or CHC could be used to improve graft quality and ameliorate IRI in kidneys retrieved from deceased donors.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2014. , 77 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 993
National Category
URN: urn:nbn:se:uu:diva-222020ISBN: 978-91-554-8939-7OAI: oai:DiVA.org:uu-222020DiVA: diva2:711319
Public defence
2014-05-31, Gustavianum Auditorium Minus, Akademigatan 3, uppsala, 09:15 (Swedish)
Available from: 2014-05-08 Created: 2014-04-07 Last updated: 2014-06-30
List of papers
1. Standardized experimental brain death model for studies of intracranial dynamics, organ preservation, and organ transplantation in the pig
Open this publication in new window or tab >>Standardized experimental brain death model for studies of intracranial dynamics, organ preservation, and organ transplantation in the pig
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2011 (English)In: Critical Care Medicine, ISSN 0090-3493, E-ISSN 1530-0293, Vol. 39, no 3, 512-517 p.Article in journal (Refereed) Published
Abstract [en]

OBJECTIVES:: Brain death impairs organ function and outcome after transplantation. There is a need for a brain death model to allow studies of organ viability and preservation. For neurointensive care research, it is also of interest to have a relevant brain death model for studies of intracranial dynamics and evaluation of cerebral monitoring devices. Therefore, the objective was to develop a standardized clinically relevant brain death model. METHODS:: Six pigs of both sexes (10-12 wks old; mean weight, 24.5 ± 1.4 kg) were included. Mean arterial blood pressure, heart rate, intracranial pressure, intracranial compliance, cerebral perfusion pressure, and brain tissue oxygenation (BtiPo2) were recorded during stepwise elevation of intracranial pressure by inflation of an epidural balloon catheter with saline (1 mL/20 mins). Brain death criteria were decided to be reached when cerebral perfusion pressure was <0 mm Hg for 60 mins and at least 10 mL saline was inflated epidurally. BtiPo2 and arterial injections of microspheres were used for confirmation of brain death. RESULTS:: A gradual volume-dependent elevation of intracranial pressure was observed. After 10 mL of balloon infusion, mean intracranial pressure was 89.8 ± 9.7 (sd) mm Hg. Intracranial compliance decreased from 0.137 ± 0.069 mL/mm Hg to 0.007 ± 0.001 mL/mm Hg. The mean arterial pressure decreased and the heart rate increased when the intracranial volume was increased to between 5 and 6 mL. All animals showed cerebral perfusion pressure ≤0 after 7 to 10 mL of infusion. In all animals, the criteria for brain death with negative cerebral perfusion pressure and BtiPo2 ∼0 mm Hg were achieved. Only a negligible amount of microspheres were found in the cerebrum, confirming brain death. The kidneys showed small foci of acute tubular necrosis. CONCLUSIONS:: The standardized brain death model designed in pigs simulates the clinical development of brain death in humans with a classic pressure-volume response and systemic cardiovascular reactions. Brain death was convincingly confirmed.

brain death, experimental animal model, intracranial pressure, cerebral perfusion pressure, brain tissue oxygenation, organ preservation
National Category
Medical and Health Sciences
urn:nbn:se:uu:diva-143532 (URN)10.1097/CCM.0b013e318206b824 (DOI)000287480000013 ()21187748 (PubMedID)
Available from: 2011-01-21 Created: 2011-01-21 Last updated: 2015-06-15Bibliographically approved
2. Oxygen-charged HTK-F6H8 emulsion reduces ischemia: reperfusion injury in kidneys from brain-dead pigs
Open this publication in new window or tab >>Oxygen-charged HTK-F6H8 emulsion reduces ischemia: reperfusion injury in kidneys from brain-dead pigs
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2012 (English)In: Journal of Surgical Research, ISSN 0022-4804, E-ISSN 1095-8673, Vol. 178, no 2, 959-967 p.Article in journal (Refereed) Published
Abstract [en]


Prolonged cold ischemia is frequently associated with a greater risk of delayed graft function and enhanced graft failure. We hypothesized that media, combining a high oxygen-dissolving capacity with specific qualities of organ preservation solutions, would be more efficient in reducing immediate ischemia-reperfusion injury from organs stored long term compared with standard preservation media.


Kidneys retrieved from brain-dead pigs were flushed using either cold histidine-tryptophan-ketoglutarate (HTK) or oxygen-precharged emulsion composed of 75% HTK and 25% perfluorohexyloctane. After 18 h of cold ischemia the kidneys were transplanted into allogeneic recipients and assessed for adenosine triphosphate content, morphology, and expression of genes related to hypoxia, environmental stress, inflammation, and apoptosis.


Compared with HTK-flushed kidneys, organs preserved using oxygen-precharged HTK-perfluorohexyloctane emulsion had increased elevated adenosine triphosphate content and a significantly lower gene expression of hypoxia inducible factor-1 alpha, vascular endothelial growth factor, interleukin-1 alpha, tumor necrosis factor-alpha, interferon-alpha, JNK-1, p38, cytochrome-c, Bax, caspase-8, and caspase-3 at all time points assessed. In contrast, the mRNA expression of Bcl-2 was significantly increased.


The present study has demonstrated that in brain-dead pigs the perfusion of kidneys with oxygen-precharged HTK-perfluorohexyloctane emulsion results in significantly reduced inflammation, hypoxic injury, and apoptosis and cellular integrity and energy content are well maintained. Histologic examination revealed less tubular, vascular, and glomerular changes in the emulsion-perfused tissue compared with the HTK-perfused counterparts. The concept of perfusing organs with oxygen-precharged emulsion based on organ preservation media represents an efficient alternative for improved organ preservation.

Brain death, Emulsion, Kidney transplantation, Organ preservation, Oxygen carrier, Pig
National Category
Medical and Health Sciences
urn:nbn:se:uu:diva-188407 (URN)10.1016/j.jss.2012.06.031 (DOI)000311090700064 ()

De två första författarna delar förstaförfattarskapet.

Available from: 2012-12-18 Created: 2012-12-17 Last updated: 2014-06-30Bibliographically approved
3. Modifying the vessel walls in porcine kidneys during machine perfusion
Open this publication in new window or tab >>Modifying the vessel walls in porcine kidneys during machine perfusion
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2014 (English)In: Journal of Surgical Research, ISSN 0022-4804, E-ISSN 1095-8673, Vol. 191, no 2, 455-462 p.Article in journal (Refereed) Published
Abstract [en]

Background: Endothelial glycocalyx regulates the endothelial function and plays an active role in maintaining vascular homeostasis. During ischemia/reperfusion, the glycocalyx is rapidly shed into the blood stream. A heparin conjugate (CHC; Corline systems AB, Uppsala, Sweden) consists of 70 heparin molecules that have the capacity to adhere strongly to biological tissues expressing heparin affinity. We hypothesized that CHC could be used to restore disrupted glycocalyx in vivo in kidneys from brain-dead pigs.

Materials and Methods: Brain death was induced in male landrace pigs (n=6) by inflating a balloon catheter in the epidural space until obtaining negative cerebral perfusion. The recovered kidneys (n=5+5) were perfused by hypothermic machine perfusion (HMP) using two Lifeport kidney transporters (Organ Recovery Systems, Chicago, IL, USA). 50 mg CHC (including 25 mg biotinylated CHC) or 50 mg unfractionated heparin (control) was added to the perfusion fluid in the respective machines. In one case, the kidneys were used only for dose escalation of CHC with the same procedure.

Results: CHC was detected by immunofluorescence and confocal microscopy in the inner surface of vessel walls. The binding of CHC in the kidney was confirmed indirectly by consumption of CHC from the perfusion fluid.

Conclusions: In this first attempt, we show that CHC may be used to coat the vessel walls of perfused kidneys during HMP, an approach that could become useful in restoring endothelial glycocalyx of kidneys recovered from deceased donors to protect vascular endothelium and possibly ameliorate ischemia/reperfusion injuries.

National Category
urn:nbn:se:uu:diva-221875 (URN)10.1016/j.jss.2014.04.006 (DOI)000341358100027 ()24819743 (PubMedID)
Available from: 2014-04-07 Created: 2014-04-06 Last updated: 2014-10-28Bibliographically approved
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