Induction of Necrosis and DNA Fragmentation during Hypothermic Preservation of Hepatocytes in UW, HTK, and Celsior Solutions

Autor: S.L. Abrahamse, Robin Hartman, R.A.F.M. Chamuleau, Thomas M. van Gulik, Pieter Van Runnard Heimel
Přispěvatelé: Tytgat Institute for Liver and Intestinal Research, Surgery
Jazyk: angličtina
Rok vydání: 2003
Předmět:
Male
0301 basic medicine
Adenosine
Necrosis
Cell Culture Techniques
Tetrazolium Salts
lcsh:Medicine
Apoptosis
Cell Count
Disaccharides
Potassium Chloride
Electrolytes
chemistry.chemical_compound
Cryoprotective Agents
0302 clinical medicine
Glutamates
Hypothermia
Induced
Insulin
Mannitol
Cells
Cultured
Caspase 3
Chemistry
Liver Diseases
Glutathione
Caspases
Tissue Transplantation
DNA fragmentation
Female
Trypan blue
medicine.symptom
Cell Division
Programmed cell death
Cell Survival
Allopurinol
Organ Preservation Solutions
Biomedical Engineering
DNA Fragmentation
Models
Biological
Andrology
03 medical and health sciences
Raffinose
Lactate dehydrogenase
medicine
Animals
Histidine
Viability assay
Cryopreservation
Transplantation
L-Lactate Dehydrogenase
lcsh:R
Cell Biology
Thiazoles
Glucose
030104 developmental biology
Hepatocytes
Procaine
030217 neurology & neurosurgery
Zdroj: Cell Transplantation, Vol 12 (2003)
Cell Transplantation, 12(1), 59-68
Cell transplantation, 12(1), 59-68. Cognizant Communication Corporation
ISSN: 1555-3892
0963-6897
Popis: Donor cells can be preserved in University of Wisconsin (UW), histidine-tryptophan-ketoglutarate (HTK), or Celsior solution. However, differences in efficacy and mode of action in preventing hypothermia-induced cell injury have not been unequivocally clarified. Therefore, we investigated and compared necrotic and apoptotic cell death of freshly isolated primary porcine hepatocytes after hypothermic preservation in UW, HTK, and Celsior solutions and subsequent normothermic culturing. Hepatocytes were isolated from porcine livers, divided in fractions, and hypothermically (4°C) stored in phosphate-buffered saline (PBS), UW, HTK, or Celsior solution. Cell necrosis and apoptosis were assessed after 24- and 48-h hypothermic storage and after 24-h normothermic culturing following the hypothermic preservation periods. Necrosis was assessed by trypan blue exclusion, lactate dehydrogenase (LDH) release, and mitochondrial 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) reduction. Apoptosis was assessed by the induction of histone-associated DNA fragments and cellular caspase-3 activity. Trypan blue exclusion, LDH release, and MTT reduction of hypothermically preserved hepatocytes showed a decrease in cell viability of more than 50% during the first 24 h of hypothermic preservation. Cell viability was further decreased after 48-h preservation. DNA fragmentation was slightly enhanced in hepatocytes after preservation in all solutions, but caspase-3 activity was not significantly increased in these cells. Normothermic culturing of hypothermically preserved cells further decreased cell viability as assessed by LDH release and MTT reduction. Normothermic culturing of hypothermically preserved hepatocytes induced DNA fragmentation, but caspase-3 activity was not enhanced in these cells. Trypan blue exclusion, LDH leakage, and MTT reduction demonstrated the highest cell viability after storage in Celsior, and DNA fragmentation was the lowest in cells that had been stored in PBS and UW solutions. None of the preservation solutions tested in this study was capable of adequately preventing cell death of isolated porcine hepatocytes after 24-h hypothermic preservation and subsequent 24-h normothermic culturing. Culturing of isolated and hypothermically preserved hepatocytes induces DNA fragmentation, but does not lead to caspase-3 activation. With respect to necrosis and DNA fragmentation of hypothermically preserved cells, UW and Celsior were superior to PBS and HTK solutions in this model of isolated porcine hepatocyte preservation.
Databáze: OpenAIRE
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