Summary
Primary hepatocyte cultures have been used to evaluate data concerning hypoxic liver cell injury. To show the suitability of this method in liver preservation studies hepatocyte cultures were incubated under different conditions: warm normoxia (37°C, pO2 > 70 mm Hg), warm hypoxia (37°C, pO2 < 0.1 mm Hg), cold normoxia (4°C, pO2 > 70 mm Hg) and cold hypoxia (4°C, pO2 < 0.1 mmHg). Incubations were performed in Euro Collins solution (EC), University of Wisconsin solution of Belzer (UW) and histidine ketoglutarat tryptophan solution of Bretschneider (HTK) as well as in Krebs Henseleit buffer (KH) for control incubations. During 12 h of incubation hepatocyte cultures under warm normoxia lost viability continously in EC, UW and HTK while in KH they remained stable. Under warm normoxia all cultures lost 50% of their viability during 12 h of incubation while in cold normoxia loss of viability was mild but significant. Under cold anoxia which is the standard condition of liver preservation the cultured hepatocytes remained unchanged for 12 h in KH, UW and HTK, while in EC most of the cells were dead after 6 h. It is concluded that incubations of primary hepatocyte cultures under different pO2 and temperatures are well suited to contribute to liver preservation studies on a preclinical level and thus may help to save animal experiments.
Zusammenfassung
Primäre Leberzellkulturen haben bisher bei biochemischen Untersuchungen zum Pathomechanismus des hypoxischen Leberzellschadens Anwendung gefunden. Um die Eignung der Methode für Untersuchungen zur Leberkonservierung zu prüfen, wurden Leberzellkulturen bei normothermer Normoxie (37°C, pO2 > 70 mm Hg), normothermer Hypoxie (37°C, pO2 < 0,1 mm Hg), hypothermer Normoxie (4°C, pO2 > 70 mm Hg) and hypothermer Hypoxie (4°C, pO2 < 0,1 mm Hg) inkubiert. Bei jeder Inkubationsbedingung wurde Euro-Collins-Lösung (EC), University-of-Wisconsin-Lösung nach Belzer (UW) and Histidin-Tryptophan-Ketoglutarat-Lösung nach Bretschneider (HTK) als Inkubationsmedium verwendet; Kontrollinkubationen erfolgten in Krebs-Henseleit-Puffer (KH). Das Absterbeverhalten der kultivierten Hepatozyten zeigte während 12stündiger Inkubation einen raschen Viabilitätsverlust bei warmer Normoxie in EC, HTK and UW. In KH blieben die Zellen nahezu vollstandig vital. Bei normothermer Hypoxie starben die Leberzellen in allen Methen während der 12stündigen Inkubation zu mindestens 50% ab, während bei hypothermer Normoxie der Vitalitätsverlust deutlich verlangsamt war. Hypotherme Hypoxie schließlich zeigte in KH, HTK and UW eine nahezu vollständig erhaltene Zellviabilität ℏer 12 h, während in EC nach 6 h die meisten Zellen abgestorben waren. Durch diese Ergebnisse zeigt sich die Methode der Inkubation von primären Leberzellkulturen unter verschiedenen Sauerstoffpartialdrucken und Temperaturen als geeignet zur Evaluierung von Konservierungsmedien und ihrer Komponenten in der präklinischen Phase unter Einsparung von Tierversuchen.
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Viebahn, R., de Groot, H., Lauchart, W. et al. Primäre hepatozytenkulturen als modell zur experimentellen untersuchung der leberkonservierung. Langenbecks Arch Chir 376, 268–272 (1991). https://doi.org/10.1007/BF00188266
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DOI: https://doi.org/10.1007/BF00188266