STUDIES ON THE ELECTROLYTE-COMPOSITION OF LIVER PRESERVATION SOLUTIONS USED FOR HEPATOCYTES IN AN IN-VITRO PRESERVATION AND REOXYGENATION MODEL

The cation and anion content in liver preservation solutions have been investigated in order to justify the use of ''intracellular'' or ''extracellular'' electrolyte compositions. Various concentrations of sodium and potassium with chloride or lactobionate as anions and with added calcium and/or magnesium were made up as preservation solutions and incubated with in vitro adherent cultures of pig hepatocytes. In vitro hypoxia and hypothermia (4-degrees-C, PO2 < 0.1 mmHg) for 24 h, with reoxygenation for 3 h, in standard culture medium was used as a model for preservation. Measurements of cell viability and detachment rate by light microscopy and of LDH and GOT liberation were used as parameters of cell damage. Cell swelling was estimated in suspension cultures of isolated hepatocytes. When chloride was used as the anion, significant cell toxicity from potassium concentrations over 75 mM was found within 6 h of preservation. Enzyme liberation decreased with increasing content of sodium cations in the preservation solution. Calcium ions had a protective effect at a concentration of 0.8 mM. Addition of magnesium to an ''intracellular'' ion composition minimized the toxic effect of potassium cations. Using lactobionate as an impermeant anion, there was no difference between the sodium and the potassium salt and the choice of cation had no effect on enzyme leakage or cell volume. An ''extracellular'' solution with high sodium chloride content and 0.8 mM calcium resulted in better preservation than was obtained with lactobionate solutions. With chloride as the anion, a significant increase in cell swelling was found when potassium replaced sodium in the solutions. Cell swelling decreased with increasing concentration of sodium cations. Calcium and magnesium ions had no effect on the cell volume. When the impermeant lactobionate was used as the anion a ce shrinking effect was observed, which was independent of the choice of cation. High potassium chloride concentrations in preservation solutions lead to depolarization of the hepatocyte membrane, followed by a flux of chloride anions and water into the cells. Reduced activity of Na-K-ATPase results in less cell swelling when an ''extracellular'' solution with high sodium chloride content is used. The influx of permeable anions seems to be responsible for hypoxic cell swelling and cell alteration, because when an impermeant anion is used the choice of cation does not affect the outcome of preservation.