[Addition of an osmotic agent to liver preservative solutions in a model of in vitro preservation of hepatocytes]

Abstract

To avoid hypoxic cell swelling during liver preservation followed by reduced perfusion in the reoxygenation period, osmotic substances such as mannitol, sucrose and raffinose, and the impermeant anion lactobionate are used in established liver preservation solutions. The various osmotic agents were investigated at concentrations of 60, 140, 260 and 300 mM, the solutions being kept isotonic by substitution with sodium and potassium chloride to 300 mosmol/l. Cultures of adherent pig hepatocytes were incubated in an in vitro model of cold hypoxia (4 degrees C, PO2 < 0.1 mmHg) for 24 h and reoxygenated with standard culture medium for 3 h. After each incubation period, light microscopy was performed to estimate cell viability and detachment rate. LDH and GOT liberation were also measured. To estimate the change in cell volume, isolated hepatocytes were incubated in suspension for 24 h of cold hypoxia. The cell volumes were compared after centrifugation and measurement of the pellet and the solute levels. Rising concentrations of osmotic substances resulted in increasing liberation of LDH and GOT. The levels of LDH and GOT release from cultures incubated with 60 mmol/l sucrose or raffinose were comparable to those in a preservation solution of "extracellular" ion composition. Addition of mannitol to the preservation solution resulted in cell damage. At high concentrations, sucrose did not affect the hepatocytes as much as raffinose. While mannitol can permeate the hepatocytes and lead to cell swelling, a cell-shrinking effect was observed when sucrose was used, and even more pronounced cell shrinking was seen with raffinose, to which the hepatocyte membrane is known to be permeable.(ABSTRACT TRUNCATED AT 250 WORDS)