Determination of Optimal Incubation Media and Suitable Slice Diameters in Precision-Cut Liver Slices: Optimization of Tissue Slice Culture, Part 2

Abstract

To validate tissue slice methodology in experiments from one laboratory to another, it is important to optimize slice viability. The effects of different culture media and slice diameters were investigated in this study. Rat and mouse liver slices (200 μm thick) were placed in either Krebs/Hepes, Kxebs/bicarbonate, Waymouth's/Hepes, or Waymouth's/bicarbonate to assess the effects of nutrient-enriched and nutrient-deprived media as well as the effects of a Hepes-or bicarbonate-based medium on slice viability. Overall, an enriched and bicarbonate-based medium maintained slice viability better. Determination of suitable slice diameters was achieved by incubating 6- and 8-mm-diameter rat, mouse, and dog liver slices in Waymouth's/bicarbonate media for 24 h. For both rat and mouse liver slices, the 8-mm-diameter slices were better maintained than the 6-mm slices, whereas both diameters were equivalent for dog liver slices. Various nutrient-enriched media were also compared for their ability to maintain rat liver slice viability in culture. Of the 15 media tested, nine maintained slice K⁺ levels above 80 μmol/g wet weight for 3 days. Rat liver slice viability could be maintained for 5 days in culture when 8-mm slices were incubated in Waymouth's/bicarbonate medium in dynamic organ culture. Liver slice viability was assessed by measuring K⁺ retention, protein synthesis, and lactate dehydrogenase leakage. This study shows that incubation parameters do affect liver slice viability and that these parameters need to be optimized.

Key Words: :

• Optimal slice viability
• Incubation media
• Slice diameter