Abstract
The present study was undertaken to evaluate the use precision-cut liver slices as a system for studying xenobiotic metabolism. Comparison of two culture systems, namely the dynamic organ culture system (DOCS), and the multiwell plate (12-well) system, using morphological and biochemical parameters, showed that the 12-well plate system is superior to the DOCS. The present study also demonstrates that increasing the number of slices from one to two per incubation does not lead to a proportional increase in metabolic activity, and this approach is, therefore, not suitable for scaling up the system. Cytochrome P450 activity was lost in precision-cut rat and human liver slices during long culture periods with a substantial loss occurring during the first 24 hours of culture, but the rate of decline was isoform-specific. The loss in cytochrome P450 activities was not always mirrored by their apoprotein levels, the majority of which were still detectable even after 72 hours of culture. Determination of cytosolic and microsomal protein content revealed that the latter declined more rapidly in culture as compared with the cytosolic protein. The rat and human cytosolic Phase II drug metabolising enzymes were far more stable than the microsomal Phase II enzymes and the Phase I cytochromes P450 in long-term culture. Although the activity of all enzymes declined with culture time, the rate of loss differed among the various enzymes. Histological examination of commercially available human liver slices showed that the cells were preserved well at time of receipt and during culture. Human liver slices metabolised 7-ethoxycoumarin for up to 72 hours, although there was a significant decline in the levels of metabolites after 24 hours of culture. A large inter- and intra-variation was also noted. Precision-cut liver slices were finally used to investigate the role of individual cytochrome P450 isoforms in the metabolism of the endogenous substrate, melatonin. The present study clearly demonstrates that the CYP1A family is responsible of the metabolism of exogenous levels of melatonin.