Maintenance of monooxygenase activities and detection of cytochrome P-450-mediated cytotoxicity in Mongolian gerbil hepatocyte cultures

Abstract

1. Hepatocytes were isolated from untreated and phenobarbitone (PB)-treated Mongolian gerbils by lobe perfusion. Yields were approx. 20 ± 10⁶ cells/g liver and viability was 95 ± 1%.

2. PB treatment significantly increased the total cytochrome P-450 content, and the 7-ethoxycoumarin O-deethylase, p-nitrophenol hydroxylase and coumarin 7-hydroxylase activities, relative to those of untreated gerbils, measured in homogenates of freshly isolated hepatocytes.

3. After 24 h in culture the cytochrome P-450 content of hepatocyte homogenates from both untreated and PB-treated gerbils was 40–45% that of the corresponding values of freshly isolated hepatocytes. This decrease was accompanied by selective losses of cytochrome P-450-dependent enzyme activities.

4. Erythromycin and benzphetamine N-demethylase, and p-nitrophenol hydroxylase, activities were well maintained over 24 h in culture, whilst 7-ethoxycoumarin O-deethylase and coumarin 7-hydroxylase activities were poorly maintained. In general, the stability of the monooxygenase activities measured was improved by PB treatment of gerbils.

5. The toxicity of coumarin, precocene I and precocene II to gerbil hepatocyte cultures was dose-dependent. Precocene II was significantly more toxic to hepatocytes cultured from PB-treated, compared with untreated, gerbils.

6. Gerbil hepatocyte cultures would seem to be appropriate for investigating species differences in metabolism-mediated cytotoxicity.