Application of isolated hepatocytes to studies of drug metabolism in large food animals

Abstract

A definitive hazard assessment of xenobiotics translocated through food animals into edible products such as meat or milk requires a complete analysis of metabolism in food animals. However, large animal metabolism studies present many experimental difficulties. None of several in vitro alternatives such as subcellular fractions has been established as an acceptable predictor of in vivo metabolism. The feasibility of using isolated hepatocytes to predict the metabolism of xenobiotics, both quantitatively and qualitatively, in large ruminant animals (e.g. cattle) is being studied in our laboratory. A procedure was developed for isolating hepatocytes aseptically from the caudate process of the liver which was obtained surgically from 100-125 kg calves. A modified two-step vascular perfusion procedure provides hepatocyte suspensions that are typically greater than or equal to 85% viable and greater than or equal to 1 X 10(7) viable hepatocytes/g of liver (wet wt). Xenobiotic metabolism has been evaluated in suspensions and primary cultures using aldrin epoxidation, ethoxycoumarin O-deethylation, and 7-hydroxycoumarin glucuronidation and sulfation. Metabolic activities are relatively short-lived in suspensions less than or equal to 4 h, but quite stable up to 10 h when cultured on collagen-coated plates in chemically defined medium. Bovine hepatocytes behave similarly in culture to rodent hepatocytes. Although primary culturing of hepatocytes is more difficult than suspensions, primarily due to the asepsis requirements, it is the method of choice for xenobiotic metabolism determinations in isolated hepatocytes of cattle.