An in vitro model for peroxisome proliferation utilizing primary hepatocytes in sandwich culture

Abstract

Peroxisome proliferators comprise a group of structurally diverse chemicals which share as a common biologic effect the induction of peroxisomal fatty acid degrading enzymes. Concomitantly, the number and size of peroxisomes within hepatocytes increases. Following chronic administration some peroxisome proliferators act as non-genotoxic hepatocarcinogens in susceptible species such as rodents. To establish an in vitro model for the toxicological investigation of peroxisome proliferation, primary hepatocytes of rats, dogs and humans were cultivated in an organotypic cell culture model (sandwich model). By employing a panel of diverse compounds in this model a graded response was observed in the induction of carnitine acetyl transferase (CAT), the activity of which was determined as an endpoint. The following results were obtained in the order of decreasing inducing potential for rat hepatocytes: FOE 3798>nafenopin>fenofibrate (<clofibrate>ciprofibrate>bezafibrate ⪢ DEHP≈ETYA>DEHA. Induction of CAT activity was generally higher than reported in earlier cell culture systems, probably reflecting the effect of the extracellular matrix provided by the collagen gel sandwich. In parallel, transcription of the rat CYP4A1 gene was induced by a similar order of magnitude as measured by TaqMan RT-PCR. In accordance with literature data, human and dog hepatocytes did not display such a strong and graded response but rather were not susceptible to this effect. In addition, 3H-thymidine incorporation data demonstrated that nafenopin was able to induce DNA synthesis in rat hepatocytes whereas it did not in human hepatocytes.