ATLA::Alternatives to Laboratory Animals

Volume 29, Number 2

Toxic effects of chromium acetate hydroxide on cells cultivated in vitro.

ATLA 29, 179-192, March/April 2001

Karen De Smet,¹ Christophe Cavin,² Antoine Vercruyesse¹ and Vera Rogiers¹

¹Department of Toxicology, Vrije Universiteit Brussel, Laarbeeklaan 103, 1090 Brussels, Belgium; ²Nestec Ltd, Nestlé Research Centre, Vers-chez-les-Blanc, 1000 Lausanne 26, Switzerland

SUMMARY

Albumin secretion, expression of cytochrome P450-dependent mono-oxygenases (CYPs) and their inducibility by well-known inducers were evaluated during 1 week in collagen type I gel sandwich and immobilisation cultures of adult primary rat hepatocytes. Albumin secretion increased during culture and, following an initial decrease, CYP biotransformation activities remained stable for at least 7 days. Better preservation results were observed in the collagen gel sandwich culture than in the immobilisation model. The inducibility of Cyps by &zlig;-naphthoflavone (β-NF), 3-methylcholanthrene (3-MC), phenobarbital (PB) and dexamethasone (DEX) was studied in both collagen gel hepatocyte cultures. Exposure of the cells to either 5 µM 3-MC or 25 µM β-NF, added to the culture medium. resulted in strong increases of CYP1A1/2 activity in both culture models. Treatment with PB (3.2 mM) resulted in an increase in the CYP2B activitv and a higher hydroxylation of testosterone in the 16(alpha)-position (CYP2B1/2 and CYP2C11), the 7(alpha)-position (CYP2A1/2), and the 6β-position (CYP3Al). DEX (10 µM) markedly increased testosterone 6β- and 7(alpha)-hydroxylation. Expression and induction experiments on CYP proteins exposed to these molecules confirmed the results of the CYP activity measurements. The patterns of CYP induction in collagen gel cultures of rat hepatocytes were similar to those observed in vivo. Consequently, collagen gel cultures and, more specifically, collagen gel sandwich cultures seem to be suitable as in vitro models for evaluating xenobiotics as potential inducers of CYP-enzymes.

Keywords: sandwich culture, immobilisation culture, phase 1 biotransformation, in vitro, liver cell culture, phenobarbitol, dexamethasone, 3-methylcholanthrene, β-naphthoflavone