Effects of epidermal growth factor on CYP inducibility by xenobiotics, DNA replication, and caspase activations in collagen I gel sandwich cultures of rat hepatocytes

• Published in: Biochemical pharmacology 2001-05, Vol.61 (10), p.1293-1303
• Main Authors: De Smet, Karen, Loyer, Pascal, Gilot, David, Vercruysse, Antoine, Rogiers, Vera, Guguen-Guillouzo, Christiane
• Format: Article
• Language: English
• Subjects: Animals; Apoptosis; Biological and medical sciences; c- fos; c- jun; c- myc; Caspase 3; Caspases - metabolism; Cdk1; Cell cycle; Cell Cycle - drug effects; Cell physiology; Cell Survival - drug effects; Collagen - pharmacology; Cyclin D1; CYP1A1; CYP2B; Cytochrome P-450 CYP1A1 - drug effects; Cytochrome P-450 CYP1A1 - metabolism; Cytochrome P-450 CYP2B1 - drug effects; Cytochrome P-450 CYP2B1 - metabolism; DNA Replication - drug effects; Drug Interactions; Enzyme Activation; Enzyme Induction - drug effects; Epidermal Growth Factor - pharmacology; Fundamental and applied biological sciences. Psychology; Hepatocytes - drug effects; Hepatocytes - enzymology; In Vitro Techniques; Male; Molecular and cellular biology; Rats; Rats, Sprague-Dawley; Responses to growth factors, tumor promotors, other factors; Serum Albumin - metabolism; Xenobiotics - pharmacology
• ISSN: 0006-2952; 1873-2968
• DOI: 10.1016/S0006-2952(01)00612-8

In this study, we investigated the combined effects of EGF and collagen I gel on the phenotype of cultured rat hepatocytes and we focussed our investigations on the regulation of xenobiotic-mediated induction of CYP, cell cycle progression and activation of capases 8 and 3. We found that EGF, added to basal culture medium or phenobarbital (3.2 mM) containing medium, provoked a moderate decrease of CYP1A1 and CYP2B1/2 activities. However, EGF did not exert any inhibitory effect on 3-methylcholantrene (5 μM) and β-naphtoflavone (25 μM) induction of CYP1A1 activities. In collagen gel sandwich cultures, hepatocytes remained arrested in mid-G1 phase of the cell cycle, even in the presence of EGF. In conventional primary cultures, caspases 8 and 3 were activated at 3 and 5 days after plating respectively. In collagen gel sandwich cultures, we found that neither collagen I nor EGF prevented activation of caspase 8 while collagen I gel inhibited activation of caspase 3, preventing spontaneous apoptosis of cultured rat hepatocytes. In contrast, EGF transiently increased caspase 3 activity at day 1 after plating. Altogether, our data demonstrate that collagen I gel triggers intracellular signals which strongly affect cultured hepatocyte phenotype, leading to a cell cycle arrest in G1 phase and long-term survival through the inhibition of caspase 3 activation and that EGF-free medium improves survival and liver-specific gene expression in hepatocytes maintained in collagen I gel sandwich cultures.