Effect of extracellular matrix on the expression of peroxisome proliferation associated genes in cultured rat hepatocytes

Abstract

The purpose of this study was to determine whether the extracellular matrix used in hepatocyte culture would alter gene expression induced by the peroxisome proliferator ciprofibrate (CIP). We compared the activities and mRNA levels of two enzymes associated with peroxisome proliferation—fatty acyl-CoA oxidase (FAO), the first enzyme in the peroxisomal β-oxidation pathway, and lauric acid hydroxylase (LAH), which represents the activity of the cytochrome P450 4A subfamily—in rat hepatocytes cultured on different plates: plastic, collagen-coated, thin and thick Matrigel, and collagen gel plates. CIP increased FAO activity about fivefold in collagen gel plates and sixfold in thick Matrigel plates compared to a fourfold increase in other plates; LAH was increased about threefold in thin Matrigel plates and fourfold in thick Matrigel and collagen gel plates compared to only a twofold increase in plastic and collagen-coated plates. The mRNA level for FAO was highest in hepatocytes cultured on collagen gel and Matrigel plates compared to those cultured on plastic and collagen-coated plates. The P-450 4A1 mRNA expression, however, was highest in the collagen gel plates, with lower expression in the thick Matrigel, collagen-coated and plastic plates. DNA synthesis and the DNA binding activity of the transcription factor AP-1 were also examined in response to epidermal growth factor (EGF) and CIP. Without the addition of EGF, DNA synthesis was significantly higher on collagen-coated plates than on collagen gel plates. The DNA binding activity of AP-1 was also induced after 24 hr culture in collagen-coated plates, whereas it was not detected in collagen gel plates. After the addition of EGF, the DNA binding activity of AP-1 was increased in both collagen-coated plates and collagen gel plates. CIP did not increase the DNA binding activity of AP-1 in either plate. These results demonstrate that components of the extracellular matrix influence the induction of peroxisome proliferator-induced enzyme activities and mRNA levels by CIP, with the highest induction seen in collagen gel and thick Matrigel plates. Furthermore, the induction of cell proliferation and AP-1 DNA binding activity are influenced by the extracellular matrix.