Influence of Matrigel-overlay on constitutive and inducible expression of nine genes encoding drug-metabolizing enzymes in primary human hepatocytes

Abstract

1. Previous studies reported that rat hepatocytes overlaid with extracellular matrix components (Matrigel) maintain the expression and responsiveness of drug-metabolizing enzymes. However, whether Matrigel provides similar advantages in human hepatocytes remains largely uncertain.

2. The influence of Matrigel-overlay on the constitutive and phenobarbital- and oltipraz-inducible expression of nine biotransformation enzymes, cytochrome P450s 1A1, 1A2, 2B6, 3A4, and glutathione S-transferases A1, A2, M1, T1, P1, in primary human hepatocytes was evaluated.

3. Hepatocytes from five livers were maintained on a rigid collagen substratum with or without Matrigel overlay and treated for 48 h with two doses of each inducer. Quantitative RT-PCR, and for selected genes, immunoblot and enzyme activity analyses, demonstrated that human hepatocytes overlaid with Matrigel showed consistently higher constitutive and inducible expression of biotransformation genes. 4. Phenobarbital-mediated responsiveness of cytochrome P450 2B6, a potential indicator of hepatocyte differentiation status, was markedly higher in overlaid relative to non-overlaid hepatocytes. 5. It is concluded that an Matrigel overlay facilitates the maintenance and induction of xenobiotic metabolizing enzymes in primary cultures of human hepatocytes.

Key words::

• Extracellular matrix
• Matrigel
• phenobarbital
• oltipraz
• hepatocytes
• human
• cytochrome P450
• glutathione S-transferases