Influence of Matrigel-overlay on constitutive and inducible expression of nine genes encoding drug-metabolizing enzymes in primary human hepatocytes.
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.
Gross-Steinmeyer
K
K
Department of Environmental and Occupational Health Sciences, Center for Ecogenetics & Environmental Health, University of Washington, Seattle, WA 98105, USA.
Stapleton
P L
PL
Tracy
J H
JH
Bammler
T K
TK
Lehman
T
T
Strom
S C
SC
Eaton
D L
DL
D000328:Adult; D000818:Animals; D001711:Biotransformation; D003094:Collagen; D019363:Cytochrome P-450 CYP1A1 / Q000378:metabolism; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D004338:Drug Combinations; D004790:Enzyme Induction; D015971:Gene Expression Regulation, Enzymologic; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D022781:Hepatocytes / Q000201:enzymology*; D006801:Humans; D007797:Laminin; D008875:Middle Aged; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D011509:Proteoglycans; D012377:Rodentia
D004338:Drug Combinations; D007797:Laminin; D004364:Pharmaceutical Preparations; D011509:Proteoglycans; C059485:matrigel; D003094:Collagen; D003577:Cytochrome P-450 Enzyme System; D019363:Cytochrome P-450 CYP1A1; D005982:Glutathione Transferase