CYP3A4 inducible model for in vitro analysis of human drug metabolism using a bioartificial liver

Hepatology. 2003 Mar;37(3):665-73. doi: 10.1053/jhep.2003.50094.


CYP3A is responsible for approximately 50% of the therapeutic drug-metabolizing activity in the liver. The present study was undertaken to establish the CYP3A4 inducible model for analysis of human drug metabolism using a bioartificial liver composed of the functional hepatocellular carcinoma cell (HCC) line FLC-5. A radial-flow bioreactor (RFB), which is a carrier-filled type bioreactor, was used for 3-dimensional perfusion culture of FLC-5 cells. The CYP3A4 messenger RNA (mRNA) expression level 48 hours after rifampicin treatment in the RBF was approximately 100 times higher than that in a monolayer culture. Western blot analysis also demonstrated an increase in expression of the CYP3A protein. When testosterone, a substrate for CYP3A4, was added to the rifampicin-treated cell culture, 6 beta-hydroxy testosterone as a metabolite was formed. Electrophoretic mobility shift assay (EMSA) with a CYP3A4 ER6 probe demonstrated that relatively high molecular weight complex containing pregnane X receptor (PXR)/retinoid X receptor alpha(RXR alpha), compared with that in the monolayer culture, is possibly generated in the RFB culture of FLC-5 treated with rifampicin. Similarly, the assay with a probe of HNF-4 alpha-binding motif indicated the formation of a large protein complex in the RFB culture. Because it is known that PXR transactivates CYP3A4 gene via its response element and expression of PXR is regulated by HNF-4 alpha, the large complexes binding to response elements of PXR or HNF-4 alpha in the RFB culture may contribute to up-regulation of CYP3A4 mRNA. In conclusion, the bioartificial liver composed of human functional HCC cell line was useful in studying drug interactions during induction of human CYP3A4.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Basic Helix-Loop-Helix Leucine Zipper Transcription Factors
  • Bioreactors
  • Carcinoma, Hepatocellular
  • Chromatography, High Pressure Liquid
  • Cytochrome P-450 CYP3A
  • Cytochrome P-450 Enzyme System / biosynthesis*
  • Cytochrome P-450 Enzyme System / genetics
  • DNA-Binding Proteins*
  • Electrophoretic Mobility Shift Assay
  • Enzyme Induction
  • Gene Expression
  • Gene Expression Regulation / drug effects
  • Hepatocyte Nuclear Factor 4
  • Humans
  • Liver / enzymology*
  • Liver Neoplasms
  • Microscopy, Electron
  • Microsomes, Liver / enzymology
  • Models, Biological*
  • Pharmaceutical Preparations / metabolism*
  • Phosphoproteins / pharmacology
  • Polymerase Chain Reaction
  • Pregnane X Receptor
  • RNA, Messenger / analysis
  • Receptors, Cytoplasmic and Nuclear / genetics
  • Receptors, Steroid / genetics
  • Rifampin / pharmacology
  • Testosterone / metabolism
  • Transcription Factors / pharmacology
  • Tumor Cells, Cultured


  • Basic Helix-Loop-Helix Leucine Zipper Transcription Factors
  • DNA-Binding Proteins
  • Hepatocyte Nuclear Factor 4
  • MLX protein, human
  • Pharmaceutical Preparations
  • Phosphoproteins
  • Pregnane X Receptor
  • RNA, Messenger
  • Receptors, Cytoplasmic and Nuclear
  • Receptors, Steroid
  • Transcription Factors
  • Testosterone
  • Cytochrome P-450 Enzyme System
  • CYP3A protein, human
  • Cytochrome P-450 CYP3A
  • CYP3A4 protein, human
  • Rifampin