Z Gastroenterol 2007; 45 - A2_13
DOI: 10.1055/s-2007-967803

Comparing cytochtome P450 activities of blood monocyte derived Neo-Hepatocytes and primary hepatocytes

S Ehnert 1, A Müller 2, P Godoy 2, U Böcker 2, S Siegmund 2, M Singer 2, K Breitkopf 2, A Nüssler 3, S Dooley 2
  • 1Abt. Mol. Alkoholforschung Gastroenterologie, Medizin II, Universitätskrankenhaus Mannheim, Universität Heidelberg, Mannheim
  • 2Dept. of Medicine II, Mol. Alcohol Research in Gastroenterology, University Hospital Mannheim, University of Heidelberg, Germany, Mannheim
  • 3Dept. of Cell therapy, Fresenius Biotech GmbH, Bad Homburg, Germany

The human body is constantly under assault by chemicals encountered in the environment. Altering their molecular structure to increase polarity is essential for the elimination from the body. Most bio-activation reactions are catalyzed by a group of hepatic microsomal monooxygenases. Its reaction specificity is defined by cytochrome P450 (CYP), a family of structurally related hemoproteins. Thus, CYPs are essential for the determination of pharmacologic and toxic effects, which are conventionally measured in primary human hepatocytes, whose availability is limited due to donor organ scarcity. This raises the demand for alternatives. Hepatocyte-like (NeoHep) cells generated by trans-differentiation of monocytes offer great perspectives.

NeoHep cells were prepared from peripheral blood monocytes. These cells were compared to primary hepatocytes in terms of their morphology and metabolic behavior. CYP (1A2, 2A6, 2B6, 2C9, 2E1, and 3A4) expression and activity was investigated by RT-PCR, Westernblot, and fluorescence-based microscale activity assays.

After 13–15 days of differentiation NeoHep cells form a confluent layer with cell-cell contact displaying the typical hexagonal shape of primary hepatocytes, expressing hepatocyte marker genes like transferrin and connexin 32. The basal CYP activity is increasing throughout differentiation, reaching a stable level after approximately 15 days. Over a culture period of 5 days primary mouse hepatocytes undergo typical morphologic changes in collagen monolayer culture and basal activity of all CYP isoforms decreases. Responsiveness to inducers (e.g. 3-methylcholanthrene) and inhibitors (e.g. Nifedipine) of CYP activity is comparable in both cell types.

Our data reveal phenotypic and metabolic similarities of NeoHep cells and primary hepatocytes. Hence NeoHep cells might be used as an alternative for primary hepatocates in measuring bioactivation of substances.