Alkoxyresorufin O-dealkylase assay using a rat hepatocyte spheroid microarray.

Hepatocyte multicellular aggregates (spheroids), which maintain high expression of liver functions, have been advocated as a useful culture technique for various cell-based assays. In this study, we investigated the drug metabolic function of a hepatocyte spheroid microarray (HSM) chip, which contained an array of 672 spheroids of primary rat hepatocytes within a 100-mm(2) region in the center of a poly(methylmethacrylate) plate (24 x 24 mm) and used an alkoxyresorufin (ethoxy-, methoxy-, pentoxy- and benzyloxyresorufin) O-dealkylase assay system. Ethoxyresorufin O-dealkylase (EROD) activity of the HSM chip initiated by 3-methylcholanthrene (3-MC), an inducer of cytochrome P450 enzymes, was 5- to 10-fold higher than that of monolayer hepatocytes, with activity being maintained for at least 2 weeks. We also demonstrated that 3-MC induced EROD, methoxyresorufin O-dealkylase (MROD) and benzyloxyresorufin O-dealkylase (BROD) activities in the HSM chip, while sodium phenobarbital (P450 inducer) induced pentoxyresorufin O-dealkylase (PROD), BROD, EROD and MROD activities. Induction of these activities was confirmed by increased gene expression of the related P450 enzymes. These results showed that the HSM chip had a good response to P450 inducers and that function was maintained for long periods of time. The HSM chip therefore may be a promising cellular platform for drug metabolic assays using hepatocytes. Copyright 2009 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Citation

Yusuke Sakai, Tomoko Tanaka, Junji Fukuda, Kohji Nakazawa. Alkoxyresorufin O-dealkylase assay using a rat hepatocyte spheroid microarray. Journal of bioscience and bioengineering. 2010 Apr;109(4):395-9

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PMID: 20226384

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