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The extent of phase I and phase II reactions is affected by the choice of enzyme used to prepare rat hepatocytes

Sinclair, J.A. and Henderson, C.J. and Martin, I.K. and Grant, M.H. and Tettey, J.N.A. (2009) The extent of phase I and phase II reactions is affected by the choice of enzyme used to prepare rat hepatocytes. Chemico-Biological Interactions, 179 (2-3). pp. 256-262. ISSN 0009-2797

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Abstract

The preparation of hepatocytes using the two-stage perfusion technique usually involves the use of collagenase (CII) alone or in combination with dispase (C/D) or trypsin inhibitor (CA/TI) as digestion enzymes. The effect of CII, C/D and CA/TI on cell viability, yield, cytochrome P450 mediated oxidation of testosterone, glucuronidation and sulfation of 7-hydroxycoumarin, glutathione content, glutathione-S-transferase activity and glutathione-conjugation capacity of hepatocytes has been assessed. Cytochrome P450 mediated oxidation of testosterone was significantly (p < 0.05) decreased with CII isolated hepatocytes (81.7 +/- 3.3 nmol/10(6) cells, mean +/- S.E.M., n = 3), compared with those isolated using CA/TI (96.6 +/- 1.9 nmol/10(6) cells) or C/D (95.1 +/- 2.1 nmol/10(6) cells). In contrast, glutathione conjugation of the non-specific substrate 1-chloro-2,4-dinitrobenzene was significantly (p < 0.05) increased with CII isolated hepatocytes (56.9 +/- 5.9 nmol/10(6) cells, mean +/- S.E.M., n = 3), compared with those isolated using CA/TI (36.0 +/- 3.7 nmol/10(6) cells) or C/D (31.6 +/- 3.7 nmol/10(6) cells). These findings have significant implications for the interpretation of metabolism data derived from hepatocytes in suspension, particularly in terms of glutathione conjugation of potentially toxic reactive intermediates of xenobiotic metabolism. Indeed, data presented show that the presence of trypsin inhibitor in the preparation of isolated rat hepatocytes significantly affects the formation of glutathione conjugates of reactive intermediate products of troglitazone metabolism.

Item type: Article
ID code: 18717
Notes: PMID: 19330883
Keywords: hepatocytes, collagenase, dispase, trypsin inhibitor, glutathione, reactive intermediates, Bioengineering, Engineering (General). Civil engineering (General), Toxicology
Subjects: Technology > Engineering (General). Civil engineering (General) > Bioengineering
Technology > Engineering (General). Civil engineering (General)
Department: Faculty of Engineering > Bioengineering
Faculty of Science > Strathclyde Institute of Pharmacy and Biomedical Sciences
Related URLs:
    Depositing user: Ms Ashley Urie
    Date Deposited: 13 Apr 2010 14:50
    Last modified: 05 Sep 2014 03:04
    URI: http://strathprints.strath.ac.uk/id/eprint/18717

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