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Differential maintenance of cytochrome P450 enzymes in cultured precision-cut human liver slices

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Author: Renwick, A.B. · Watts, P.S. · Edwards, R.J. · Barton, P.T. · Guyonnet, I. · Price, R.J. · Tredger, J.M. · Pelkonen, O. · Boobis, A.R. · Lake, B.G.
Type:article
Date:2000
Institution: Centraal Instituut voor Voedingsonderzoek TNO TNO Voeding
Source:Drug Metabolism and Disposition, 10, 28, 1202-1209
Identifier: 280408
Keywords: Nutrition · Apoprotein · Cytochrome P450 · Cytochrome p450 2a6 · Cytochrome P450 2B6 · Cytochrome P450 2C19 · Cytochrome p450 2c8 · Cytochrome P450 2C9 · Cytochrome P450 3A4 · Cytochrome P450 3A5 · Debrisoquine 4 hydroxylase · Dimethylnitrosamine demethylase · Ethoxyresorufin · O Deethylase · Peptide antibody · Phenacetin o deethylase · Tolbutamide methylhydroxylase · Unclassified drug · Enzyme activity · Enzyme metabolism · Human tissue · Liver cell culture · Liver slice · Protein stability · School child · Xenobiotic metabolism · Adolescent · Adult · Aged · Child · Child, Preschool · Culture Media · Culture Techniques · Cytochrome P-450 Enzyme System · Enzyme Stability · Female · Humans · Immunoblotting · Isoenzymes · Liver · Male · Microsomes, Liver · Middle Aged

Abstract

The maintenance of the major hepatic cytochrome P450 (CYP) enzymes has been studied in precision-cut human liver slices cultured for up to 72 h in supplemented RPMI 1640 medium. The relative apoprotein levels of 11 CYP enzymes were determined using a panel of antipeptide antibodes. In addition, 7-ethoxyresorufin O-deethylase, tolbutamide methylhydroxylase, debrisoquine 4-hydroxylase, and testosterone 6β-hydroxylase activities were determined as enzymatic markers for CYP1A2, CYP2C9, CYP2D6, and CYP3A4, respectively. There was a large variation in the rate of decline of different CYP levels with time in culture. Based on the rate of decrease, CYP enzymes could be separated into two groups, with CYP2C9, CYP2D6, CYP3A4, and CYP4A11 being relatively stable (half-lives between 70 and 104 h), compared with CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C19, CYP2E1, and CYP3A5, which were relatively unstable (half-lives between 23 and 36 h). Enzyme activities decreased at rates similar to those of their corresponding apoproteins. There was also a large difference in the stability of individual CYP enzymes from different liver donors, particularly for the most rapidly declining CYP enzymes. Similar losses of CYP enzymes were found when human river slices were cultured in supplemented Williams' medium E for 72 h, except that CYP2E1 apoprotein levels were better maintained. Because of the variable decreases of CYP enzymes, xenobiotic metabolism studies are best performed with freshly cut rather than cultured human liver slices. Chemicals/CAS: Culture Media; Cytochrome P-450 Enzyme System, 9035-51-2; Isoenzymes