Abstract
An hepatocyte culture system was developed for potential use in toxicological studies in vitro. Rat hepatocytes were isolated by two-step collagenase perfusion and cultured on Vitrogen-coated Permanox(TM) dishes in a modified Chee's medium containing 1 μM dexamethasone and 1% dimethylsulfoxide. The cells remained highly viable for at least 10 d as determined by lactate dehydrogenase release and total protein levels. Albumin secretion into the medium, as a measure of differentiated function, was maintained at elevated levels over the course of 10 d in culture. A number of CYP activities were determined by the analysis of testosterone metabolism in freeze-thawed cells, diazepam metabolism in live cells, and specific assays for CYP 1A1/2, 2B1/2, 2E1, and 3A. Results of these assays indicated that a wide range of CYP isozymes were maintained, some activities were enhanced under the conditions of culture and some activities were inducible. Activities of the phase 11 enzymes, glutathione S-transferase and UDP- glucuronosyltransferase, and glutathione levels were also maintained in the cultured hepatocytes for at least 6 d. These results strongly support the use of this hepatocyte culture system for in vitro toxicological studies.
Original language | English (US) |
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Pages (from-to) | 211-220 |
Number of pages | 10 |
Journal | In Vitro Cellular and Developmental Biology - Animal |
Volume | 32 |
Issue number | 4 |
DOIs | |
State | Published - Apr 1996 |
Keywords
- cytochrome P450
- in vitro toxicology
- metabolism
- rat hepatocytes
ASJC Scopus subject areas
- Developmental Biology
- Cell Biology