Comparison of mRNA expression profiles of drug-metabolizing enzymes and transporters in fresh and cryopreserved cynomolgus monkey hepatocytes
| Autor: | Akiko Koeda, Misaki Yamashita, Tamihide Matsunaga, Shota Mizuno, Anna Nakanishi, Katsunori Nakamura, Yoko Sakai, Takahiro Iwao |
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| Rok vydání: | 2018 |
| Předmět: |
Male
Cell Survival Mrna expression Pharmaceutical Science Real-Time Polymerase Chain Reaction 030226 pharmacology & pharmacy Cryopreservation Andrology 03 medical and health sciences Mice 0302 clinical medicine Animals Pharmacology (medical) Viability assay Aspartate Aminotransferases RNA Messenger Gene Cells Cultured 030304 developmental biology Pharmacology chemistry.chemical_classification 0303 health sciences L-Lactate Dehydrogenase Chemistry Gene Expression Profiling Spheroid Transporter Alanine Transaminase 3T3 Cells gamma-Glutamyltransferase Macaca fascicularis Enzyme Hepatocytes Female Liver function |
| Zdroj: | Drug metabolism and pharmacokinetics. 34(4) |
| ISSN: | 1880-0920 |
| Popis: | In this study, freshly isolated and cryopreserved cynomolgus monkey hepatocytes were seeded on Cell-able® plates with feeder cells to form spheroids and were cultured for 28 days. As a control, hepatocytes were also cultured with or without feeder cells on collagen-coated plates. We verified the mRNA expression levels of drug-metabolizing enzyme-related genes and the leakage of enzymes (AST, ALT, LDH, and γ-GTP) as indicators of cell survival. As a result, the patterns of target mRNA expression in fresh and cryopreserved hepatocytes were very similar during the culture period between culture methods. mRNA expression levels were highly maintained at day 28 using the 3D spheroid and co-culture methods, demonstrating that these methods are useful for maintenance of liver function. Leakage of AST and ALT was higher at day 3 but decreased at day 14. LDH was not detected, suggesting that the cell viability was also maintained during the culture period. Furthermore, the functional differences between fresh and cryopreserved hepatocytes were not clearly detected. The co-culture method was useful for long-term culture not requiring 3D structure, and the 3D spheroid culture method was effective as well. With these techniques, cynomolgus monkey hepatocytes are expected to exhibit smaller individual differences and high reproducibility. |
| Databáze: | OpenAIRE |
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