Details

Autor(en) / Beteiligte

• Hoffmann, Stefan A.
• Müller-Vieira, Ursula
• Biemel, Klaus
• Knobeloch, Daniel
• Heydel, Sandra
• Lübberstedt, Marc
• Nüssler, Andreas K.
• Andersson, Tommy B.
• Gerlach, Jörg C.
• Zeilinger, Katrin

Titel

Analysis of drug metabolism activities in a miniaturized liver cell bioreactor for use in pharmacological studies

Ist Teil von

• Biotechnology and bioengineering, 2012-12, Vol.109 (12), p.3172-3181

Ort / Verlag

Hoboken: Wiley Subscription Services, Inc., A Wiley Company

Erscheinungsjahr

2012

Link zum Volltext

Quelle

MEDLINE

Beschreibungen/Notizen

• Based on a hollow fiber perfusion technology with internal oxygenation, a miniaturized bioreactor with a volume of 0.5 mL for in vitro studies was recently developed. Here, the suitability of this novel culture system for pharmacological studies was investigated, focusing on the model drug diclofenac. Primary human liver cells were cultivated in bioreactors and in conventional monolayer cultures in parallel over 10 days. From day 3 on, diclofenac was continuously applied at a therapeutic concentration (6.4 µM) for analysis of its metabolism. In addition, the activity and gene expression of the cytochrome P450 (CYP) isoforms CYP1A2, CYP2B6, CYP2C9, CYP2D6, and CYP3A4 were assessed. Diclofenac was metabolized in bioreactor cultures with an initial conversion rate of 230 ± 57 pmol/h/106 cells followed by a period of stable conversion of about 100 pmol/h/106 cells. All CYP activities tested were maintained until day 10 of bioreactor culture. The expression of corresponding mRNAs correlated well with the degree of preservation. Immunohistochemical characterization showed the formation of neo‐tissue with expression of CYP2C9 and CYP3A4 and the drug transporters breast cancer resistance protein (BCRP) and multidrug resistance protein 2 (MRP2) in the bioreactor. In contrast, monolayer cultures showed a rapid decline of diclofenac conversion and cells had largely lost activity and mRNA expression of the assessed CYP isoforms at the end of the culture period. In conclusion, diclofenac metabolism, CYP activities and gene expression levels were considerably more stable in bioreactor cultures, making the novel bioreactor a useful tool for pharmacological or toxicological investigations requiring a highly physiological in vitro representation of the liver. Biotechnol. Bioeng. 2012; 109: 3172–3181. © 2012 Wiley Periodicals, Inc. The suitability of a novel miniaturized hollow fiber bioreactor for pharmacological studies on primary human hepatocytes was investigated. Metabolism of diclofenac, activity and expression of pharmacologically relevant cytochrome P450 enzymes and general metabolism were considerably more stable over the culture period of 10 days compared to conventional monolayer culture. Morevover, bioreactor culture promoted formation of neo‐tissue from single cell suspensions with spatial expression patterns resembling native liver tissue.