生物工程与生物医学杂志

Organotypic Cultures of Hepg2 Cells for In Vitro Toxicity Studies

Abstract

Daniel Mueller, Anika Koetemann and Fozia Noor

Tissue engineering of human liver cells in a three dimensional cell culture system could improve pharmacological studies in terms of drug metabolism, drug toxicity or adverse drug effects by mimicking the in vivo situation. In this study, we produced 3D organotypic cultures of HepG2 cells using the hanging drop method. 250 – 8000 seeded cells formed organotypic cultures within 2–3 days which increased in size during the first week. Viability and metabolic parameters (glucose, lactate) were analyzed during almost three weeks of cultivation. Liver specific albumin production was higher in the organotypic cultures as compared to both monolayer and collagen-sandwich cultures. Amino acid quantification revealed high production of glutamate as well as uptake of glutamine, alanine and branched-chain amino acids. CYP1A induction capacity was significantly improved by organotypic cultivation. The acute toxicity (24 h) of tamoxifen, an anti-cancer drug, was lower in the 3D cultures as compared to monolayer and collagen-sandwich cultures. This could be explained by a higher drug efflux through membrane transporter (MRP-2). We conclude that the engineered HepG2 cultures could be used for the investigation of CYP450 induction, anti-cancer drug effects and for the study of chemotherapy resistance. Applied to other cell types such as the human primary cells these 3D organotypic cultures may have potential in long term toxicity screening of compounds.