Development of Scaffolds with Adjusted Stiffness for Mimicking Disease-Related Alterations of Liver Rigidity.

Drug-induced liver toxicity is one of the most common reasons for the failure of drugs in clinical trials and frequent withdrawal from the market. Reasons for such failures include the low predictive power of in vivo studies, that is mainly caused by metabolic differences between humans and animals, and intraspecific variances. In addition to factors such as age and genetic background, changes in drug metabolism can also be caused by disease-related changes in the liver. Such metabolic changes have also been observed in clinical settings, for example, in association with a change in liver stiffness, a major characteristic of an altered fibrotic liver. For mimicking these changes in an in vitro model, this study aimed to develop scaffolds that represent the rigidity of healthy and fibrotic liver tissue. We observed that liver cells plated on scaffolds representing the stiffness of healthy livers showed a higher metabolic activity compared to cells plated on stiffer scaffolds. Additionally, we detected a positive effect of a scaffold pre-coated with fetal calf serum (FCS)-containing media. This pre-incubation resulted in increased cell adherence during cell seeding onto the scaffolds. In summary, we developed a scaffold-based 3D model that mimics liver stiffness-dependent changes in drug metabolism that may more easily predict drug interaction in diseased livers.

Cell-Type-Specific Quantification of a Scaffold-Based 3D Liver Co-Culture.

In order to increase the metabolic activity of human hepatocytes and liver cancer cell lines, many approaches have been reported in recent years. The metabolic activity could be increased mainly by cultivating the cells in 3D systems or co-cultures (with other cell lines). However, if the system becomes more complex, it gets more difficult to quantify the number of cells (e.g., on a 3D matrix). Until now, it has been impossible to quantify different cell types individually in 3D co-culture systems. Therefore, we developed a PCR-based method that allows the quantification of HepG2 cells and 3T3-J2 cells separately in a 3D scaffold culture. Moreover, our results show that this method allows better comparability between 2D and 3D cultures in comparison to the often-used approaches based on metabolic activity measurements, such as the conversion of resazurin.