Satisfactory performance of hepatocytes in three-dimensional culture on innovative biopolymeric scaffolds

The introduction in recent years of innovative biomaterials as scaffolds to replace physiological matrix components has led to significant advances in culture techniques, in terms of cell survival, quantitative expansion, maintenance of differentiated phenotype and specific functions of cells. ese bio-artificial tissues can be used as functional substitutes for damaged organs, as models to study complex biological processes, or for tissue-specific toxicology tests. e aim of our research was to identify the most suitable biomaterial for technological applications with hepatocytes, e.g. tissue therapy, cellular transplantation, metabolic analysis of new drugs or screening for potentially toxic substances. Since the possibility to improve the performance of these systems depends strongly on the methods used to create the scaffolds, here we analyzed porous matrices made of gelatin or blends of gelatin and glycosaminoglycans, obtained with different methods for the culture of the C3A cell line, considered a good model of human hepatocytes. Scaffolds were obtained using either a concentrated emulsion-templating technique known as high internal phase emulsion (HIPE) or a gas foaming technique; the latter method uses an inert gas instead of the internal liquid phase toluene, avoiding the use of organic solvent and allowing the creation of scaffolds with larger pores and interconnections. By analyzing cell adhesion, viability, ultrastructural morphology, production of albumin and urea and induction of cytochrome P450-3A4 as the main metabolic markers of hepatocyte functions, it was possible to compare the performances of matrices prepared with the different methods. Taken together our results suggest that the morphology of the scaffolds (surface porosity, void/ interconnection size) is less crucial than the utilization of potentially harmful molecules during their creation.