Microfluidic 3D HepG2 cell culture: Reproducing hepatic tumor gene and protein expression in in vitro scaffolds

Abstract

As for 3D cell culture in general, it is also true for tumor cells that 3D conditions create a spatial microenvironment that is stimulated by concentration gradients of nutrients and gases. Cellular vitality and metabolic activity were examined in 2D and scaffold‐based 3D cultures of HepG2 cells. We compared gene and protein expression levels of tumor‐ and hepato‐specific markers such as alpha‐fetoprotein, albumin, integrin beta 1, insulin‐like growth factor , vascular endothelial growth factor A, and matrix metalloproteinase 11. Although similar biological functions were influenced under 3D static and perfused conditions, their gene expression levels differed. Immunofluorescence stainings detected increased integrin beta 1 expression while alpha‐fetoprotein was downregulated both in static and perfused 3D cultures of HepG2 cells. We hypothesized that improved mass flow in the bioreactors enables the cells to retain their nutrient supply, which is indicated by moderate or even downregulated expression of genes related to angiogenesis and apoptosis. Potential hypoxic stress in static 3D cultures of HepG2 cells resulted in upregulated vascular endothelial growth factor A expression and enhanced caspase 3 activity. Tuning the mass flow in 3D tumor cultures may lead to promising tools to investigate the metabolism and differentiation state of tumor tissues in vitro.