The effect of fibroblast-secreted factor on primary heparocytes

Abstract

Primary hepatocytes are widely used in the studies of drug metabolism, toxicology, gene expression and the bioartificial liver. However, hepatocytes rapidly lose their specific function in vitro. Hepatocytes viability and liver-specific function can be maintained for several weeks in vitro when cultivated with other cell types. The potential mediators of this co-culture interaction are thought to be: receptors, gap junctions, cell-cell contacts, secreted extracellular matrix (ECM), exosomes and soluble growth factors. Even though there have been many efforts to investigate the hepatocyte co-culture mechanism, the precise mechanism remains unknown. It has been suggested that direct contact is required between hepatocytes and fibroblasts in co-culture. In addition, the use of conditioned medium has long been known for the inability to examine the role of soluble factor degradation or dilution and the potential for its effects on hepatocytes. In this study, we demonstrated that liver function can be modulated by treating with condensed fresh fibroblast-conditioned medium (F-CM) in varying concentrations and without direct contact. Contrary to previous study, our conclusion shows that the concentration of fibroblast secreted factors is a critical point, and that employing direct contact between hepatocytes and fibroblasts is not required. A certain threshold concentration of F-CM is required. In addition to this, other insoluble factors may exist such as extracellular vesicles (EVs) which are known to be one kind of communicator playing diverse roles in intercellular communication. To further study the effect of exosomes on hepatocytes, we treated exosomes, one type of EV, on hepatocytes by separating exosomes in the conditioned medium of fibroblasts. We confirmed that exosomes released by fibroblasts have no effect on hepatocytes. This mechanism can be useful for hepatic tissue engineering as a novel therapeutic approach to treat damaged or diseased liver tissue. We describe a new method of hepatocytes culture for the retention of viability and functions by treating hepatocytes with fresh condensed F-CM.