Human hepatocyte cell lines proliferating as cohesive spheroid colonies in alginate markedly upregulate both synthetic and detoxificatory liver function

Background/Aims: Bio-artificial liver support systems for treatment of hepatic failure require maintained expression of hepatocyte function in vitro. We studied cultures of human hepatocyte cell-lines proliferating within alginate beads, investigating the hypothesis that 3-dimensional cohesive colonies of hepatocyte cell-lines would achieve polarity and cell-to-cell contact resulting in upregulation of function. Methods: HepG2 and HHY41 human cell lines in alginate beads were cultured for >20 days. Results: Proliferation was maintained for 20 days. Production of albumin, prothrombin, fibrinogen, alpha-1-acid glycoprotein and alpha-1-antitrypsin was maintained throughout, maximal at days 8–10, when upregulation was 300–1100% compared with monolayer cultures at similar cell number per unit volume. Detoxificatory functions: ethoxyresorufin deethylase activity, androstenedione metabolism, and urea synthesis from arginine was also increased several-fold. Function returned to pre-freezing levels within 18 h of thawing after cryopreservation of cells in alginate. Electron microscopy revealed spherical colonies of cells of cuboidal shape, with cell-to-cell contact via desmosomes and junctional complexes, abundant microvilli, and cytoplasmic appearances suggesting transcriptionally active hepatocytes. Conclusion: Hepatocyte cell-lines, proliferating in alginate express a range of liver-specific functions at levels approaching those found in vivo, relevant to their use in liver support systems.