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

• Published in: Journal of hepatology Vol. 34; no. 1; pp. 68 - 77
• Main Authors: Khalil, Marianne, Shariat-Panahi, Ali, Tootle, Rosemary, Ryder, Tim, McCloskey, Paschal, Roberts, Eve, Hodgson, Humphrey, Selden, Clare
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier B.V 2001; Elsevier
• Subjects: 3-Dimensional; Alginate; Alginates; Androstenedione - metabolism; Animal cells; Bio-artificial liver; Biological and medical sciences; Biotechnology; Cell Division; Cell Line; Cryopreservation; Cytochrome P-450 Enzyme System - metabolism; Detoxification; Establishment of new cell lines, improvement of cultural methods, mass cultures; Eukaryotic cell cultures; Fundamental and applied biological sciences. Psychology; Glucuronic Acid; Hepatocytes - physiology; Hexuronic Acids; Human hepatocyte; Humans; Inactivation, Metabolic; Liver - metabolism; Liver specific protein; Liver, Artificial; Methods. Procedures. Technologies; Spheroids, Cellular; Up-Regulation; Urea - metabolism; Alginate; Human hepatocyte; Bio-artificial liver; 3-Dimensional; Liver specific protein; Detoxification; Human; Biotechnology; Cell culture; Artificial organ; Hepatic disease; Alginates; Experimental study; In vitro; Fulminant; Medium enrichment; Liver function; Liver failure; Hepatocyte; Reinforcement; Digestive diseases
• ISSN: 0168-8278; 1600-0641
• DOI: 10.1016/S0168-8278(00)00080-5

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.