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


CONCLUSIONS
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.
Khalil
M
M
Division of Medicine, Imperial College School of Medicine, Hammersmith Hospital, London, UK.
Shariat-Panahi
A
A
Tootle
R
R
Ryder
T
T
McCloskey
P
P
Roberts
E
E
Hodgson
H
H
Selden
C
C
D000464:Alginates; D000735:Androstenedione / Q000378:metabolism; D002455:Cell Division; D002460:Cell Line; D015925:Cryopreservation; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D020723:Glucuronic Acid; D022781:Hepatocytes / Q000502:physiology*; D006603:Hexuronic Acids; D006801:Humans; D008658:Inactivation, Metabolic; D008099:Liver / Q000378:metabolism*; D019164:Liver, Artificial; D018874:Spheroids, Cellular; D015854:Up-Regulation; D014508:Urea / Q000378:metabolism
D000464:Alginates; D006603:Hexuronic Acids; D000735:Androstenedione; D020723:Glucuronic Acid; D014508:Urea; D003577:Cytochrome P-450 Enzyme System