Mr. Shanks et al., AN IMPROVED POLARIZED RAT HEPATOMA HYBRID CELL-LINE - GENERATION AND COMPARISON WITH ITS HEPATOMA RELATIVES AND HEPATOCYTES IN-VIVO, Journal of Cell Science, 107, 1994, pp. 813-825
Studies of hepatocyte polarity, an important property of liver epithel
ial cells, have been hampered by the lack of valid in vitro models. We
report here that a new polarized hepatoma-derived hybrid cell line, c
alled WIF-B, has improved characteristics to those of its parent, WIF1
2-1. This latter line originated from the fusion of non-polarized rat
hepatoma Fao cells with human fibroblasts (WI-38) and selection for a
polarized phenotype. We generated the WIF-B line by growing WIF12-1 ce
lls as unattached aggregates for three weeks and selecting for survivo
rs. Karyotype analysis showed a broad chromosome pattern in the initia
l WIF-B population, but this pattern stabilized after a few passages.
The growth and phenotypic properties of these cells were quite differe
nt from those of their polarized WIF12-1 parent. WIF-B cells attained
a 4-fold higher maximal density in monolayer culture, survived at this
density for >5 days rather than 1 day, and exhibited two to three tim
es more apical structures during this period (80 to 95%). We compared
several parameters of liver differentiation in the WIF-B cells with th
ose of a related hybrid clone, WIF12-E, which is extinguished for most
liver-specific functions, and,vith the common hepatoma parent, Fao. B
y immunoblot analysis, the levels of expression of eight plasma membra
ne proteins were higher in the WIF-B cells than in either of the other
two cell lines and ranged from 10 to 200% of those in vivo. Two plasm
a membrane proteins were not detected in WIF12-E cells. By immunofluor
escence, the apical membrane proteins in WIF-B displayed different cel
lular localizations than in either of the other two cell lines. In WIF
-B cells, apical proteins were confined to a plasma membrane region th
at we have identified as the apical domain by several criteria (Ihrke,
G., Neufeld, E. D., Meads, T., Shanks, M. R., Cassio, D., Laurent, M.
, Schroer, T.A., Pagano, R. E. and Hubbard, A. L. J. Cell Biol., 123,
1761-1765). The same molecules were distributed over the entire plasma
membrane of Fao and WIF12-E cells and also (for Fao cells) in intrace
llular punctate structures that did not colocalize with the majority o
f structures containing a secretory protein, albumin. Our results indi
cate that the WIF-B dells are more highly differentiated than any of t
heir ancestors (Fao or WIF12-1 cells) and thus, are promising candidat
es for in vitro studies of hepatocyte polarity.