Cellular interactions promote tissue-specific function, biomatrix deposition and junctional communication of primary cultured hepatocytes

Abstract

Hepatocytes, prepared from normal adult rat liver, were seeded onto a collagen substratum and cultured alone or in the presence of rat liver endothelial cells. When hepatocytes were cultured alone in a hormonally defined serum-free medium, decreased albumin production and rapid morphological deterioration of bile canaliculi structures and gap junctions occurred within 4 to 5 days. In contrast, hepatocytes cocultured with liver mesenchymal cells remained morphologically intact and biochemically functional for at least 4 weeks. They reorganized into small islands, continued to secrete high levels of albumin, did not express alpha-fetoprotein (a fetal marker), and remained strongly dye coupled. All of the hepatocytes synthesized albumin and retained their gap junctional channels. No junctional communication was observed between hepatocytes and endothelial cells. Long fibers containing fibronectin, Type I collagen and laminin distributed over the hepatocytes were induced in coculture but never appeared in hepatocytes cultured alone. Moreover, supplementation of the hormonally defined medium with phenobarbital and dimethyl sulfoxide, both of which improve the life span and functional activities of cultured hepatocytes, failed to induce reticulin fiber formation in pure culture of hepatocytes. The modulation of albumin secretion, biomatrix deposition and junctional communication observed in hepatocytes cultured with sinusoidal liver cells was also obtained when hepatocytes were in association with various epithelial or mesenchymal cells [rat liver epithelial cells (T51B), mouse embryonic fibroblasts (NIH 3T3), human or rat dermal fibroblasts and bovine aorta endothelial cells (AG 4762)].

Liver cell models in in vitro toxicology.

“…The enhanced survival and function of hepatocytes cocultured with rat liver epithelial cells have been confirmed by many investigators (134,(138)(139)(140)(141)(142)(143); the induction of CYP2B1 and CYP2B2 by phenobarbital has also been demonstrated (137,144). Moreover, other cells have been effective, including both liver endothelial cells and nonhepatic cells (145)(146)(147). Studies from our laboratory (148,149) have shown that the presence of an integral plasma membrane glycoprotein is a prerequisite to obtain longterm survival of differentiated hepatocytes in coculture.…”

Liver endothelial cells promote LDL-R expression and the uptake of HCV-like particles in primary rat and human hepatocytes

“…8. Hepatocytes cultured in a collagen double-gel have been shown to exhibit native cell-cell contacts such as E-cadherin and bile canaliculi, 34 but do not express sinusoidal receptors such as EGF-R 34 and LDL-R. Hepatocytes cocultured with 3T3 fibroblasts exhibit hepatic cell-cell contacts such as connexin-32 44 and bile canaliculi, 24 but also do not express the EGF-R and LDL-R. On the other hand, hepatocytes cocultured with LSEC show both traditional polarity markers, 45,46 and express a high level of sinusoidal receptors (EGF-R, LDL-R) at the interface between the hepatocytes and the LSEC. At least part of the interaction between hepatocytes and LSECs has been shown to be mediated by growth factors.…”

Long-Term Coculture Strategies for Primary Hepatocytes and Liver Sinusoidal Endothelial Cells

“…[19][20][21] In the liver sinusoid, LSECs and hepatocytes are arranged in layers with the intervening space occupied by the extracellular matrix (ECM) of the perisinusoidal space (space of Disse); collagen I is one of the major components in this matrix. [22][23][24][25] While hepatocytes are primarily responsible for various secretion and metabolic functions of the liver; LSECs line the sinusoids, isolating hepatocytes from the sinusoid flow and are first to be exposed to various toxic and benign factors circulating through the hepatic sinusoids.…”