Differential transcriptional characteristics of small and large biliary epithelial cells derived from small and large bile ducts

Abstract

Biliary epithelial cells (BEC) are morphologically and functionally heterogeneous. To investigate the molecular mechanism for their diversities, we test the hypothesis that large and small BEC have disparity in their target gene response to their transcriptional regulator, the biliary cell-enriched hepatocyte nuclear factor HNF6. The expression of the major HNF (HNF6, OC2, HNF1b, HNF1a, HNF4a, C/EBPb, and Foxa2) and representative biliary transport target genes that are HNF dependent were compared between SV40-transformed BEC derived from large (SV40LG) and small (SV40SM) ducts, before and after treatment with recombinant adenoviral vectors expressing HNF6 (AdHNF6) or control LacZ cDNA (AdLacZ). Large and small BEC were isolated from mouse liver treated with growth hormone, a known transcriptional activator of HNF6, and the effects on selected target genes were examined. Constitutive Foxa2, HNF1a, and HNF4a gene expression were 2.3-, 12.4-, and 2.6-fold, respectively, higher in SV40SM cells. This was associated with 2.7-and 4-fold higher baseline expression of HNF1a-and HNF4a-regulated ntcp and oatp1 genes, respectively. Following AdHNF6 infection, HNF6 gene expression was 1.4-fold higher (P ϭ 0.02) in AdHNF6 SV40SM relative to AdHNF6 SV40LG cells, with a corresponding higher Foxa2 (4-fold), HNF1a (15-fold), and HNF4a (6-fold) gene expression in AdHNF6-SV40SM over AdHNF6-SV40LG. The net effects were upregulation of HNF6 target gene glucokinase and of Foxa2, HNF1a, and HNF4a target genes oatp1, ntcp, and mrp2 over AdLacZ control in both cells, but with higher levels in AdH6-SV40SM over AdH6-SV40LG of glucokinase, oatp1, ntcp, and mrp2 (by 1.8-, 3.4-, 2.4-, and 2.5-fold, respectively). In vivo, growth hormone-mediated increase in HNF6 expression was associated with similar higher upregulation of glucokinase and mrp2 in cholangiocytes from small vs. large BEC. Small and large BEC have a distinct profile of hepatocyte transcription factor and cognate target gene expression, as well as differential strength of response to transcriptional regulation, thus providing a potential molecular basis for their divergent function. heterogeneity; bile transport; hepatocyte nuclear factors; hepatocyte nuclear factor 6

Growth Hormone Mediates Its Protective Effect in Hepatic Apoptosis through Hnf6

“…Primer sequences are listed in Table 1. Hnf6 , Hnf1b , Hnf4a , Oc2 , Foxa2 , C/epb primer sequences have been reported previously [6,15,17]. Relative gene expression was calculated using the mathematical delta delta method by PE Applied Biosystems.…”

Pathobiology of biliary epithelia

“…Large cholangiocytes are thought to produce the majority of bile, while the rest (10–30%) is produced by the hepatocytes in rats and humans respectively [35,36]. However, it has been demonstrated that both small and large cholangiocytes express bile acid transporters (e.g., sodium/taurocholate co -transporting polypeptide [ ntcp ], organic anion-transporting polypeptide [ oatp1 ] and multidrug resistance protein 2 [ mrp2 ]) and aquaporins, suggesting that both may play a substantive role in bile acid transport and production [37,38]. Regardless, it is clear that the apical and basolateral membranes of large cholangiocytes possess specialized functions via the presence of different ion carriers and channels (Fig.…”

Physiology of Cholangiocytes

“…Recent observations, however, suggest that the functional heterogeneity of large and small cholangiocytes is more complex, and that both large as well as small cholangiocytes may contribute to bile modification (106,283). …”