Hepatocyte nuclear factor-4α promotes differentiation of intestinal epithelial cells in a coculture system

Abstract

Normal cellular models able to efficiently recapitulate intestinal epithelial cell differentiation in culture are not yet available. The aim of this work was to establish and genetically characterize a mesenchymal-epithelial coculture system to identify transcriptional regulators involved in this process. The deposition of rat intestinal epithelial cells on human intestinal mesenchymal cells led to the formation of clustered structures that expanded shortly after seeding. These structures were composed of polarized epithelial cells with brush borders and cell junction complexes. A rat GeneChip statistical analysis performed at different time points during this process identified hepatocyte nuclear factor-4α (HNF-4α) and hepatocyte nuclear factor-1α (HNF-1α) as being induced coincidently with the apparition of polarized epithelial structures. Stable introduction of HNF-4α in undifferentiated epithelial cells alone led to the rapid induction of HNF-1α and several intestinal-specific markers and metabolism-related genes for which mRNA was identified to be upregulated during epithelial differentiation. HNF-4α was capable to transactivate the calbindin 3 gene promoter, a process that was synergistically increased in the presence of HNF-1α. When HNF-4α-expressing cells were plated on mesenchymal cells, an epithelial monolayer formed rapidly with the apparition of dome structures that are characteristics of vectorial ion transport. Forced expression of HNF-1α alone did not result in dome structures formation. In sum, this novel coculture system functionally identified for the first time HNF-4α as an important modulator of intestinal epithelial differentiation and offers an innovative opportunity to investigate molecular mechanisms involved in this process.

Modification in Oxidative Stress, Inflammation, and Lipoprotein Assembly in Response to Hepatocyte Nuclear Factor 4α Knockdown in Intestinal Epithelial Cells

“…Surprisingly, limited work has been devoted to the gut even though HNF4␣ is at the crossroads between intestinal morphogenesis and function. As reported previously, HNF4␣ plays a role in colon development (17), intestinal epithelial cell differentiation, and phenotype expression (18,19). Interestingly, not only did HNF4␣ regulate the expression of many genes such as apolipoproteins (apo) (20 -25), but it also has been found associated with susceptibility to abnormal intestinal permeability and inflammation (26).…”

Hepatocyte Nuclear Factor-4α Promotes Gut Neoplasia in Mice and Protects against the Production of Reactive Oxygen Species

“…Hnf4α plays major roles in epithelial cell polarization by targeting specific genes encoding cell junction molecules in various tissues, including the intestine (10,11,15,17,32). Moreover, Hnf4α acts as a morphogen with its capacity to trigger microvillus formation in nonepithelial cell lines (12,13).…”

“…The Wnt/β-catenin pathway is mostly active in crypt stem cell/progenitors, and Hnf4α is induced during intestinal epithelial cell differentiation in culture (11). To explore the apparent opposite relationship between Hnf4α expression and sustained β-catenin activity, we monitored the expression level of Hnf4α along the intestinal crypt-villus axis as well as in intestinal Apc Min polyps well characterized by harboring sustained activation of the Wnt/β-catenin pathway (28).…”

“…The specific role of Hnf4α in controlling intestinal epithelial functions has emerged recently. Hnf4α can polarize intestinal epithelial cells in culture (11) and is potent for the epithelialization of different cell types (10,12,13). Maturation of colonic epithelial cells was shown to be dependent on Hnf4α when conditionally removed during early embryonic colonic development (14).…”

Genome-wide Analysis of CDX2 Binding in Intestinal Epithelial Cells (Caco-2)

“…Previous work by Lussier et al (44) has shown that CDX2 is essential in initiating differentiation and that this is coordinated with the activation of HNF4␣ and HNF1␣ expression, which in turn requires CDX2 action. Furthermore, we have previously shown that HFN4␣ regulates the expression of CDX2; thus, a loop that is necessary for the initiation and completion of differentiation of intestinal cells appears to exist (12).…”