The past years have witnessed an increasing number of reports relative to homeobox genes in endoderm-derived tissues. In this review, we focus on the caudal-related Cdx-1 and Cdx-2 homeobox genes to give an overview of the in vivo, in vitro, and ex vivo approaches that emphasize their primary role in intestinal development and in the control of intestinal cell proliferation, differentiation, and identity. The participation of these genes in colon tumorigenesis and their identification as important actors of the oncogenic process are also discussed.

Cell deformation occurs in many critical biological processes, including cell extravasation during immune response and cancer metastasis. These cells deform the nucleus, its largest and stiffest organelle, while passing through narrow constrictions in vivo and the underlying mechanisms still remain elusive. It is unclear which biochemical actors are responsible and whether the nucleus is pushed or pulled (or both) during deformation. Herein we use an easily-tunable poly-L-lactic acid micropillar topography, mimicking in vivo constrictions to determine the mechanisms responsible for nucleus deformation. Using biochemical tools, we determine that actomyosin contractility, vimentin and nucleo-cytoskeletal connections play essential roles in nuclear deformation, but not A-type lamins. We chemically tune the adhesiveness of the micropillars to show that pulling forces are predominantly responsible for the deformation of the nucleus. We confirm these results using an in silico cell model and propose a comprehensive mechanism for cellular and nuclear deformation during confinement. These results indicate that microstructured biomaterials are extremely versatile tools to understand how forces are exerted in biological systems and can be useful to dissect and mimic complex in vivo behaviour.

The effects of long-term starvation on the activities of sucrase, lactase, and aminopeptidase, and on their respective mRNA were determined in the small intestine of thyroidectomized and sham-operated adult rats. Thyroidectomy reduced the protein loss at the level of the intestinal brush border membranes during starvation. Prolonged fasting caused a significant decrease in sucrase activity, but thyroidectomy partly prevented this effect. However, the amount of the corresponding mRNA dropped during long term starvation without incidence of thyroidectomy. Lactase activity in the brush border membranes was increased by starvation, and thyroidectomy caused a further elevation of the enzyme activity. Simultaneously, lactase mRNA content rose only slightly compared to the enzyme activity. Aminopeptidase activity and mRNA content decreased during starvation and thyroidectomy did not prevent this process. These results indicate that intestinal hydrolases respond non-coordinately to long-term food deprivation. In addition, the thyroid status of the animals has a direct influence on the adaptation of several brush border hydrolases to starvation. This suggests that the drop in plasma thyroid hormones during fasting allows a better maintenance of protein content and of hydrolase activities in the brush border membranes of the small intestine. These adaptive processes seemed to be partly controlled at a post-transcriptional level.

We have determined the sequence of a 2784 bp rat genomic fragment originating from the 5' region of the gene coding for intestinal lactase-phlorizin hydrolase. The fragment overlaps the gene exon 1, part of the intron 1 and the 5'-upstream segment including a TATA-like box. Over 155 bp, the upstream segment shows 72% similarity with the corresponding sequence in human. Far upstream, the rat sequence exhibits a Calcium Responsive Element and putative binding sites for AP2, C/EBP, and CTF/NF. The intron contains a T-rich sequence that may cause DNA helix distortion.

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In segmented organs, homeobox genes are involved in axial patterning and cell identity. Much less is known about their role in non-segmented endoderm derivatives such as the digestive epithelium. Using a xenograft model of fetal intestinal anlagen implanted under the skin of nude mice, we have investigated whether the expression of five homeobox genes (HoxA-4, HoxA-9, HoxC-8, Cdx-1 and Cdx-2) is modified when intestinal epithelium undergoes normal development or displays heterodifferentiation in association with heterotopic mesenchyme. In homotypic associations of fetal endoderm and mesenchyme that recapitulate normal development, the overall pattern of homeobox gene expression was maintained: HoxA-9 and HoxC-8 were the highest in the colon and ileum, respectively, and HoxA-4 was expressed all along the intestine; Cdx-1 and Cdx-2 exhibited an increasing gradient of expression from small intestine to colon. Yet, grafting per se caused a faint upregulation of HoxA-9 and HoxC-8 in small intestinal regions in which these genes are not normally expressed, while the endoderm-mesenchyme dissociation-association step provoked a decay of Cdx-1 in the colon. In heterotopic associations of colonic endoderm with small intestinal mesenchyme, the colonic epithelium exhibited heterodifferentiation to a small intestinal-like phenotype. In this case, we observed a decay of HoxA-9 expression and an upregulation of HoxC-8. Additionally, heterodifferentiation of the colonic epithelium was accompanied by a downregulation of Cdx-1 and Cdx-2 to a level similar to that found in the normal small intestine. To demonstrate that mesenchyme-derived cells can influence Cdx-1 and Cdx-2 expression in the bowel epithelium, fetal jejunal endoderm was associated with intestinal fibroblastic cell lines that either support small intestinal-like or colonic-like morphogenesis. A lower expression of both homeobox genes was shown in grafts presenting the small intestinal phenotype than in those showing glandular colonic-like differentiation. Taken together, these results suggest that homeobox genes participate in the control of the positional information and/or cell differentiation in the intestinal epithelium. They also indicate that the level of Cdx-1 and Cdx-2 homeobox gene expression is influenced by epithelial-mesenchymal cell interactions in the intestinal mucosa.

In previous experiments we showed that intestinal development was dependent upon epithelial-mesenchymal cell interactions. The aim of this study was to investigate the possible role of retinoic acid (RA), a morphogenetic and differentiating agent, on the gut epithelial-mesenchymal unit. For this purpose we first analyzed the effects of a physiological dose of RA on 14-day fetal rat intestine using short-term organ culture experiments, or long-term grafts under the skin of nude mice. In these conditions, RA accelerated villus outgrowth and epithelial cell differentiation as assessed by the onset of lactase expression, and it also stimulated muscle and crypt formation. In order to analyze potential effects of RA mediated by mesenchymal cells, we isolated and characterized gut mucosa mesenchyme-derived cell cultures (mesenchyme-derived intestinal cell lines, MIC). These cells were shown to express mRNAs for retinoid binding proteins similar to those expressed in situ in the intestinal mesenchyme. MIC cells co-cultured with 14-day intestinal endoderms promoted endodermal cell adhesion and growth, and the addition of exogeneous RA enhanced epithelial cell polarization and differentiation assessed by cytokeratin and lactase immunostaining. Such a differentiating effect of RA was not observed on endodermal cells when cultured without a mesenchymal feeder layer or maintained in conditioned medium from RA-treated MIC cells. In the co-cultures, immunostaining of laminin and collagen IV with polyclonal antibodies, as well as alpha1 and beta1 laminin chains mRNAs (analyzed by RT-PCR) increased concurrently with the RA-enhanced differentiation of epithelial cells. It is worth noting that this stimulation by RA was also obvious on the mesenchymal cells cultured alone. These results show that RA plays a role in intestinal morphogenesis and differentiation. In addition, they indicate that RA acts on the mesenchymal cell phenotype and suggest that RA may modify the mesenchymal-epithelial cell interactions during intestinal development.

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