Nikè V. J. A. Büller

Blimp1 regulates the transition of neonatal to adult intestinal epithelium.

In many mammalian species, the intestinal epithelium undergoes major changes that allow a dietary transition from mothers milk to the adult diet at the end of the suckling period. These complex developmental changes are the result of a genetic programme intrinsic to the gut tube, but its regulators have not been identified. Here we show that transcriptional repressor B lymphocyte-induced maturation protein 1 (Blimp1) is highly expressed in the developing and postnatal intestinal epithelium until the suckling to weaning transition. Intestine-specific deletion of Blimp1 results in growth retardation and excessive neonatal mortality. Mutant mice lack all of the typical epithelial features of the suckling period and are born with features of an adult-like intestine. We conclude that the suckling to weaning transition is regulated by a single transcriptional repressor that delays epithelial maturation.

Loss of Indian hedgehog activates multiple aspects of a wound healing response in the mouse intestine

BACKGROUND & AIMS Indian Hedgehog (Ihh) is expressed by the differentiated epithelial cells of the small intestine and signals to the mesenchyme where it induces unidentified factors that negatively regulate intestinal epithelial precursor cell fate. Recently, genetic variants in the Hh pathway have been linked to the development of inflammatory bowel disease. METHODS We deleted Ihh from the small intestinal epithelium in adult mice using Cyp1a1-CreIhh(fl/fl) conditional Ihh mutant mice. Intestines were examined by immunohistochemistry, in situ hybridization, and real-time polymerase chain reaction. RESULTS Deletion of Ihh from the intestinal epithelium initially resulted in a proliferative response of the intestinal epithelium with lengthening and fissioning of crypts and increased Wnt signaling. The epithelial proliferative response was associated with loss of bone morphogenetic protein and Activin signaling from the epithelium of the villus and crypts, respectively. At the same stage we observed a substantial influx of fibroblasts and macrophages into the villus core with increased mesenchymal transforming growth factor-β signaling and deposition of extracellular matrix proteins. Prolonged loss of Ihh resulted in progressive leukocyte infiltration of the crypt area, blunting and loss of villi, and the development of intestinal fibrosis. CONCLUSIONS Loss of Ihh initiates several events that are characteristic of an intestinal wound repair response. Prolonged loss resulted in progressive inflammation, mucosal damage, and the development of intestinal fibrosis. Ihh is a signal derived from the superficial epithelial cells that may act as a critical indicator of epithelial integrity.

Rage signalling promotes intestinal tumourigenesis

Development of colon cancer is a multistep process that is regulated by intrinsic and extrinsic cellular signals. Extrinsic factors include molecular patterns that are derived from either pathogens (PAMPs) or cellular damage (DAMPs). These molecules can promote tumourigenesis by activation of the innate immune system, but the individual contribution of ligands and their receptors remains elusive. The receptor for advanced glycation end products (Rage) is a pattern recognition receptor that binds multiple ligands derived from a damaged cell environment such as Hmgb1 and S100 protein. Here we show that Rage signalling has a critical role in sporadic development of intestinal adenomas, as ApcMin/+ Rage−/− mice are protected against tumourigenesis.

Hedgehog signaling and maintenance of homeostasis in the intestinal epithelium.

Homeostasis of the rapidly renewing intestinal epithelium depends on a balance between cell proliferation and loss. Indian hedgehog (Ihh) acts as a negative feedback signal in this dynamic equilibrium. We discuss recent evidence that Ihh may be one of the key epithelial signals that indicates epithelial integrity to the underlying mesenchyme.

Stromal Indian Hedgehog Signaling Is Required for Intestinal Adenoma Formation in Mice

BACKGROUND & AIMS Indian hedgehog (IHH) is an epithelial-derived signal in the intestinal stroma, inducing factors that restrict epithelial proliferation and suppress activation of the immune system. In addition to these rapid effects of IHH signaling, IHH is required to maintain a stromal phenotype in which myofibroblasts and smooth muscle cells predominate. We investigated the role of IHH signaling during development of intestinal neoplasia in mice. METHODS Glioma-associated oncogene (Gli1)-CreERT2 and Patched (Ptch)-lacZ reporter mice were crossed with Apc(Min) mice to generate Gli1CreERT2-Rosa26-ZSGreen-Apc(Min) and Ptch-lacZ-Apc(Min) mice, which were used to identify hedgehog-responsive cells. Cyp1a1Cre-Apc (Apc(HET)) mice, which develop adenomas after administration of β-naphthoflavone, were crossed with mice with conditional disruption of Ihh in the small intestine epithelium. Apc(Min) mice were crossed with mice in which sonic hedgehog (SHH) was overexpressed specifically in the intestinal epithelium. Intestinal tissues were collected and analyzed histologically and by immunohistochemistry and quantitative reverse-transcription polymerase chain reaction. We also analyzed levels of IHH messenger RNA and expression of IHH gene targets in intestinal tissues from patients with familial adenomatous polyposis (n = 18) or sessile serrated adenomas (n = 15) and normal colonic tissue from control patients (n = 12). RESULTS Expression of IHH messenger RNA and its targets were increased in intestinal adenomas from patients and mice compared with control colon tissues. In mice, IHH signaling was exclusively paracrine, from the epithelium to the stroma. Loss of IHH from Apc(HET) mice almost completely blocked adenoma development, and overexpression of SHH increased the number and size of adenomas that developed. Loss of IHH from Apc(HET) mice changed the composition of the adenoma stroma; cells that expressed α-smooth muscle actin or desmin were lost, along with expression of cyclooxygenase-2, and the number of vimentin-positive cells increased. CONCLUSIONS Apc mutant epithelial cells secrete IHH to maintain an intestinal stromal phenotype that is required for adenoma development in mice.

Role of mast cells in colorectal cancer development, the jury is still out.

The link between inflammation and colorectal cancer development is becoming increasingly clear. It had long been recognized that patients with inflammatory bowel disease are at an increased risk of colon cancer. Evidence from experimental animals now also implicates the innate immune system in the development of sporadically occurring intestinal adenomas, the precursors to colorectal cancer. Here we discuss the interaction between the immune system and the adenoma to carcinoma sequence with a special emphasis on the role of mast cells which may play a key role in adenoma development. This article is part of a Special Issue entitled: Mast cells in inflammation.

Stromal Hedgehog signalling is downregulated in colon cancer and its restoration restrains tumour growth

A role for Hedgehog (Hh) signalling in the development of colorectal cancer (CRC) has been proposed. In CRC and other solid tumours, Hh ligands are upregulated; however, a specific Hh antagonist provided no benefit in a clinical trial. Here we use Hh reporter mice to show that downstream Hh activity is unexpectedly diminished in a mouse model of colitis-associated colon cancer, and that downstream Hh signalling is restricted to the stroma. Functionally, stroma-specific Hh activation in mice markedly reduces the tumour load and blocks progression of advanced neoplasms, partly via the modulation of BMP signalling and restriction of the colonic stem cell signature. By contrast, attenuated Hh signalling accelerates colonic tumourigenesis. In human CRC, downstream Hh activity is similarly reduced and canonical Hh signalling remains predominantly paracrine. Our results suggest that diminished downstream Hh signalling enhances CRC development, and that stromal Hh activation can act as a colonic tumour suppressor.

ER stress induces epithelial differentiation in the mouse oesophagus

Objective Stress in the endoplasmic reticulum (ER) leads to activation of the unfolded protein response (UPR). Xbp1, a key component of the UPR has recently been linked to the risk of developing oesophageal squamous cell carcinoma, suggesting an important role for the UPR in the oesophageal epithelium. Here we examined the role of ER stress and the UPR in oesophageal epithelial homoeostasis. Design We examined the expression of components of the UPR in the oesophageal epithelium. We used a pharmacological approach and a genetic approach to examine the effects of ER stress in vivo in the mouse oesophagus. The oesophagus of these mice was examined using immunohistochemistry and real-time reverse transcription (RT)-PCR. Results Components of the UPR were heterogeneously expressed in the basal layer of the epithelium. Induction of ER stress by 24-h treatment with thapsigargin resulted in depletion of proliferating cells in the basal layer of the oesophagus and induced differentiation. We next activated the UPR by inducible deletion of the major ER chaperone Grp78 in Ah1Cre-Rosa26-LacZ-Grp78−/− mice in which mutant cells could be traced by expression of LacZ. In these mice LacZ-positive mutant cells in the basal layer lost their proliferative capacity, migrated towards the oesophageal lumen and were replaced by LacZ-negative non-mutant cells. We observed no apoptosis in mutant cells. Conclusions These results show that ER stress induces epithelial differentiation in precursor cells in the oesophageal epithelium. This UPR induced differentiation may serve as a quality control mechanism that protects against oesophageal cancer development.

Rage mediated DAMP signaling in intestinal tumorigenesis

In the intestine, a large variety of pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) can instigate innate immune responses, which have been shown to promote colorectal carcinogenesis. We have recently demonstrated an important role for the receptor for advanced glycation end products (Rage) in intestinal adenoma formation. Rage is a receptor for DAMPs that are present in several proteins produced in intestinal adenomas. We found that Rage signaling upholds a pro-inflammatory milieu through a feed-forward loop that stimulates the production of its own ligands.

Indian Hedgehog Suppresses a Stromal Cell–Driven Intestinal Immune Response

Background & Aims Upon intestinal epithelial damage a complex wound healing response is initiated to restore epithelial integrity and defend against pathogenic invasion. Epithelium-derived Indian Hedgehog (Ihh) functions as a critical sensor in this process. Signaling occurs in a paracrine manner because the receptor for Ihh is expressed only in the mesenchyme, but the exact Hedgehog target cell has remained elusive. The aim of this study was to elucidate further the nature of this target cell in the context of intestinal inflammation. Methods Hedgehog activity was modulated genetically in both cell type–specific and body-wide models and the resulting animals were analyzed for gene expression profiles and sensitivity for dextran sodium sulfate (DSS) colitis. To characterize the Hedgehog target cell, Gli1-CreERT2-Rosa26-ZsGreen animals were generated, which express ZsGreen in all Hedgehog-responsive cells. These cells were characterized using flow cytometry and immunofluorescence. Results Loss of Indian Hedgehog from the intestinal epithelium resulted in a rapid increase in expression of inflammation-related genes, accompanied by increased influx of immune cells. Animals with epithelium-specific deletion of Ihh or lacking the Hedgehog receptor Smoothened from Hedgehog target cells were more sensitive to DSS colitis. In contrast, specific deletion of Smoothened in the myeloid compartment did not alter the response to DSS. This suggests that Hedgehog signaling does not repress intestinal immunity through an effect on myeloid cells. Indeed, we found that Hedgehog-responsive cells expressed gp38, smooth muscle actin, and desmin, indicating a fibroblastic nature. Ihh signaling inhibited expression of C-X-C motif chemokine ligand 12 (CXCL12) in fibroblasts in vitro and in vivo, thereby impairing the recruitment of immune cells. Conclusions We show that epithelium-derived Indian Hedgehog signals exclusively to fibroblasts in the intestine. Loss of Ihh leads to a rapid immune response with up-regulation of fibroblast-derived CXCL12, and migration of immune cells into the lamina propria.