Enterocyte proliferation and signaling are constitutively altered in celiac disease.

Celiac disease (CD) occurs frequently, and is caused by ingestion of prolamins from cereals in subjects with a genetic predisposition. The small intestinal damage depends on an intestinal stress/innate immune response to certain gliadin peptides (e.g., A-gliadin P31-43) in association with an adaptive immune response to other gliadin peptides (e.g., A-gliadin P57-68). Gliadin and peptide P31-43 affect epithelial growth factor receptor (EGFR) signaling and CD enterocyte proliferation. The reason why the stress/innate immune and proliferative responses to certain gliadin peptides are present in CD and not in control intestine is so far unknown. The aim of this work is to investigate if, in CD, a constitutive alteration of enterocyte proliferation and signaling exists that may represent a predisposing condition to the damaging effects of gliadin. Immunofluorescence and immunohistochemistry were used to study signaling in CD fibroblasts and intestinal biopsies. Western blot (WB) analysis, immunoprecipitation, and quantitative PCR were also used. We found in CD enterocytes enhancement of both proliferation and Epidermal Growth Factor Receptor (EGFR)/ligand system. In CD enterocytes and fibroblasts we found increase of the phosphorylated downstream signaling molecule Extracellular Signal Regulated Kinase (ERK); block of the ERK activation normalizes enterocytes proliferation in CD mucosa. In conclusion the same pathway, which gliadin and gliadin peptide P31-43 can interfere with, is constitutively altered in CD cells. This observation potentially explains the specificity of the damaging effects of certain gliadin peptides on CD intestine.

A celiac cellular phenotype, with altered LPP sub-cellular distribution, is inducible in controls by the toxic gliadin peptide P31-43.

Celiac disease (CD) is a frequent inflammatory intestinal disease, with a genetic background, caused by gliadin-containing food. Undigested gliadin peptides P31-43 and P57-68 induce innate and adaptive T cell-mediated immune responses, respectively. Alterations in the cell shape and actin cytoskeleton are present in celiac enterocytes, and gliadin peptides induce actin rearrangements in both the CD mucosa and cell lines. Cell shape is maintained by the actin cytoskeleton and focal adhesions, sites of membrane attachment to the extracellular matrix. The locus of the human Lipoma Preferred Partner (LPP) gene was identified as strongly associated with CD using genome-wide association studies (GWAS). The LPP protein plays an important role in focal adhesion architecture and acts as a transcription factor in the nucleus. In this study, we examined the hypothesis that a constitutive alteration of the cell shape and the cytoskeleton, involving LPP, occurs in a cell compartment far from the main inflammation site in CD fibroblasts from skin explants. We analyzed the cell shape, actin organization, focal adhesion number, focal adhesion proteins, LPP sub-cellular distribution and adhesion to fibronectin of fibroblasts obtained from CD patients on a Gluten-Free Diet (GFD) and controls, without and with treatment with A-gliadin peptide P31-43. We observed a "CD cellular phenotype" in these fibroblasts, characterized by an altered cell shape and actin organization, increased number of focal adhesions, and altered intracellular LPP protein distribution. The treatment of controls fibroblasts with gliadin peptide P31-43 mimics the CD cellular phenotype regarding the cell shape, adhesion capacity, focal adhesion number and LPP sub-cellular distribution, suggesting a close association between these alterations and CD pathogenesis.

An undigested gliadin peptide activates innate immunity and proliferative signaling in enterocytes: the role in celiac disease.

BACKGROUND: On ingestion of gliadin, the major protein component of wheat and other cereals, the celiac intestine is characterized by the proliferation of crypt enterocytes with an inversion of the differentiation/proliferation program. Gliadins and A-gliadin peptide P31-43, in particular, act as growth factors for crypt enterocytes in patients with celiac disease (CD). The effects of gliadin on crypt enterocyte proliferation and activation of innate immunity are mediated by epidermal growth factors (EGFs) and innate immunity mediators [interleukin 15 (IL15)]. OBJECTIVE: The aim of this study was to determine the molecular basis of proliferation and innate immune response to gliadin peptides in enterocytes. DESIGN: The CaCo-2 cell line was used to study EGF-, IL15-, and P31-43-induced proliferation. Silencing messenger RNAs and blocking EGF receptor and IL15 antibodies have been used to study proliferation in CaCo-2 cells and intestinal biopsy samples from patients with CD and control subjects. RESULTS: In the CaCo-2 cell model, IL15 and EGF cooperated to induce proliferation in intestinal epithelial cells at both the transcriptional and posttranscriptional levels, and the respective receptors interacted to activate each other's signaling. In addition, the effects of the P31-43 peptide on CaCo-2 cell proliferation and downstream signaling were mediated by cooperation between EGF and IL15. The increased crypt enterocyte proliferation in intestinal biopsy samples from patients with CD was reduced by EGF receptor and IL15 blocking antibodies only when used in combination. CONCLUSIONS: EGF receptor/IL15R-α cooperation regulates intestinal epithelial cell proliferation induced by EGF, IL15, and the gliadin peptide P31-43. Increased proliferation of crypt enterocytes in the intestine of CD patients is mediated by EGF/IL15 cooperation.