Gliadin Peptide P31-43 Localises to Endocytic Vesicles and Interferes with Their Maturation

BACKGROUND: Celiac Disease (CD) is both a frequent disease (1:100) and an interesting model of a disease induced by food. It consists in an immunogenic reaction to wheat gluten and glutenins that has been found to arise in a specific genetic background; however, this reaction is still only partially understood. Activation of innate immunity by gliadin peptides is an important component of the early events of the disease. In particular the so-called "toxic" A-gliadin peptide P31-43 induces several pleiotropic effects including Epidermal Growth Factor Receptor (EGFR)-dependent actin remodelling and proliferation in cultured cell lines and in enterocytes from CD patients. These effects are mediated by delayed EGFR degradation and prolonged EGFR activation in endocytic vesicles. In the present study we investigated the effects of gliadin peptides on the trafficking and maturation of endocytic vesicles. METHODS/PRINCIPAL FINDINGS: Both P31-43 and the control P57-68 peptide labelled with fluorochromes were found to enter CaCo-2 cells and interact with the endocytic compartment in pulse and chase, time-lapse, experiments. P31-43 was localised to vesicles carrying early endocytic markers at time points when P57-68-carrying vesicles mature into late endosomes. In time-lapse experiments the trafficking of P31-43-labelled vesicles was delayed, regardless of the cargo they were carrying. Furthermore in celiac enterocytes, from cultured duodenal biopsies, P31-43 trafficking is delayed in early endocytic vesicles. A sequence similarity search revealed that P31-43 is strikingly similar to Hrs, a key molecule regulating endocytic maturation. A-gliadin peptide P31-43 interfered with Hrs correct localisation to early endosomes as revealed by western blot and immunofluorescence microscopy. CONCLUSIONS: P31-43 and P57-68 enter cells by endocytosis. Only P31-43 localises at the endocytic membranes and delays vesicle trafficking by interfering with Hrs-mediated maturation to late endosomes in cells and intestinal biopsies. Consequently, in P31-43-treated cells, Receptor Tyrosine Kinase (RTK) activation is extended. This finding may explain the role played by gliadin peptides in inducing proliferation and other effects in enterocytes from CD biopsies

Involvement of innate immunity in the development of inflammatory and autoimmune diseases.

Initial events and effector mechanisms of most inflammatory and autoimmune diseases remain largely unknown. Dysfunction of the innate and adaptive immune systems associated with mucosae (the major interface between the organism and its environment, e.g., microbiota, food) can conceivably cause impairment of mucosal barrier function and development of localized or systemic inflammatory and autoimmune processes. Animal models help in elucidating the etiology and pathogenetic mechanisms of human diseases, such as the inflammatory bowel diseases, Crohn's disease and ulcerative colitis, severe chronic diseases affecting the gut. To study the role of innate immunity and gut microbiota in intestinal inflammation, colitis was induced by dextran sulfate sodium (DSS) in mice with severe combined immunodeficiency (SCID). Conventionally reared (microflora-colonized) SCID mice displayed severe inflammation like that seen in immunocompetent Balb/c mice, whereas only minor changes appeared in the intestinal mucosa of DSS-fed gnotobiotic germ-free SCID mice. The presence of microflora facilitates the inflammation in DSS-induced colitis that develops in immunodeficient SCID mice, that is, in the absence of T and B lymphocytes. Celiac disease, a chronic autoimmune small bowel disorder, afflicts genetically susceptible individuals with wheat gluten intolerance. We showed that, in contrast with any other food proteins, wheat gliadin and its peptic fragments activate mouse macrophages and human monocytes to produce proinflammatory cytokines through the nuclear factor-kappaB signaling pathway. Activation of innate immunity cells by food proteins or components from gut microbiota thus could participate in the impairment of intestinal mucosa and the development of intestinal and/or systemic inflammation

Review article: future research on coeliac disease - a position report from the European multistakeholder platform on coeliac disease (CDEUSSA).

BACKGROUND: CDEUSSA is a Specific Support Action project from the Sixth Framework Programme Priority of the European Union (EU). Its aim is to bring together basic and applied research in the area of coeliac disease (CD). This paper reviews the main issues that are a result of the CDEUSSA initiative. AIM: To identify the major issues in need of investigation in the areas of clinical aspects, treatment, prevention and public health. METHODS: Key stakeholders, representing a wide range of knowledge with crucial importance for CD research and practice, have participated in two workshops aimed at identifying and proposing to the EU, as high priority research, topics in the areas of clinical aspects, treatment, prevention and public health. RESULTS: In public health, the overall goal should be to improve quality of life of the European population by implementing primary prevention strategies, early diagnosis and improved treatments for CD. New treatment strategies need to be developed. The option of primary prevention should be fully explored, which requires combined epidemiological, clinical and basic scientific research efforts. Such studies should also consider the importance of gene-environment interactions in the development of CD. Increased knowledge is needed on the natural history of CD. Diagnostic criteria need to be revised. CONCLUSIONS: To achieve these goals, a collaboration of the stakeholders is fundamental, including research and patient organizations, as well as industries within both diagnostics and food production