In vivo and ex vivo examination of the safety of oats in coeliac disease

Abstract

Coeliac disease is a chronic autoimmune disorder of the small intestine characterised by infiltration of intraepithelial lymphocytes, crypt hyperplasia and villous atrophy. It develops in genetically susceptible individuals following exposure to gluten. The only treatment is dietary and all sources of gluten must be permanently removed from the diet. On the basis of studies carried out in the 1930s and 1940s, wheat, barley, rye and oats were deemed to be harmful for coeliac patients and the exclusion of these four grains was the basis of the gluten free diet. However, the true toxicity of oats has more recently been questioned and although a number of studies have indicated oats to be safe for coeliac patients there have also been reports of adverse effects as a result of oats addition to the gluten free diet. The gluten free diet is difficult to maintain and often high in fat and low in fibre, vitamins and other nutrients. The addition of oats to this diet would have both lifestyle and nutritional benefits. The aim of this thesis was to thoroughly examine the safety of oats in the gluten free diet. In the first part of this study, a large group of treated and newly diagnosed coeliac patients added pure oats to their gluten free diet for one year. During this time parameters including coeliac serology, symptoms and histology were examined for any signs of disease activation. In addition, the expression of Ki67, CD3 and CD8 were compares between patient biopsies taken at the start and the end of the study, to look for more subtle signs of change. No signs of disease activation were seen, indicating that pure oats can be safely included in the coeliac diet. In the next part of the study, the IN Cell Analyzer 1000, a system designed for automated analysis of cell-based assays, was adapted for use with tissue sections. This enabled us to attain a numerical measure of the amount of positive staining present on a tissue section. This was used to measure the expression of tTG and SM α-actin in biopsies taken from coeliac patients at the start and end of the oats study, as well biopsies from Marsh grade 0 and Marsh grade 3 coeliac patients. tTG expression is known to be upregulated in active coeliac disease; this was seen in the Marsh grade 3 biopsies but not in the biopsies from the oats study. Likewise, expression of SM α-actin was increased in the Marsh grade 3 biopsies but not in the biopsies from the oats study. This is the first time the expression of tTG and SM α-actin has been assessed in the investigation of oats tolerance in coeliac disease and provides further evidence of a lack of oats toxicity. Additionally, we demonstrated for the first time the co-expression of tTG and SM α-actin in myofibroblasts in the coeliac mucosa. Furthermore, this co-expression was found to be increased in the Marsh grade 3 biopsies compared to the Marsh grade 0 and oats study biopsies. In the last part of the study, duodenal biopsies from coeliac and control patients were cultured in the presence of PT avenin or PT gliadin. They were then stained for the expression of E-cadherin and cytokeratin 20 and imaged on a confocal microscope. Cytokeratin 20 staining was too inconsistent to draw any conclusions; however, E-cadherin staining showed changes in three out of five coeliac patients following culture with avenin. The concentration of suPAR was measured in culture supernatants and was increased in the same three coeliac patients. In addition, suPAR concentration was significantly increased in all coeliac biopsy supernatants compared to controls, regardless of culture condition. These final results indicate caution may still be necessary when introducing oats to the gluten free diet; however, this is based on the examination of a small number of patients.