P.06.7 GLIADIN EFFECT ON THE OXIDATIVE BALANCE AND DNA DAMAGE IN CACO-2 CELL LINE

Abstracts of the 22nd National Congress of Digestive Diseases / Digestive and Liver Disease 48S2 (2016) e67–e231

Results: Results. Smad7 protein, but not RNA, expression was increased in RCD samples compared to normal controls, while there was no difference between ACD or ICD and normal controls. In RCD duodenal mucosa, Smad7-positive cells were abundant in both the epithelial and lamina propria compartments. TGFb1 protein content did not differ among groups. However, TGFb1-associated Smad2/3 phosphorylation was less pronounced in RCD as compared to controls. Knockdown of Smad7 in RCD biopsy samples reduced RNA expression of interleukin (IL)-15, IL-6 and TNFa, all cytokines that are over-produced in RCD mucosa. Conclusions: Conclusions. High Smad7 associates with defective TGFb1 signalling in RCD, thus suggesting a novel mechanism by which the ongoing mucosal inflammation is sustained and perpetuated in this disorder.

P.06.6 SUBTYPES OF CHRONIC GASTRITIS IN PATIENTS WITH COELIAC DISEASE BEFORE AND AFTER GLUTEN-FREE DIET Gabrieli D.*, Valerii G., Ciccone F., Di Ruscio M., Serva D., Capannolo A., Viscido A., Melideo D., Frieri G., Necozione S., Latella G. Università degli studi dell’Aquila, L’Aquila, Italy Background and aim: Chronic gastritis appears to be more common in patients with coeliac disease (CD). Aim of this study is to evaluate the frequency of lymphocytic gastritis (LG), chronic active gastritis (CAG) and chronic inactive gastritis (CIG) in a cohort of patients with CD, and their histological changes after treatment with a gluten-free diet. Material and methods: A five-year prospective study including all consecutive patients with a new diagnosis of CD performed at our GI Unit, in the period between January 2010 and January 2015. All gastric and duodenal biopsy specimens at the time of the diagnosis of CD and at the first endoscopic control after 18-24 months on gluten free diet were analyzed. CD diagnosis was made in the presence of anti-tissue transglutaminases and/or anti-endomisial antibodies associated with specific alterations at histological evaluation of duodenal biopsies, according to the modified Marsh-Oberhuber classification. Gastric lesions were classified according to the Updated Sydney System. Giemsa staining was used for histological diagnosis of Helicobacter pylori infection and immunohistochemical staining for the diagnosis of LG, defined as a dense proliferation of intraepithelial lymphocytes (more than 25 lymphocytes per 100 epithelial cells). Anti-gastric parietal cell antibodies were assayed by enzyme-linked immunosorbent assay (ELISA). Demographic, clinical, and laboratory data were collected. Results: 250 patients with CD were enrolled (191 F, 59 M, mean age 34 years at the diagnosis). At the time of CD diagnosis, histological examination showed normal gastric mucosa in 78 patients (31.2%), LG in 32 (12.8%), CAG in 74 (29.6%), and CIG in 66 (26.4%). Out of 32 patients with LG, 20 (62.5%) were H. pylori negative and all of them showed an improvement of gastritis after gluten free diet. Out of 74 patients with CAG, 30 (40.5%) were H. pylori negative and one third of them showed an improvement of gastritis after gluten free diet. Out of 66 patients with CIG, 63 (95.4%) patients were H. pylori negative. LG is significantly associated to histological improvement after gluten-free diet compared to other types of gastritis (p= 0.0039). Conclusions: Subtypes of gastritis have different probability to be influenced by the gluten-free diet. LG is present in a significant number (13%) of CD patients and seems to improve as well as duodenal lesions after gluten free diet. Two thirds of LG are not associated with H. pylori infection. Both CAG and CIG are also

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significantly associated with coeliac disease, despite less influenced by gluten free diet.

P.06.7 GLIADIN EFFECT ON THE OXIDATIVE BALANCE AND DNA DAMAGE IN CACO-2 CELL LINE Monguzzi E.*2, Marabini L.1, Roncoroni L.3, Doneda L.3, Conte D.2, Elli L.3 Department of Pharmacological and Biomolecular Sciences, Università degli Studi di Milano, Milano, Italy, 2Department of Pathophysiology and Transplantation, Università degli Studi di MIlano, Milano, Italy, 3 Gastroenterology and Endoscopy Unit and Center for Prevention and Diagnosis of Celiac Disease, Fondazione IRCCS Cà Granda - Ospedale Maggiore Policlinico - Milan, Milano, Italy

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Background and aim: Wheat has been identified as a key environmental factor in different human disorders (celiac disease, food allergies and non-celiac gluten sensitivity). As its components (gliadins) can induce different cellular and biological noxious alterations, present study was aimed at evaluating the gliadin effect on oxidative/reductive balance and assessing the possible oxidative/ genotoxic damage. Material and methods: Caco-2 cells were exposed for 12-24 h to increasing concentrations (from 250 to 1000μg/mL) of digested gliadin, then investigating : i) cytotoxicity (MTT test); ii) oxidative balance (DCFDA ROS evaluation); iii) DNA damage by means of comet assay (Alkaline and with ENDO III e Fpg enzymes) to identify single and double strand DNA breaks; iv) transglutaminase type 2 (TG2) activity in different cellular compartments (including nucleus, cytoskeleton, membranes and cytoplasm) with ELISA assay. Results: Gliadin treatment did not significantly reduce cell viability after 24h at different concentration (250µg/mL-1000μg/mL). Conversely, gliadin induced a significant increase of ROS production after 12h of exposition at the concentrations of 500μg/ml and 1000μg/mL. Alkaline comet analysis revealed a DNA damage in all of the analyzed parameters (Tail length, Tail moment and % DNA), the damage being particularly evident at the dose of 1000μg/mL. Moreover, comet analysis with enzymes showed an increase in the delta tail moment at 1000μg/mL, consist of oxidative origin of the damage. After gliadin treatment, TG2 activity increased from 191% to 310% into the cytoskeleton and from 117% to 153% into the nucleus. The analysis of cellular compartments evidenced a TG2 translocation from the cytoplasm to the nucleus and cytoskeleton suggesting a proapoptotic process. Conclusions: Present findings demonstrated with a gliadin-induced genotoxic damage associated to both an oxidative imbalance and an apoptotic process.

P.06.8 SERUM 25-HYDROXYCHOLECALCIFEROL LEVELS AT CELIAC DISEASE DIAGNOSIS PREDICT BONE MINERAL DENSITY RECOVERY AFTER GLUTEN EXCLUSION Efthymakis K.*, Serio M., Sanelli R., Milano A., Laterza F., Bonitatibus A., Neri M. Department of Medicine and Ageing Sciences, “G. D’Annunzio” University and Foundation, Chieti, Italy Background and aim: Celiac disease (CD) patients at presentation variably show reduced bone mineral density (BMD) and altered bone-related serology, including serum 25-hydroxycholecalciferol (25[OH)]VitD). Gluten-free diet (GFD) has been shown to promote repair in children and to a smaller extent in adults. However, complete bone mineral recovery is uncertain and predictive markers are lacking.