OC.09.6 HUMAN DERIVED ENTEROGLIAL CELLS TOLL-LIKE RECEPTORS MRNA EXPRESSION AND MODULATION BY PATHOGENS AND PROBIOTIC BACTERIA

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only the difference in IL-6 expression reached the level of significance (p = 0.0243; Mann-Whitney). Levels of IL6 (0.0008±0.0003; 0.0002-0.0015), IFNG (0.0020±0.0009; 0-0.0040), and IL21 (0.0010±0.0005; 0-0.0021) detected in specimens from GS subjects did not significantly differ from those in control subjects or CD. Conclusions: IL-6 is expressed at significantly increased levels in the CD but not GS mucosa, a finding in line with its role in Th17-cell differentiation. These findings support the idea that, relative to CD, GS is associated with more limited involvement of the adaptive immune system. # E. Small bowel diseases - 1. Malabsorption syndromes

OC.09.4 SEROLOGICAL EXPRESSION OF ADULT COELIAC DISEASE IS PROPORTIONAL TO THE EXTENT OF MUCOSAL DAMAGE A. Licata ∗ , C. Randazzo, M. Cappello, G. Butera, F. Barbaria, G. Rizzuto, P. Almasio, A. Craxì Cattedra di Gastroenterologia, Di.Bi.M.I.S., Università di Palermo, Palermo Background and aim: The clinical spectrum of coeliac disease (CD) includes “typical” and “atypical” presentations and occasionally discovered silent forms. Duodenal villous atrophy may be partial or subtotal and disease distribution in the ileum patchy or diffuse. We aimed to define the relation of serological markers of CD (IgA anti-actin antibodies: AAA; antitransglutaminase antibodies: TGA; anti-endomysial antibodies: AEA) with the presentation and severity of intestinal mucosal damage. Material and methods: All consecutive out- and in-patients (n=88) presenting between January 2004 and December 2008 with a final diagnosis of CD by NIH Consensus criteria were investigated. The following items were recorded: abdominal pain, diarrhoea, malabsorption, iron deficiency anemia, raised aminotransferases. TGA were assessed by EIA, and AEA and AAA by indirect immunofluorescence. Three or more baseline intestinal biopsies were taken and scored according to Marsh as: I-IIIA partial villous atrophy (PVA); IIIB-C total villous atrophy (TVA). Twenty subjects with a final diagnosis of irritable bowel syndrome undergoing upper G.I. endoscopy acted as controls. Results: Sixty-eight patients had a “de novo“ diagnosis of CD. Fifteen (22.1%) were men; the mean age was 36.5 (range 15-80 yrs). TGA/AEA were positive in 49 (72.1%) patients and in none of 20 controls. Typical disease was present in 33 (48.6%) patients, 26 (38.2%) showing an atypical presentation and 9 (13.2%) silent CD. Thirty (44.3%) patients had PVA and 38 (55.7)% TVA. Patients with typical pattern had abdominal pain, diarrhea and malabsorption; patient with atypical pattern showed high levels of aminotransferases, iron deficiency anemia or autoimmune disease (e.g. hypothryroidism, m. graves, diabetes type I, dermatitis herpetiforme) and ovarian disorders. AAA IgA were positive in 12 (17.6%) CD patients and 2 (10.0%) out of 20 controls. Presence of AEA and/or TGA and/or AAA was significantly associated to more severe mucosal damage (92.5% in TVA vs. 53.5% in PVA, p < 0.001). Twelve patients (17.6%) with AAA presented with TVA (p < 0.012). Conclusions: The severity of mucosal damage is the main factor governing the detectability of serological markers of CD. Although useful for mass screening, the sensitivity of serological testing is questionable among atypical or asymptomatic patients. Endoscopy remains the gold standard for diagnosis in these patients. # E. Small bowel diseases - 1. Malabsorption syndromes

OC.09.5 PREVALENCE OF SMALL INTESTINAL BACTERIAL OVERGROWTH IN PARKINSON’S DISEASE M. Gabrielli ∗ ,1 , P. Bonazzi 2 , E.C. Lauritano 1 , E. Scarpellini 1 , A. Bentivoglio 1 , F. Soleti 1 , L. Di Maurizio 1 , A. Fasano 1 , M. Petracca 1 , G. Gasbarrini 1 , P. Tonali 1 , A. Gasbarrini 1 1 Policlinico

Universitario “A. Gemelli”, Roma; 2 Policlinico Universitario “Ospedali Riuniti”, Ancona Background and aim: Autonomic disturbances such as dysphagia, delayed gastric emptying and constipation are common findings in patients with idio-

pathic Parkinson’s disease (IPD). They may favour the occurrence of small intestinal bacterial overgrowth (SIBO). SIBO is a malabsorption syndrome characterized by an increase in microrganisms = 106 bacteria/mL of jejunal juice. It may cause either impaired levo-dopa absorption or increased gastrointestinal symptoms in patients with IPD. Aim of the study was to assess the prevalence of SIBO in a population of IPD patients with respect to controls without IPD. Material and methods: IPD patients were enrolled from two different tertiary Neurologic Centers (Catholic University of Rome, 42 patients, and University of Ancona, 30 patients). Mean age of enrolled patients was 66±10 years (males: 38). Disease duration, disease severity assessed by Unified PD Rating Scale (UPDRS-III), and L-dopa equivalent daily dose (LEDD, mg/d) were recorded in all patients. The control population consisted of 36 subjects without IPD and similar for sex and age enrolled from out-patients of Internal Medicine Department of the Catholic University of Rome. All subjects underwent glucose BT (GBT) to detect SIBO under standard conditions. The presence of SIBO was based on the occurrence of a peak of H2 values >12 parts per million above the basal value after ingestion of glucose 75 grams. Results: No significant difference about age, sex, and clinical characteristics of IPD patients from the two centres were found. A total of 39 out of 72 IPD patients showed positivity to GBT (54.2%) with respect to 3 out of 36 in the control group (8.3%), the difference resulting statistically significant (p<0.0001, OR 1.86, CI 3.65-46.27). Disease duration was the only factor associated to the GPT positivity status in IPD patients at multivariate analysis. Conclusions: Results from the present study show that SIBO is highly prevalent in Parkinson’s Disease, especially in patients with longer disease duration. Impaired intestinal motility could be responsible of such findings. Further studies are needed to assess the effect of SIBO decontamination on gastrointestinal symptoms and levo-dopa absorption in patients with IPD. # E. Small bowel diseases - 2. Intestinal microflora and infections

OC.09.6 HUMAN DERIVED ENTEROGLIAL CELLS TOLL-LIKE RECEPTORS MRNA EXPRESSION AND MODULATION BY PATHOGENS AND PROBIOTIC BACTERIA F. Turco ∗ ,1 , C. Cirillo 1 , G. Sarnelli 1 , A. Mango 1 , M. Cammarota 2 , M. Giuliano 2 , M. Cartenì 2 , R. Cuomo 1 1

Department of Clinical and Experimental Medicine, University of Naples Federico II, Naples; 2 Department of Experimental Medicine, Biotechnology and Molecular Biology Section, Second University of Naples, Naples Background and aim: Enteric glial cells (EGC) are involved in intestinal inflammation and can be activated by exogenous stimuli, and foreign antigens (i.e. gliadin). Toll-Like Receptors (TLRs) are immune receptors able to recognize pathogen conserved molecular patterns. Whether EGC express TLRs or respond to bacterial stimuli is unknown. We aimed to investigate if TLRs are expressed by human EGC and if their expression is differentially modulated by pathogens or probiotic bacteria. Material and methods: Pure EGC were obtained according to method previously described by our group. Briefly, myenteric plexus preparations were isolated from ileum of patients undergoing surgery and enzimatically dissociated. Ganglia were plated and cell cultures were grown to subconfluence. After 21 days, EGC were purified using Dynal Magnet® and characterized by immunofluorescence to confirm the absence of contaminating cells (i.e. fibroblasts). TLRs 1, 2, 4, 7 and 9 mRNA expression were evaluated at baseline and after 24 hours (h) incubation of EGC with the probiotic strain Lactobacillus Paracasei F19 (LP F19) and the pathogen Enteroinvasive Escherichia Coli (EIEC). The bacteria/cells ratio was 0.1/1 and 100/1. Quantitative Real Time PCR was performed to evaluate specific mRNAs fold increases respect to untreated controls. Data were obtained with 3 experiments and expressed as mean ± SD. Results: Human purified EGC physiologically expressed TLR 1, 2, 4, 7 and 9 mRNAs, with TLR 1 and 4 transcripts being the most expressed. With a bacteria/cells ratio of 0.1/1, EIEC induced a higher significant increase than LP F19 of TLR 2 (4.1±0.1 and 2.7±0.4; p<0.05), TLR 7 (8.0±0.2 and 0.6±0.2; p<0.05) and TLR 9 (8.8±1.7 and 1.0±0.2; p<0.05). Conversely,

Abstracts / Digestive and Liver Disease 42S (2010) S61–S192 only a trend to an increased expression of TLR 4 was observed with LP F19 respect to EIEC (2.4±0.1 and 1.3±0.4; p=NS). At 100/1 bacteria/cell ratio EIEC determined an 80% of cellular mortality, while LP F19 induced an higher expression of TLR 7 (17.8±4.1) and 9 (21.2±4.9) respect to lowest bacteria/cell ratio, while the other TLRs remained unmodified. Conclusions: We showed, for the first time, that pure cultures of human EGC express TLRs’ mRNA and are able to discriminate between pathogen and probiotic bacteria. This data support the hypothesis that enteric glia is involved in the host-pathogen interaction. # E. Small bowel diseases - 3. Probiotics

OC.10.1 NEUROPLASTIC CHANGES IN THE COLONIC MUCOSA OF PATIENTS WITH IRRITABLE BOWEL SYNDROME G. Barbara 1 , C. Cremon ∗ ,1 , L. Gargano 1 , V. Vasina 2 , G. Dothel 1 , G. Carini 1 , R. De Giorgio 1 , V. Stanghellini 1 , R. Cogliandro 1 , M. Tonini 3 , F. De Ponti 2 , R. Corinaldesi 1 1 Department

of Clinical Medicine and Center for Applied Biomedical Research, University of Bologna, Bologna; 2 Department of Pharmacology, University of Bologna, Bologna; 3 Department of Physiological and Pharmacological Sciences, University of Pavia, Pavia Background and aim: Immune and non-immune mediators released by gut mucosa play a role in the pathophysiology of irritable bowel syndrome (IBS). We hypothesize that soluble mediators induce phenotypic changes in gut mucosal innervation in IBS patients. The aims of the present study were to evaluate in IBS patients and healthy controls (HC): 1) Phenotypic changes in the colonic innervation, including the expression of the marker of neuronal sprouting growth-associated protein 43 (GAP-43); 2) source and expression of nerve growth factor (NGF); 3) impact of colonic mediators and NGF on neuronal cell differentiation and sprouting in human cultured neuronal cells. Material and methods: Mucosal biopsies were obtained from the descending colon of 18 IBS patients and 8 HC. The expression of neuronal specific enolase (NSE), GAP-43 and NGF expression were assessed by quantitative immunohistochemistry. Mucosal mediator release was quantified on colonic biopsies by means of specific immuno-enzymatic assay and/or semi-quantitative western blot. The impact of mucosal supernatants on nerve sprouting and differentiation was assessed on cultured neuronal cell lines (SH-Sy5Y) by means of morphometric analysis. Results: IBS patients showed a significant increase in the area of lamina propria occupied by NSE+ fibers as compared to HC (2.52% vs 1.48%; P<0.05). A significant increase in GAP-43+ fibers was documented in IBS patients in comparison with HC (1.75% vs 1.05%; P<0.05). NGF immunoreactivity was significantly increased in IBS patients (1.31% vs 0.79%; P<0.05). The majority of NGF+ cells also expressed tryptase immunolabeling, suggesting that mucosal mast cells represent the main source of NGF. NGF mucosal concentration and protein expression was significantly increased in most IBS samples in comparison with HC. Mucosal supernatants obtained from IBS patients induced a 60% neuronal sprouting on cultured neuronal cell lines, which was similar to that obtained with the control stimulus retinoic acid (68%) and significantly higher than that obtained with HC (41%) (P<0.05). Neuronal sprouting was partly NGF-dependent, as it was significantly reduced by NGF neutralization. Conclusions: Nerve fiber density and growth are increased in IBS patients. NGF, mainly from mast cell source, is also increased and participates to neuronal sprouting. Our results provide a new mechanism which may contribute to the pathophysiology of IBS. # C. Functional disorders - 2. IBS

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OC.10.2 MUSCULAR COORDINATION AND BOWEL TRAINING ARE MORE EFFECTIVE IN NORMAL- AND SLOW-TRANSIT CONSTIPATION THAN IN PELVIC FLOOR DYSSYNERGIA M. Baracca ∗ , I. Vantini, A. Amodio, A. Massella, A. Gabbrielli, A. Garribba, A. Rostello, L. Frulloni, L. Benini Department of Biomedical Sciences, Gastroenterology Unit, University of Verona, Verona Background and aim: Specific types of constipation may deserve different treatments. Muscular coordination training was described to benefit patients with pelvic floor dyssynergia. Aim: the study was aimed at investigating the effects of muscular coordination and bowel training on specific types of chronic constipation. Material and methods: Eighty-seven patients with chronic constipation (31 normal transit, 30 slow-transit; 18 pelvic floor dyssynergia (PFD), 8 slowtransit with PFD), after 4-week run-in period, underwent a 16-week course of muscular coordination and bowel training. Colonic transit time was assessed at baseline. Anorectal manometry, relaxation of anal sphincter, rectal balloon expulsion test (BET) were carried-out at baseline and after a 16-week treatment period. Clinical parameters, spontaneous and complete bowel movements (SCBM), satisfaction rates (Likert scale), and relief from Rome III criteria for constipation were assessed at baseline and after 16 weeks. Statistical analysis was carried-out by parametric and non-parametric methods, and logistic regression analysis. Results: SCBM/week, straining-associated movements, call for stool and consistency, laxatives and enemas consumption improved in all groups (p<0.05). Balloon expulsion test, but not manometric anal relaxation during straining, improved in 50% of PFD patients (p=0.03). Per cent of fully or partially satisfied patients and patients showing relief from the Rome III criteria for constipation were equally and significantly (p<0.05) greater in both normal (77 and 61% respectively) and slow-transit constipation (70 and 60% respectively)(also if associated with PFD) than in pelvic floor dyssynergia (39% and 16.7% respectively). At logistic regression analysis high number of enemas/week at baseline (p=0.01), lower bowel movement/week and BET failure (p=0.04) predicted a worse outcome. Conclusions: Muscular coordination and bowel training benefit patients with normal- and slow-transit constipation, where is more effective than in those with pelvic floor dyssynergia, whose underlying defecation disorder was not successfully reversed. # F. Colorectal diseases - 1. Disorders of defecation

OC.10.3 LONG-TERM PROGNOSTIC IMPACT OF ISOLATED TUMOUR CELLS IN PN0 COLORECTAL CANCER: A PROSPECTIVE MONOINSTITUTIONAL STUDY C. Mescoli ∗ , L. Albertoni, S. Pucciarelli, L. Giacomelli, M. Fassan, R. Clemente, D. Nitti, M. Rugge Ospedale Padova, Padova Background and aim: The prevalence of Isolated Tumour Cells (ITC) in regional lymph-nodes (LNs) from pN0 colorectal cancer (CRC) is controversial and it has never been prospectively assessed in large series of consecutive and prospectively collected patients. Material and methods: 5016 LNs from 308 consecutive, prospectively collected pN0-CRCs, were considered (mean number of LNs/patient: 16.2; range: 1-107). ITC prevalence was assessed (MNF116 immunohistochemistry, DakoDenmark). From each collected LN, two histological sections (5 micron thick; 70-100 micron apart) were considered. Cancer outcome was clinically assessed as disease-free survival (mean follow-up = 58.4 months, range = 1-77). Results: ITC were documented in 156/308 (50.5%) patients (ITC+ve LNs: 496/5016 [9.9%]). The mean number of harvested LNs significantly differed in ITC+ve versus ITC-ve group (18 versus 13). The ITC-status significantly correlated with both p-Stage and vascular tumour invasion. CRC relapsed in 21/308 (7%) cases. Fiftheen out the 21 cancer relapses (70%) were docu-