P168 PREVENTION OF TNBS-INDUCED COLONIC FIBROSIS IN RATS BY BOSWELLIA AND SCUTELLARIA EXTRACTS

Abstracts of the 3rd ECCO Congress, Lyon, France, February 28–March 1, 2008 with Crohn' s disease (CD) submitted to curative ileal resection there is a high recurrence rate of Crohn' s lesions in the neoterminal ileum we hypothesize that colo-neoterminal ileum reflux could play a relevant role in determining the inflammatory process in the preanastomotic segment. Small Intestine Contrast Ultrasonography (SICUS) [2], performed after distension of small bowel lumen with macrogol solution, in addition to accurately assess presence, number, extension, and sites of small bowel lesions [3] enables to visualize the direction of flow of the contrast solution through the intestine. In particular SICUS can detect forward and backward flux through the colo-ileal anastomosis. Aim: To assess in CD patients submitted to curative ileal resection the presence of colo-ileal reflux and whether it is associated with the presence of lesion recurrence. Methods and Patients: Thirty two consecutive patients with CD (M19, age 44±14,7 yrs) were submitted to surgery for obstruction (20 pts), fistulae (5 pts), both (5 pts), and perforation (2 pts). After surgery, clinical evaluation, SICUS, ileo-colonscopy, and endoscopy Rutgeerts score were performed at 1 year intervals. Continuous US observation of the tract including neo-terminal ileum, ileo-colonic anastomosis, and anastomotic colon was performed for 20 min and recorded on video-tape after the ingestion of 375 ml macrogol solution. Results: During a follow-up period of 46.5±21.5 months, 11/32 (34%) of patients (7 M, age 39±14 yrs) did not have any recurrent lesions and were asymptomatic; 21/32 (66%) patients (12 M, age 46.7±15 yrs) developed lesion recurrence at SICUS/endoscopy at the level of neoterminal ileum. Drug treatment, mainly mesalazine and azathioprine, age at diagnosis, gender and length of ileal resection did not differ between patients with and without recurrence. Colo-ileal reflux was detected in 59% of the investigated patients; in 75% of patients with and in 36% without CD lesion recurrence. Conclusions: These preliminary results show that colo-ileal reflux is a frequent occurrence after ileo-colon anastomosis and support the hypothesis that colo-ileal reflux can be involved in the inflammation process and Crohn' s lesion recurrence in the neoterminal ileum after curative ileal resection. References: 1. Pallotta et al Neurogastroenterology 2006. 2. Pallotta et al Lancet 1999. 3. Pallotta et al IBD 2005.

P168 PREVENTION OF TNBS-INDUCED COLONIC FIBROSIS IN RATS BY BOSWELLIA AND SCUTELLARIA EXTRACTS G. Latella 1 , R. Sferra 1, A. Vetuschi 1 , G. Zanninelli 1, A. D' Angelo 1 , V. Catitti 1 , E. Gaudio 2 , R. Caprilli 2 . 1 University of L' Aquila, L' Aquila, Italy; 2 University of Rome "La Sapienza", Rome, Italy Background: Currently,no effective preventive measures or medical therapies are available for intestinal fibrosis and thus surgery remains the only available strategy in the management of fibrostenotic enteropathies, especially Crohn' s disease. The aim of this study was to evaluate the efficacy of a combined therapy of anti-inflammatory Boswellia and antifibrotic Scutellaria extracts on the development of colonic fibrosis in rats. Methods: Chronic colonic inflammation-associated fibrosis was induced in rats by intracolonic administration of 2,4,5-trinitrobenzene sulphonic acid (TNBS). Sixty-four healthy male Sprague-Dawley rats were assigned to 5 groups: 8 controls, 14 TNBS, 14 TNBS orally treated with Boswellia extracts (50mg/kg/day), 14 TNBS orally treated with Scutelaria (150mg/kg/day), and 14 TNBS orally treated with both Boswellia (50mg/kg/day) and Scutellaria (150mg/kg/day) extracts.The colon was removed after 21 days of treatment and assessed by macroscopic, histological, morphometric and immunohistochemical (IHC) analyses. For IHC, alpha-smooth muscle actin (α-SMA), collagen types I-III, connective tissue growth factor (CTGF), transforming growth factor-beta1 (TGF-β1), Smad3, Smad7, and CD3 antibodies were used. Results: Combined oral administration of Boswellia and Scutellaria significantly improved the course and macroscopic findings of TNBS-induced chronic colitis assessed by disease activity index, colon weight, length, adhesions, strictures, dilatation, thickness, edema, ulcerations, and extension of damage. The histological severity of the colonic fibrosis was also notably improved by the treatment and associated with a significant reduction in the colonic expression of α-SMA, collagen I-III, CTGF, TGF-β1, Smad3, and Smad7. Conclusions: These data demonstrate that the prophylactic administration of anti-inflammatory Boswellia and antifibrotic Scutellaria extracts is effective in preventing colonic fibrosis in TNBS-induced colitis. Their antifibrotic mechanism of action seems to be mediated by the inhibition of TGFβ1/Smad3 pathway.

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P169 LEUKOCYTAPHERESIS FOR ULCERATIVE COLITIS - CORRELATION OF CLINICAL RESPONSE WITH IMMUNE REGULATION J. Cummings 1 , B.J. Singh 2 , S. Aryasingha 3, H. Uhlig 4 , P. Trzonkowski 2 , F. Powrie 5 , D.P. Jewell 1 . 1 University of Oxford, Oxford, United Kingdom; 2 Nuffield Department of Surgery, Oxford, United Kingdom; 3 John Radcliffe Hospital, Oxford, United Kingdom; 4 University Children' s Hospital Leipzig, Leipzig, Germany; 5 Sir William Dunn School of Pathology, Oxford, United Kingdom Introduction: Leukocytapheresis (LCAP) has been shown to be a safe and effective treatment for ulcerative colitis (UC). The mechanisms of action are unknown. A cellular pathway which may be affected by LCAP involves regulatory T cells (Treg). FOXP3+CD4+CD25+ T cells have been shown to be potent suppressors of immune responses. Aims and methods: The aim of this study was to assess the effect of leukocytapheresis on Treg cells and correlate the changes with clinical response. Patients who had not achieved remission (Colitis Activity Index (CAI) >4) after at least 4 weeks of >20mg of oral prednisolone or who had relapsed when the prednisolone dose was reduced to <10mg per day were eligible for the study. Inclusion criteria included a CAI of 5-12 and disease extent >15 cm. Twelve patients underwent 1 treatment session a week for 5 weeks. Response to treatment was assessed by CAI, ESR and CRP. Sigmoidoscopy and biopsy were performed at the beginning and end of the study and the endoscopic index (EI) was calculated. Outcome measures were compared using a paired t-test. Results are given as mean ± SEM. The frequency of CD4, CD8 and regulatory FOXP3+CD4+ T cells from PBMC was determined by flow cytometry on samples collected before and after the initial (week one) and final (week five) LCAP sessions. Immunohistochemistry for FOXP3+CD4+ was performed on snap-frozen rectal biopsies. Results: Two patients were withdrawn from the study due to worsening of their condition and underwent surgery. 11/12 patients were on immunomodulator therapy. The total activity scores (CAI+EI) improved on per protocol analysis (17.3±1.1 vs 13.4±1.3, P=0.01). LCAP was associated with a decrease in the absolute number of CD3+ T cells without a change in the ratio of CD4:CD8 T cells. There was a reduction in the proportion of CD19+cells. Over the 5 week period of treatment there was an increased proportion of naïve T cells (CD4+CD45RA+) 27.2% vs 29.9% p=0.3 and a reduction in the proportion of memory cells (CD4+CD45RO+) 46.1% vs 41.6% p=0.07. Successful outcome correlated with the proportion of FOXP3+CD4+CD28- Treg cells (r=0.66 p=0.018) and was inversely correlated with the proportion of a subset of memory cells (CD69+CD4+CD45RO+). There was no significant change seen in the number of FOXP3+ cells in the lamina propria nor was there a correlation with outcome. Conclusion: This pilot study provides evidence for the use of LCAP in refractory UC and suggests that response to therapy may be regulated through Treg cells.

P170 SUBCLINICAL INFLAMMATION, DISEASE RELAPSE AND RE-RESPONSE UPON DOSE INCREASE: AN ACCENT I SUBANALYSIS W. Reinisch 1 , G. Lang 2 , A. Fasanmade 2, A. Olson 2 , D. Esser 3 . 1 AKH Wien, Vienna, Austria; 2 Centocor R&D, Malvern, PA, United States; 3 Centocor BV, Leiden, The Netherlands Objective: We investigated whether ACCENT I patients clinically responding to scheduled infliximab therapy were more likely to experience a future disease relapse if they showed signs of persistent subclinical inflammation, using C-reactive protein (CRP) as a marker. We further investigated if response was re-established upon crossover to a higher dose. Methods: In ACCENT I, 573 patients with active CD received an infusion of 5mg/kg infliximab and were randomised into an episodic (Group 1) and two infliximab scheduled treatment groups. The scheduled treatment groups received further infusions of 5mg/kg infliximab at weeks 2 and 6 followed by 8-weekly infusions of 5mg/kg (Group 2) or 10mg/kg (Group 3). Patients who lost response after week 14 could cross over to treatment upon flare with 10mg/kg (Group 2) or 15mg/kg (Group 3) infliximab. We conducted a retrospective analysis of patients in Groups 2 and 3 (scheduled infliximab treatment) who responded at weeks 2 and 14, i.e. after an initial dose and a full induction regimen. Disease relapse at any timepoint after week 14 was analysed according to C-reactive protein (CRP) status at week 14 (high, ≥0.8mg/dL; low, <0.8mg/dL). Results: 157 patients randomised to scheduled infliximab treatment were in response at weeks 2 and 14. Patients with high CRP levels at week 14 were more likely to experience a future disease relapse than those with low levels (Group 2: 63% (10/16) vs. 49% (31/63); Group 3: 54% (15/28) vs. 24% (12/50); Overall: 57% (25/44) vs. 38% (43/113); p=0.0097 using a CochranMantel-Haenszel test stratified by treatment group). Of the 25 patients with