the final CAT activity. The increased transcriptor factor STAT1 highlights the importanceof IF- γ in CD and its correlation with OxS regulation. The increased PSKH1 identifies thetrafficking and processing of pre-mRNA as a target for CD pathogenia. 1) Beltrán B, et al.Inflamm Bowel Dis. 2009 Jul 27 (ahead of print).

[mRNA]as aM

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Genes Associated With Reduced Epithelial Permeability and Epithelial-Mesenchymal Transition: Changes in Expression Following InfliximabTherapy in Ulcerative ColitisGary Toedter, Keying Ma, Katherine Li, Colleen W. Marano, Michael Macoritto, JenniferPark, Renee Deehan Kenney, Andrea Matthews, Gary D. Wu, James D. Lewis, Paul J.Rutgeerts, Frederic Baribaud

Background:Changes in intestinal permeability have been implicated in the pathology ofulcerative colitis (UC). Objectives: To investigate mechanismsmodulating intestinal permeab-ility and epithelial-mesenchymal transition (EMT) following infliximab (IFX)treatment, weanalyzed mRNA expression in colonic biopsies obtained from UC patients(pts), normalhealthy subjects, and UC pts enrolled in the ACT1 study. ACT1 evaluated safety and efficacyof IFX in pts with UC. Methods: Colonic biopsies (n=17 UC anti-TNFα naïve; n=8 non-IBD normal; n=113 from ACT1) were analyzed for mRNA expression using AffymetrixGeneChip Human Genome U133 Plus 2.0 arrays followed by quantitative polymerase chainreaction (qPCR) confirmation of the differentially expressed genes. Causal network modelingwas utilized to generate models describing the biological signaling pathways that drivechanges observed in the mRNA expression data. Results: mRNA expression results fromanti-TNFα naïve UC and normal samples analyzed using network modeling determinedthat genes known to affect intestinal permeability were deregulated in UC pts. Analysis ofthe ACT1 sample expression data demonstrated that treatment with IFX restored geneexpression toward, but not completely to, normal in IFX-responder pts. Pts in ACT1 whowere non-responders or placebo treated (including those diagnosed as clinical responders)did not restore these functions. qPCR analysis of the identified genes (Table) confirmed thechanges in gene expression observed in IFX treated pts from the ACT1 study. Conclusion:Using the combination of expression analysis and network modeling, we found that respond-ers to IFX restore mRNA expression levels in genes involved in intestinal epithelial permeabil-ity and fibrotic processes leading to EMT to levels comparable with those seen in normalsubjects. IFX non-responders and placebo treated pts have no equivalent restoration in geneexpression involved in these functions.

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P-Glycoprotein (P-GP) Functional Activity in Peripheral Blood Lymphocytes(PBL) and Colonic Biopsies of Ulcerative Colitis (UC)Alicia M. Sambuelli, Catalina M. Cortada, Maricel I. Bellicoso, Valeria Cismondi, AnibalH. Gil, Silvia M. Negreira, Sergio P. Huernos, Pablo R. Tirado, Silvina A. Goncalves, AnaM. Cabanne, Marta A. Carballo

BACKGROUND: The multidrug resistant gene (MDR1) encodes the P-gp, a transmembraneefflux pump, over-expressed in resistant cancer-cells, decreasing intracellular drug concentra-tion. Regarding IBD, MDR maps in a susceptibility genome site and P-gp is expressed inapical intestinal epithelial cells (EC) and immune cells (PBL, mucosal lymphocytes). P-gphyperfunction was a factor proposed for IBD refractoriness, but a deficient function to pump-out harmful toxins was postulated for pathogenesis. AIM: to study P-gp activity in UC fromPBL and colonic biopsies. MATERIALANDMETHODS: P-gp functional activity was evaluatedin PBL (with cell-populations immunomarcation) of UC (n 49) and healthy controls (HC,n 78) with simultaneous assessment in intraepithelial lymphocytes (IEL) and epithelial cells(EC), isolated from colonoscopic biopsies (UC 18, HC 11). Rhodamine123 (a fluorescentP-gp substrate) efflux was studied by flow cytometry, in absence and presence of P-gpmodulators Verapamil 100 μM, Valspodar 1μM. Data were expressed by the behaviour oftwo markers defined by % of cells with different fluorescence levels: M1 (high fluorescence,

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low P-gp pump activity) and M2 (low fluorescence, high activity). Patients were categorizedin responders (RESP: n 26) or refractory (REFR: n 23) to current treatment (mesalazine,steroids, 6-MP) and activity was scored (DAI). RESULTS (mean±SD): Irrespective of kindof current treatment, significant differences were observed in PBL, showing increased P-gpfunctional activity in REFR vs RESP (M2: 60.0±15.9 vs 47.4±10.7 p<0.002, Median test)and HC (47.9±12.3 p=0.009), but not between RESP vs HC. Reassessment in respondersshowed P-gp reduction. Interestingly, an UC subset assessed before any treatment (n 10)showed increased activity vs HC (M2: 54.2±5.8 p<0.04) and RESP (p<0.02). In the inhibitionassay mean M2 was higher in REFR (26.6±22.6) vs RESP (9.1±5.6 p<0.002) and HC(12.6±8.0 p<0.001). In spite of mild global DAI correlation, in severe pts (DAI 9-12, n12)surgery was required in 3/4 cases showing M2>M1 in the inhibition assay. Results forbiopsies were: IEL T cells showed decreased P-gp activity vs HC (only basal M2: 30.3±22.4vs. 53.6±22.4, p<0.02). PBL and IEC (T cells) did not showed correlation in UC or HC. P-gp in EC was higher than HC (basal: 1096±35 vs 1515±51 p<0.01, inhibition: 1015±37 vs1794±50 p<0.002). CONCLUSIONS: We found significant P-gp activity differences betweenUC and HC. PBL and IEC did not show correlation. Because of P-gp is able for modulation,its research in UC, could improve disease management and pathogenesis knowledge, eitherif the behaviour is cause or consequence of the disease.

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CTLA4 -1661a/G and 3'Utr Long Repeat Polymorphisms are Associated WithUlcerative Colitis, Regulate CTLA4 mRNA Stability and Influence mRNA andProtein ExpressionsZhitao Chen, Steven R. Brant, Umid K. Shrestha, Ting Jiang, Feng Zhou, Bing Xia

Background and aims: Reduced cytotoxic T-lymphocyte antigen 4 (CTLA4) expression hasbeen proposed as a risk for autoimmunity. CTLA4 polymorphisms have been associatedwith autoimmune diseases. Our aim was to evaluate the association of CTLA4 polymorphismswith Chinese ulcerative colitis (UC), the polymorphisms' effects on mRNA stability and fulllength (flCTLA4) and soluble CTLA4 (sCTLA4) expressions. Methods: Three hundredChinese new patients with UC and 700 age and sex matched healthy controls were genotypedfor -1661A/G, -1722T/C and 3'UTR (AT)n repeats by PCR-based methods combined DNAsequencing. The phenotypes of Chinese UC patients were grouped according to diseaseextent and extra-intestinalmanifestations. flCTLA4 and sCTLA4mRNA expressions in colonicbiopsies and mRNA stability in peripheral blood mononuclear cells of UC patients weremeasured by quantitative PCR and half-life, respectively. The protein expressions of flCTLA4in colonic biopsies and sCTLA4 in sera of UC patients were determined by immunohisto-chemistry and enzyme-linked immunosorbent assay, respectively. Results: The frequency ofG carrier of CTLA4 -1661A/G was higher in UC patients than in healthy controls (30.7%vs. 22.0%, P = 0.004, OR = 1.57, 95% CI: 1.16-2.13). The prevalence of (AT)n repeats ofCTLA4 gene carrying long alleles (≥ 116 bp) was more common in UC patients than inhealthy controls (22.0% vs. 6.3%, P < 0.001, OR = 4.21, 95% CI: 2.79-6.33), and associatedwith extensive colitis (P = 0.008). Among UC patients, sCTLA4 mRNA expressions weredecreased in active disease compared to non-active disease (P = 0.004), long allele carriersexpressed lower levels of flCTLA4 and sCTLA4 mRNA and sCTLA4 protein in active diseasethan short allele carriers (P < 0.001; P < 0.001; P < 0.001, respectively), and mRNA withlong (AT)n repeat alleles has shorter half-life than mRNA with short alleles and , hence,are more unstable. There were no significant associations between the CTLA4 -1722T/Cpolymorphism and UC patients. Conclusions: CTLA4 gene -1661A/G and long 3'UTR (AT)nrepeat polymorphisms are associated with UC in central China. The (AT)n repeat polymorph-ism in the 3'UTR contributes to decreased mRNA stability and, hence, to reduced expressionof CTLA4. Our study suggests that CTLA4 plays an important role in susceptibility for UCin central China. Acknowledgements The project is supported by grants from the NaturalScience Foundation of China (30871149, 30470783) and from the Funding for publicwelfare of the Ministry of Public Health of China (200802156). Dr. Brant received supportfor this collaboration from the Sherlock Hibbs scholarship.

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A Highly Significant Association of Tumor Necrosis Factor SuperfamilyMember 15 Gene With Inflammatory Bowel Disease in ChinaFeng Zhou, Ting Jiang, Liuqing Ge, Zhitao Chen, Jie Zhao, Bing Xia

Background and aims: Tumor necrosis factor superfamily member 15 (TNFSF15) has beenpreviously identified as a susceptibility gene for inflammatory bowel disease (IBD) in Japaneseand European cohorts and confirmed existing a strong ethnic variation. The aim of thisstudy was to investigate an association between TNFSF15 and IBD in Chinese population.Methods: A total of 464 IBD patients including 320 UC and 144 CD and age and sexmatched 500 healthy controls were genotyped by matrix-assisted laser desorption/ionizationtime-of-flight mass spectrometry (MALDI-TOF MS) for six common single nucleotide poly-morphisms (SNP), which define the main TNFSF15 haplotypes. The association betweenthe TNFSF15 haplotypes and UC or CD was analyzed by Haploview 4.1 software. Results:Distribution of TNFSF15 haplotypes was shown in Table. We defined new haplotypes 1 to4, and found that the haplotype 3 was a risk for UC (P=0.013, OR=1.365, 95%CI 1.068-1.745) and the haplotype 1 was protective for CD (P=0.012, OR=0.707, 95%CI 0.539-0.926). Our results have shown that previously classified haplotype A was associated withUC (P=0.001, OR=1.374, 95%CI 1.125-1.50) and CD (P=0.007, OR=1.435, 95%CI 1.102-1.868), but the “protective” haplotype B was fail to show those associations. Conclusion:Our study has shown further evidence that the genetic variations of the TNFSF15 genecontribute to the susceptibility of IBD in the Chinese population. Acknowledgement Thestudy was supported by grants from the ministry of public health welfare project ofChina (200802156)Association studies of TNFSF15 haplotypes between cases and controls in the Chinese popula-tion

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