Su2073

Neonatal Colonic Irritation Produces Persistent Visceral Hypersensitivity andAltered Affective Behavior in a Mouse Model of Irritable Bowel Syndrome(IBS)Qian Li, Zachary Cordner, Liansheng Liu, Kellie L. Tamashiro, Timothy H. Moran, PankajJ. Pasricha

Introduction: A major theme in functional bowel disorders (FBD) such as irritable bowelsyndrome (IBS) is that psychological/psychiatric problems have a pathogenic role, basedboth on the observation that patients with FBD are more anxious and depressed than healthycontrol, as well as research linking stress and depression to altered gastrointestinal sensoryand motor function. However, recent experimental and epidemiological studies suggest thatprimary visceral disturbances can lead secondarily to persistent behavioral and emotionalabnormalities. The aims of the present study were to test the hypothesis that mild chemicalirritation of the colon of neonatal mice can produce changes in visceral hypersensitivity andpsychological behaviors that persist into adulthood. Methods: Ten-day old C57BL/6 mousepups received an intracolonic infusion of 20 μl of 0.5% acetic acid (IBS) or saline (control)to mimic a previously validated model of IBS in rats based on neonatal visceral irritation.Psychological behaviors and pain sensitivity of the colon were assessed when these micewere 8-14 weeks of age. Anxiety-like behaviors were tested using elevated plus maze andopen field tests. Depression-like behavior was tested by the forced swim test. Colonichypersensitivity to graded colorectal balloon distension (15, 30, 50 and 70mmHg) wasdetermined by visceromotor responses (VMR) measured by electromyography of externaloblique muscle. Resutls: Neonatal colorectal infusion of acetic acid but not saline significantlyincreased electromyographic activity in response to all of pressure colorectal distension inadults, suggesting the persistence of colonic hypersensitivity. In the elevated plus maze andopen field tests, IBS mice showed a significant reduction in time spent in the open arms ofelevated plus maze (control: 73.04±5.26s vs IBS: 53.54±6.53s, p<0.05) and in the centerof the open field (control: 73.99±6.42s vs IBS: 37.32±4.62s, p<0.05), suggesting an anxiety-like phenotype. IBS mice also showed a significant increase in immobility in the forcedswim test (control: 90.05±12.08s vs IBS: 140.43±20.06s, p<0.05), suggesting an increasein depression-like behavior. Conclusion: The present results demonstrate that transientcolonic irritation in the neonatal period can induce long-lasting depression-like and anxiety-like behaviors. These findings provide a new paradigm for the psychological co-morbidityin functional bowel disorders.

Su2074

The Effects of Adenosine A3 Receptor Agonist on Visceral Hypersensitivity ofNeonatal Maternally Separated Rat ModelsTianhua Ren, Yu Zhou

Background: The intestinal mucosa serotonin plays an important role in visceral hypersensi-tivity of irritable bowel syndrome (IBS). Adenosine A3 receptor modulates the release ofserotonin, but the role of adenosine A3 receptor in visceral hypersensitivity of IBS is notknown.Objective: To study the role of adenosine A3 receptor agonist in visceral hypersensitiv-ity of neonatal maternal separation IBS rat models. Methods: Neonatal maternally separatedSprague-Dawley male rats were used to establish visceral hypersensitivity IBD models. Afteradministration of adenosine A3 receptor agonist 2-Cl-IB-MECA, visceral pain response wasmeasured with behavior observation and abdominal withdrawl reflex scores. The expressionof c-fos protein in the lumbosacral spinal cord dorsal horn was detected using immunohisto-chemical methods. 5-HT positive cell number and 5-HT content in distal colon were quanti-tated using immunohistochemistry and ELISA respectively. Distal colon tissues were pro-cessed for routine histological assessment.Results: Compared to control group, 2-Cl-IB-MECA treated IBS rats showed significantly lower AWR scores in response to colorectaldistension (CRD), reduced number of c-fos protein immunoreactive neurons in the dorsalhorn of spinal cord, and reduced number of 5-HT positive cells and 5 - HT content indistal colon. No tissue damage or inflammation was found in the colons of both groupsbefore and after CRD. Conclusions: Adenosine A3 receptor activation may reduce colonic5-HT bioavailability and thereby attenuate visceral hypersensitivity of IBS rat models. Adeno-sine A3 receptor may be a new target for the treatment of visceral hypersensitivity of IBS.Table 1. The effect of 2-Cl-IB-MECA on visceral pain response of IBS rats

AWR scores were analyzed using Mann-Whitney U test.

S-539 AGA Abstracts

The immunoreactivity of c-fos in lumbosacral spinal dorsal horn and the 5-HT immunoreac-tive cells in colonic mucosa after colorectal distension

Su2075

Cognitive Functions and Depression in Patients With Irritable BowelSyndrome (IBS) - What Is the Connection?Per G. Farup, Knut Hestad

IBS is associated with depression and depression with impaired cognitive functions, butlittle is known about cognitive functions in patients with IBS. This study aimed at findingassociations between IBS, depression, and cognitive functions. Methods. A case-controlstudy. Patients with mild to severe depression ("depression" group) were compared withpatients with unspecified neurological symptoms ("neurological" group). Information aboutpatient characteristics, IBS (Rome III criteria) and depression (Montgomery-Aasberg depres-sion scale (MADRS) and Beck Depression Inventary (BDI-II)) were collected. Neuropsycho-logical tests were performed: Hopkin's Verbal Learning Test - R (HVLT-R, immediate verbalmemory), Trail Making A and B (simple and complex cognitive flexibility), Grooved Pegboard(eye-hand coordination and motor speed in dominant and not dominant hand), Brief VisualMemory Test Revised (BVMT-R, immediate visual memory) and Controlled Word AssociationTest (COWA, number of garments and animals). Comparisons were performed betweenthe two groups, within the groups and reported as mean (SD) and number (proportion).Multivariable analyses were performed. The study conforms to the principles of the Declara-tion of Helsinki and was approved by the Norwegian Regional Committees for Medical andHealth Research Ethics. Results. 48 (26 females) and 18 patients (11 females) with a meanage of 45.3 (14.3) and 46.1 years (13.9) (p=0.83) were included in the "depression" and"neurological" groups. The number of patients with IBS in the groups were 28 (58%) and5 (28%) (p=0.05) respectively. In the "depression" group, the only significant differencesbetween patients with and without IBS were MADRS scores; 29.3 (8.1) and 24.7 (7.3) (p=0.05) and BDI scores: 34.3 (11.9) and 24.6 (11.4) (p=0.007) respectively. In the "neurologicalgroup", no statistically significant differences were seen between patients with and withoutIBS. The table shows groups and IBS as predictors of depression and cognitive functionafter adjusting for age and gender (regression analyses). Only statistically significant associa-tions are shown. Conclusion. IBS was associated with depression but not with impairedcognitive function, whereas depression was associated with cognitive impairment. The find-ings may indicate that depression in patients with IBS differs from "true" depression withregard to comorbidity such as cognitive dysfunction.Groups and IBS as predictors of depression and cognitive function

Su2076

Bile Acid Diarrhoea Masquerades As Diarrhoea-Predominant Irritable BowelSyndrome: Results From a Dual Centre Prospective StudyImran Aziz, Saqib Mumtaz, Hassan Bholah, Fahmid U. Chowdhury, David S. Sanders,Alexander C. Ford

Background: Several studies have suggested that bile acid diarrhoea (BAD) can present withsymptoms that are compatible with diarrhoea-predominant irritable bowel syndrome (IBS-D). However, uncertainty exists as these studies have often been retrospective in nature,have not defined IBS-D according to accepted diagnostic criteria, or have included patients

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swith chronic diarrhoea in the analysis. We have therefore examined this issue in a well-characterised cohort of patients with rigorously defined IBS-D. Methods: This was a prospec-tive cross-sectional survey conducted among consecutive patients with IBS-D attendingGastroenterology clinics in two University hospitals in Sheffield and Leeds, UK. All patientscompleted questionnaires, and underwent 23-seleno-25-homo-tauro-cholic acid (SeHCAT)scanning according to local protocol, with a retention of <15% at day 7 used to confirmBAD. The degree of BAD was classed as severe if retention <5%, moderate if 5.0%-9.9%,and mild if 10.0%-14.9%. Presence of IBS-D was defined according to the Rome III criteria.Patients with other known risk factors for BAD, including previous cholecystectomy, terminalileal Crohn's disease, previous terminal ileal resection, previous pelvic or abdominal radiother-apy, coeliac disease, or microscopic colitis, were excluded. Participants also completed thepatient health questionnaire-15, a validated somatisation score, and the hospital anxiety anddepression score. Demographic data, including age, gender, lifestyle, and body mass index(BMI) were collected for all patients. The effect of all these patient characteristics on presenceor absence of BAD was examined by multivariate logistic regression analysis, with resultsexpressed as odds ratios (ORs) with 99% confidence intervals (CI). Results: This is an interimanalysis of an ongoing study. In total, 51 patients with IBS-D according to the Rome IIIcriteria have been recruited to date (37 (72.5%) female, mean age 47.0 years). In total, 14(27.5%) were found to have BAD following SeHCAT scanning. Of these, nine (17.6%) hadsevere BAD, four moderate, and one mild. Mean age, BMI, anxiety, depression, and somatisa-tion scores were not significantly different among those with, compared with those without,BAD. No predictors of presence of BAD were identified following multivariate logisticregression analysis. Conclusions: Our data suggest that more than one-in-four IBS-D patients,if investigated, will have definitive evidence of BAD. In the majority of patients this is severe.Failure to investigate patients to exclude BAD as an underlying cause of symptoms compatiblewith IBS-D results in misdiagnosis and a failure to institute effective therapy, in the formof bile acid sequestrants. This suggests that future IBS management guidelines should advocatediagnostic testing to exclude BAD before a diagnosis of IBS-D is made.

Su2077

Phasic and Tonic Motility Responses in the Colorectum After PeripheralAdministration of Corticotropin-Releasing Hormone Are Exaggerated inPatients With Irritable Bowel SyndromeMotoyori Kanazawa, Michiko Kano, Tomohiko Muratsubaki, Mao Yagihashi, Ayaka Sasaki,Yukari Tanaka, Joe Morishita, Shin Fukudo

Background and aims: Corticotropin-releasing hormone (CRH) signaling pathway in thegut may contribute to stress-related progression and exacerbation of irritable bowel syndrome(IBS) symptoms by modulating autonomic and enteric nervous systems as well as immunefunction. Using manometry, we previously reported that intravenous administration of CRHinduces colonic phasic contractions in both patients with IBS and healthy subjects and theseresponses are more prominent in IBS patients (Fukudo et al. Gut 1998). However, datashowing effect of CRH on colorectal tone and fine contractions were lacking. We tested thehypotheses that IBS patients show exaggeration of phasic and tonic motility responses inthe colorectum after exogenous CRH and that these responses are associated with IBSsymptoms. Methods: Subjects were 21 IBS patients with diarrhea (10 males, 11 females,mean age; 23 years) and 19 healthy controls (HC; 10 males, 9 females, 21 years). Colorectalmotility was recorded for 20 min as a baseline period and for 120 min after administrationof CRH (2 μg/kg) intravenously with a 600 ml capacity bag inflated at the individual operatingpressure (IOP) using an electrical barostat. Average barostat bag volumes and phasic volumeevents (PVEs) during the baseline and 0-60 min and 60-120 min periods after CRH injectionwere measured. An ordinate scale (0, none; 10, maximum) was used to assess IBS symptomsbefore administration of CRH and at the end of the study. Results: There was no significantdifference in IOP (8.8±1.5 vs. 9.6±1.6 mmHg, mean±SD), the average bag volume (131±59vs. 158±56 ml) or number of PVEs (6.3±9.2 vs. 4.9±7.8 times/h) during the baseline betweenIBS and HC. After administration of CRH, IBS patients showed less colorectal bag volume(higher smooth muscle tone; 123±61 vs. 160±50 ml for 0-60 min, p<0.05; 117±65 vs.169±55 ml for 60-120 min, p=0.01) and more phasic contractions (18.4±24.7 vs. 6.4±18.1times/h for 0-60 min, p=0.10; 18.6±17.6 vs. 7.8±11.6 times/h for 60-120 min, p<0.05)compared to HC. Changes in the bag volumes after CRH injection were also lower in IBSthan HC (93±16% vs. 103±12% for 0-60 min, p<0.05; 86±28% vs. 109±27% for 60-120min, p=0.01). In IBS patients, the baseline bag volume was negatively correlated with scoresfor abdominal pain (Rho=-0.44, p<0.05) and urge (Rho=-0.55, p=0.01) and the bag volume60-120 min after CRH injection was correlated with the urge score (Rho=-0.39, p=0.01).Conclusions: IBS patients are likely to show more increased tone and fine contractions inthe colorectum at events with peripheral CRH release. Moreover, increase in colorectalmuscle tone by CRH is associated with IBS symptoms. These findings support the idea thatmore increased peripheral CRH signaling pathway may induce exacerbation of IBS symptomsas well as abnormal central CRH system.

Su2078

Grey Matter Alterations in Medial Prefrontal Cortex Show NegativeAssociations With Subjective Reports of Worry in IBS PatientsJennifer S. Labus, Emeran A. Mayer, Aubrey D. Love, Jean Stains, Cathy Liu, Cody Ashe-McNalley, Laurie Keefer, Jeffrey M. Lackner, Bruce D. Naliboff, Kirsten Tillisch

Background. A hallmark of IBS patients is the presence of "worry", a state of anxiety oruncertainty about actual or potential problems. While this feature has been dismissed aspurely psychological, neurobiological mechanisms related to IBS pathophysiology may beinvolved. The medial prefrontal cortex (mPFC) plays a major role in emotion regulationand alterations in the structure, function and connectivity of mPFC with amygdala, insulaand nucleus accumbens have been implicated in the pathophysiology of anxiety disorders(Kim et al, 2011) and in chronic pain (Baliki et al, 2012). Aim. To determine if self-reportof "worry" is associated with structural alteration in bilateral middle frontal gyrus as indexedby volume, surface area, cortical thickness and mean curvature. Methods. Structural brainimages were obtained from 22 right handed female IBS patients (mean age = 40 y(SD=15)beginning a non-pharmacological treatment trial. All Rome III subtypes were represented

S-540AGA Abstracts

(Constipation=4, Diarrhea=8, Mixed= 9, and Unspecified=1). Segmentation and regionalparcellation was performed using Freesurfer on the UCLA Laboratory of Neuroimagingpipeline using the Destrieux atlas. Partial correlations controlling for age were used todetermine the association between the abbreviated Penn State Worry Questionnaire(PSWQ-A), a self-report measure of cognitions associated with anxiety, with morphometry of themiddle frontal sulci and gyri and the gyrus rectus (ventral medial prefrontal cortex, Brodmannarea 11). Significance was only considered after correcting for the number of tests (N=4)for each region and hemisphere using false discovery rate (q<.05). Results. Average scoreon the PSWQ-A was 28.14 (SD=6.67). Analyses indicated significant negative associationsbetween the volume of the right middle frontal gyrus, r(19)-.57, p=.008, q=.03, and theright gyrus rectus, r=-.55, p=.009, q=.03. Trends for an association with the PSWQ-A werealso seen in this small sample for the surface area of the right middle frontal gyrus, r= -.41,p=.07, surface area of the right gyrus rectus, r=-.45, p=.04, and mean curvature, an indexof gyrification, of the right and left middle frontal sulci, r=.40, p=.07 and r=.48, p=.03.Conclusions. These findings implicate a role of prefrontal brain regions (including themedial and ventromedial PFC) in the generation of worry symptoms in IBS patients, consistentwith an alteration in effectiveness of this brain region in corticolimbic inhibition. They areconsistent with previous reports in anxiety and other chronic pain disorders. The mechanismsunderlying these structural changes deserve further study.

Su2079

Decreased miR-199a/B Augments Visceral Pain Through Translational Up-Regulation of TRPV1 in IBS PatientsShehzad N. Merwat, G. Nicholas Verne, QiQi Zhou

Background: Irritable bowel syndrome (IBS) is a common gastrointestinal disorder in whichthere are few effective therapeutic agents. Worse, many IBS patients not only have abdominalpain, but also exhibit extra-intestinal somatic symptoms (e.g., back pain, migraine headaches,muscle pain) suggesting central hyperalgesic dysfunction. The mechanisms underlying thepathophysiology of pain in IBS are poorly understood. The objective of this study wasto evaluate the expression of transient-receptor-potential-vanilloid-type-1 (TRPV1) and itscomplementary miRNA in colonic tissues of diarrhea- predominant IBS (IBS-D) patientswith abdominal pain. Methods: We evaluated 45 IBS-D patients using the FBDSI and visceralpain severity score and 40 controls. A miRNA array was performed and miRNA expressionwas evaluated in human colonic tissue. To further confirm the relationship between miR-199a/b and TRPV1 expression, a Luciferase assay was performed. RESULTS: Significantlydecreased miR-199a/b expression was present in colon tissues from IBS-D patients comparedto controls that inversely correlated with FBDSI and visceral pain severity scores. Thesesame IBS-D patients had increased expression of TRPV1. Visceral pain and visceral hypersensi-tivity was modulated by miR199a/b which has a complementary site in the 3'-UTR of theTRPV1 gene. Conclusion: Visceral pain and visceral hypersensitivity in IBS-D patients mayresult from decreased miR-199a/b expression that leads to increased numbers of polymodalsensory nerve fibers expressing TRPV1. These results support the conclusion that miR-199a/b regulates abdominal pain and visceral hypersensitivity in IBS-D patients via modulationof the TRPV1 signaling pathway.

Su2080

Global DNA Methylation Analysis in Irritable Bowel SyndromeSwapna M. Joshi, Christos Polytarchou, Dimitrios Iliopoulos, Joe DeYoung, Emeran A.Mayer, Lin Chang

Background and Aims: Despite a growing consensus about the role of altered brain gutinteractions in the pathophysiology of irritable bowel syndrome (IBS), there is little under-standing about underlying molecular mechanisms. Complex interactions between host andenvironment can influence gene expression mediated by epigenetic mechanisms, such asDNA methylation. In an exploratory analysis, we aimed to identify differentially methylatedgenes between IBS patients and healthy controls (HCs) and also between various IBS bowelhabit subtypes. Methods: We compared global DNA methylation profiles of peripheral bloodmononuclear cells from 12 IBS patients (mean[±sd] age=39.8±3.4 yrs, 58 % F, 25% IBS-Constipation [IBS-C], 50% IBS-Diarrhea [IBS-D], 25% Mixed [IBS-M]) with 12 HCs(mean[±sd] age=39.8±3.8 yrs, 58% F), using Illumina HM450 array, which interrogatesDNA methylation status of > 450,000 CpG sites and > 99% of all genes. DNA methylationvalues were compared between IBS and HCs, and between various bowel habits usingWilcoxon rank-sum test. Multiple testing corrections were not applied in the initial analysis,due to limited power; however, we assumed 5% genes to be significant by chance. Geneontology (GO) signature was analyzed to identify the common function term associatedwith a gene list, using DAVID bioinformatics tool (http://david.abcc.ncifcrf.gov/). Results:Using the criteria of a mean methylation difference ≥15% and p<0.05, 29 probes weredifferent between IBS and HCs, 81 probes were different between IBS-C vs. IBS-D, 39between IBS-C vs. HCs, and 26 between IBS-D vs. HCs (total of 175 probes for 128 genes).Six genes corresponding to 10 probes (Table 1) with the lowest p values and highest DNAmethylation group differences were Synphilin-1 (SNCAIP), SCO-spondin (SSPO), RINGfinger protein 39 (RNF39) and Tubulin Polymerization Promoting (TPPP), glutathione Stransferases mu class (GSTM1, GSTM5). All 10 probes were hypermethylated in IBS. Fourout of 6 genes, SNCAIP, SSPO, RNF39 and TPP3, were associated with neuronal function.GSTM enzymes play a role in the detoxification of environmental toxins and products ofoxidative stress. GO analysis of 128 significant genes showed a significant enrichment ofGO term "neuropeptide hormone activity" (defined as "any peptide hormone that acts inthe central nervous system") (p=0.004, FDR=5.4). Conclusions: Although there were feweroverall differences in DNA methylation between IBS patients and HCs, there were markeddifferences by bowel habit subtypes. These findings warrant replication in a larger cohortand assessment of expression levels, however, our data suggests that epigenetic modificationsplay a role in IBS, and that IBS-C and IBS-D have different pathophysiologic mechanismsthat involve oxidative stress and neuropeptide hormonal activity within the brain-gut axis.Table1: Location and DNA methylation values of 10 selected probes