Search

Summary Background Patients with inflammatory bowel disease (IBD) are at higher risk for Clostridium difficile infection (CDI). Disruption of gut microbiome and interaction with the intestinal immune system are essential mechanisms for pathogenesis of both CDI and IBD. Whether genetic polymorphisms associated with susceptibility to IBD are also associated with risk of CDI is unknown. Aims To use a well-characterised and genotyped cohort of patients with UC to (i) identify clinical risk factors for CDI; (ii) examine if any of the IBD genetic risk loci were associated with CDI; and (iii) to compare the performance of predictive models using clinical and genetic risk factors in determining risk of CDI. Methods We used a prospective registry of patients from a tertiary referral hospital. Medical record review was performed to identify all ulcerative colitis (UC) patients within the registry with a history of CDI. All patients were genotyped on the Immunochip. We examined the association between the 163 risk loci for IBD and risk of CDI using a dominant genetic model. Model performance was examined using receiver operating characteristics curves. Results The study included 319 patients of whom 29 developed CDI (9%). Female gender and pancolitis were associated with increased risk, while use of anti-TNF was protective against CDI. Six genetic polymorphisms including those at TNFRSF14 [Odds ratio (OR) 6.0, P-value 0.01] were associated with increased risk while 2 loci were inversely associated. On multivariate analysis, none of the clinical parameters retained significance after adjusting for genetics. Presence of at least one high-risk locus was associated with an increase in risk for CDI (20% vs. 1%) (P = 6 × 10−9). Compared to 11% for a clinical model, the genetic loci explained 28% of the variance in CDI risk and had a greater AUROC. Conclusion Host genetics may influence susceptibility to Clostridium difficile infection in patients with ulcerative colitis.

Sleep disruption is common in inflammatory bowel diseases (IBD). However, studies demonstrating a similar prevalence in irritable bowel syndrome suggest that nighttime disruption due to diarrhea and abdominal pain may be key drivers of poor sleep quality. Whether inflammation is associated with poor sleep independently has not been examined previously.This single-center study included subjects with IBD recruited to an ongoing prospective registry who completed a questionnaire assessing sleep quality and mood. Inflammatory marker levels [C-reactive protein (CRP), erythrocyte sedimentation rate] and clinical disease activity including nighttime disruption on the day of enrollment were obtained from the medical record. Logistic regression models were used to identify predictors of sleep quality.The study included 131 subjects (72 women) with a median age of IBD diagnosis of 25 years. Twenty-three subjects (19 %) had a high C-reactive protein level (≥8 mg/dL). Poor sleep was more common in those with high CRP levels than with normal values (70 vs. 39 %, p = 0.009). This association remained significant on multivariate analysis [Odds ratio (OR) 4.12, 95 % confidence interval (CI) 1.38-12.29]. Adjusting for the presence of nighttime disruption did not significant alter this association (OR 3.16, 95 % CI 1.01-9.90). High CRP correlated with poor sleep even in patients not experiencing nocturnal symptoms (n = 101, OR 4.89, 95 % CI 1.24-19.36).High CRP is associated with poor sleep quality in IBD independent of the presence of nighttime disruptions, suggesting that a relationship exists between circulating inflammatory markers and sleep.

Crohn's disease (CD) and ulcerative colitis (UC) are chronic immunologically mediated diseases with a progressive relapsing remitting course. There is considerable heterogeneity in disease course and accurate prediction of natural history has been challenging. The phenotypic implication of increasing genetic predisposition to CD or UC is unknown.The data source for our study was a prospective cohort of CD and UC patients recruited from a tertiary referral center. All patients underwent genotyping on the Illumina Immunochip. A genetic risk score (GRS) incorporating strength of association (log odds ratio) and allele dose for each of the 163 inflammatory bowel disease (IBD) risk loci was calculated and phenotypic associations examined across GRS quartiles.Our study cohort included 1,105 patients (697 CD, 408 UC). Increasing genetic burden was associated with earlier age of diagnosis of CD (Ptrend=0.008). Patients in the highest GRS quartile were likely to develop disease 5 years earlier than those in the lowest quartile. Increasing genetic burden was also associated with ileal involvement in CD (Ptrend0.0001). The effect of genetic burden was independent of the NOD2 locus and was stronger among those with no NOD2 variants, and in never smokers. UC patients with an involved first-degree relative had a higher genetic burden, but GRS was not associated with disease phenotype in UC.Increasing genetic burden is associated with early age of diagnosis in CD, but not UC. The expanded panel of IBD risk loci explains only a fraction of variance of disease phenotype, suggesting limited clinical utility of genetics in predicting natural history.

Inflammatory bowel disease (IBD) and celiac disease are the two most common immune-mediated gastrointestinal diseases. There is limited knowledge regarding the course of IBD in those with coexisting celiac disease. We conducted this study to determine whether patients with coexisting celiac disease present a unique phenotype of IBD and to examine the frequency of co-occurrence of celiac disease and IBD in comparison with other autoimmune disorders.This was a case-control study performed at two tertiary referral centers. Cases comprised of patients with known diagnoses of celiac disease and IBD. Two random IBD controls without celiac disease were selected for each case after matching for IBD type. Disease phenotype and natural history for both Crohn's disease (CD) and ulcerative colitis (UC) were noted from medical record review, and were compared between IBD patients with and without celiac disease.We identified a total of 51 patients with IBD (22 UC, 1 indeterminate colitis, 28 CD) and celiac disease. There was no significant difference in the age, gender, or ethnicity between celiac-IBD and controls. Pancolitis was more common in celiac-UC patients as compared with controls (odds ratio (OR) 3.30, 95% confidence interval (CI) 1.05-21.50). There was also a trend toward increased use of immunomodulators (IMMs) among celiac-UC patients than in non-celiac UC controls (OR 2.83, 95% CI 0.95-8.48). No phenotypic differences were found in celiac-CD patients. There were no significant differences in IBD-related medication usage, hospitalizations, or surgeries.Patients with UC and celiac disease were more likely to have pancolitis and had a trend toward greater use of IMMs. Coexisting celiac disease did not influence natural history of CD.

Background & Aims Immunodeficiency and autoimmune sequelae, including colitis, develop in patients and mice deficient in Wiskott–Aldrich syndrome protein (WASP), a hematopoietic cell-specific intracellular signaling molecule that regulates the actin cytoskeleton. Development of colitis in WASP-deficient mice requires lymphocytes; transfer of T cells is sufficient to induce colitis in immunodeficient mice. We investigated the interactions between innate and adaptive immune cells in mucosal regulation during development of T cell–mediated colitis in mice with WASP-deficient cells of the innate immune system. Methods Naive and/or regulatory CD4 + T cells were transferred from 129 SvEv mice into RAG-2–deficient (RAG-2 KO) mice or mice lacking WASP and RAG-2 (WRDKO). Animals were observed for the development of colitis; effector and regulatory functions of innate immune and T cells were analyzed with in vivo and in vitro assays. Results Transfer of unfractionated CD4 + T cells induced severe colitis in WRDKO, but not RAG-2 KO, mice. Naive wild-type T cells had higher levels of effector activity and regulatory T cells had reduced suppressive function when transferred into WRDKO mice compared with RAG-2 KO mice. Regulatory T-cell proliferation, generation, and maintenance of FoxP3 expression were reduced in WRDKO recipients and associated with reduced numbers of CD103 + tolerogenic dendritic cells and levels of interleukin-10. Administration of interleukin-10 prevented induction of colitis following transfer of T cells into WRDKO mice. Conclusions Defective interactions between WASP-deficient innate immune cells and normal T cells disrupt mucosal regulation, potentially by altering the functions of tolerogenic dendritic cells, production of interleukin-10, and homeostasis of regulatory T cells.

Background & Aims: Controversy exists as to whether patients with inflammatory bowel disease have an underlying immunodeficiency. We have focused on a murine model of the Wiskott-Aldrich syndrome, an immunodeficiency in which autoimmunity can manifest in the form of an inflammatory bowel disease-like illness. Wiskott-Aldrich syndrome protein (WASP) deficiency in mice results in similar clinical features. Herein, we characterized the colitis in WASP-deficient mice. Methods: WASP-deficient mice were followed clinically and histologically. Immunologic studies were performed to determine the pathogenic cell population(s), the predominant cytokine expression pattern, and the role of cytokine(s) in colitis pathogenesis. Results: All WASP-deficient mice develop colitis by 6 months of age. Lymphocytes are required for disease induction, and CD4+ T cells from WASP-deficient mice are sufficient to induce disease in lymphocyte-deficient hosts. Lamina propria preparations from WASP-deficient mice demonstrated elevations in interferon-γ, interleukin (IL)-4, and IL-13 levels but decreased IL-6 and no difference in IL-17 expression in comparison with wild-type controls. Treatment with neutralizing antibody to IL-4, but not to interferon-γ, abrogated colitis development. However, mice deficient in both WASP and IL-4 showed no difference in histologic colitis scores at 24 weeks of age compared with WASP-deficient mice. Conclusions: These results demonstrate a critical role for lymphocytes and a relative T helper 2 cytokine predominance in the colitis associated with WASP-deficient mice. This is the only model of colitis with elevated T helper 2 cytokines and aberrant natural regulatory T cell function and is unique in having a human disease counterpart with similar defects.

CEACAM6 gene promoter in an open chromatin state region characterized by increased H3S10 phosphorylation level. In contrast, Caco-2 cells expressed low levels of CEACAM6 due to a compact chromatin state in CEACAM6 promoter (low level of H3S10p). AIEC infection led to an increase in CEACAM6 expression related to enhanced HIF-1a binding to CEACAM6 promoter. Abnormal H3S10 phosphorylation in CEACAM6 promoter following AIEC infection enhanced HIF-1a binding and subsequent CEACAM6 expression. Moreover, in silico analysis predicted several miRNAs that are potentially able to regulate CEACAM6 expression. Luciferase reporter assays showed that among the predictive miRNAs, miR-765 and miR-29b-2-5p directly bound to the 3'-UTR of CEACAM6 mRNA. After AIEC infection, a down-regulation of miR-765 and an up-regulation of miR-29b-2-5p in T84 cells were observed. This is consistent with the observation that miR-765 decreased CEACAM6 protein expression compared to miR-29b-2-5p after transfection in T84 cells. Conclusion: This study suggests that AIEC bacteria have the ability to modulate gene expression in the host cell for their own benefit by altering different epigenetic marks as miRNA expression profile and histones post-translational modifications enhancing CEACAM6 abnormal expression. Thus, epigenetic pathways could be new targets to prevent AIEC colonization and AIECinduced inflammation in CD patients.

dl among 84 CD cases (Pdiff=.005) and 20.0 ng/dl among 91 UC cases (Pdiff=.70). Compared to women in the lowest quartile of testosterone the multivariate-adjusted RRs for CD were 0.76 (95% CI, 0.38-1.54) for women in the second quartile; 0.45 (95% CI, 0.20-0.99) for the third quartile; and 0.51 (0.24-1.10) for the highest quartile (Plinear trend = .04). We observed that for every 5 ng/dl increase in testosterone, the risk of CD decreased by 14% (Plinear trend = .03). In contrast, we did not observe any consistent association between prediagnostic testosterone and risk of UC (Plinear trend=.75). We also did not observe any association between plasma levels of SHBG and DHEAS and risk of UC or CD (all Plinear trends > .15). Conclusion: Pre-diagnosis levels of circulating total testosterone are associated with a lower risk of CD but not UC among women. The effect of testosterone in modulating innate immunity may explain this novel finding. In addition, because oral contraceptives decrease levels of endogenous testosterone, our data provides insight into the mechanism by which exogenous hormones may influence risk of CD.

with reduced TNF release after TLR2 activation. Conclusions: In conclusion, we have developed a successful strategy to identify abnormal expression of genes in individual CD patients relevant to diminished macrophage function. In a wider context, this approach may be equally applicable in delineating the molecular pathogenesis of other complex disorders. 1. A. Franke et al., Nat. Genet. 42, 1118 (2010) 2. A. M. Smith et al., J. Exp. Med. 206, 1883 (2009)

but not in CD8αα+ IEL or LPL. Chromatin immunoprecipitation (CHIP) analysis from splenic CD8αβ T cells from TNF△ARE/+ but not wild type mice revealed selective recruitment of ATF6 and XBP1 to the Grp78 promoter. Consistent with the cytotoxic effect of TNF△ARE/ + derived CD8αβ+ T cells on the small intestinal epithelial cell line Mode-K, CHIP analysis revealed selective binding of ATF4, ATF6 and XBP1 to the granzyme B promoter in CD8αβ+ IEL from TNF△ARE/+ but not wild type mice. Finally, Grp78 knock down through the use of siRNA or the bacterial toxin EGF-SubA fusion protein completely abolished IFNγ and granzyme B expression. In conclusion, these results provide first evidence that ER stress response-associated mechanisms contribute to the cytotoxic effector function of CD8αβ+ IEL under condition of chronic ileitis in TNF△ARE/+ mice.

Background & Aims The commensal microbiota is believed to have an important role in regulating immune responsiveness and preventing intestinal inflammation. Intestinal microbes produce signals that regulate inflammation via Toll-like receptor (TLR) signaling, but the mechanisms of this process are poorly understood. We investigated the role of the anti-inflammatory cytokine interleukin (IL)-10 in this signaling pathway using a mouse model of colitis. Methods Clinical, histopathologic, and functional parameters of intestinal inflammation were evaluated in TLR4 −/− , IL-10 −/− , and TLR4 −/− × IL-10 −/− mice that were free of specific pathogens and in TLR4 −/− × IL-10 −/− mice following eradication and reintroduction of Helicobacter hepaticus. Regulatory T-cell (Treg) function was evaluated by crossing each of the lines with transgenic mice that express green fluorescent protein under control of the endogenous regulatory elements of Foxp3 . Apoptotic cells in the colonic lamina propria were detected by a TUNEL assay. Results TLR4-mediated signals have 2 interrelated roles in promoting inflammation in TLR4 −/− × IL-10 −/− mice. In the absence of TLR4-mediated signals, secretion of proinflammatory and immunoregulatory cytokines is dysregulated. Tregs (Foxp3 + ) that secrete interferon-γ and IL-17 accumulate in the colonic lamina propria of TLR4 −/− × IL-10 −/− mice and do not prevent inflammation. Aberrant control of epithelial cell turnover results in the persistence of antigen-presenting cells that contain apoptotic epithelial fragments in the colonic lamina propria of Helicobacter- infected TLR4 −/− mice. Conclusions In mice that lack both IL-10- and TLR4-mediated signals, aberrant regulatory T-cell function and dysregulated control of epithelial homeostasis combine to exacerbate intestinal inflammation.

Background & Aims: Microvillus inclusion disease is a rare and fatal congenital enteropathy, presenting with intractable secretory diarrhea shortly after birth. The complete inability to absorb nutrients from intestinal lumen demands total parenteral nutrition, and, eventually, transplantation of the small intestine. MID characteristics varies among patients and generally comprises of villous atrophy and crypt hyperplasia, and, at the cellular level, by the apical brush border atrophy, accumulation of apical proteins, lysosomes and microvilli-like inclusions in the apical cytoplasm of intestinal absorptive cells. Previously we have shown that MID enterocytes display abnormal expression of apical recycling endosomal markers, i.e. Rab11a, FIP-1 (RCP), FIP-5 (Rip11), resulting in a defective apical recycling system in MID. In this study, we aimed to identify the genetic cause and functional consequences that underlie the microvillus inclusion disease. Methods: We screened the genomic DNA of three patients diagnosed with microvillus inclusion disease, their siblings and parents. Biopsies of small intestine from MID and control patients were used to analyze the organization of organelles and localization of proteins involved in intracellular trafficking of brush border proteins. Results: In all MID patients together we have identified two substitutions, one deletion, and two protein truncating mutations in the myosin 5B gene. The MYO5B encodes for an actin filament-binding molecular motor protein that interacts with the small GTPase Rab11a, a marker of recycling endosomes, and thus facilitates the intracellular trafficking of apical proteins towards the apical membrane. We also found aberrant expression and subcellular distribution of myosin Vb protein and other key proteins that interact withmyosin Vb and/or control apical recycling endosome-mediated protein trafficking. Conclusions: The endosomal system that ensures the recycling of brush border proteins, with myosin Vb as a critical regulator, is required to develop and maintain functional apical cell surface in human enterocytes, and perturbations in this can be causally linked to microvillus inclusion disease. Mutations occurring at different positions of MYO5B gene and thus affecting different functional regions of MYO5B protein could explain the diversity of phenotypes present in MID patients. The identification of mutations in MYO5B as the cause for MID brings a major advance in setting the reliable diagnosis, enables the genetic counseling and prenatal screening, as well as paves the way for developing alternative therapeutic strategies.

3 d). TNBS caused an increase c-fos positive neurons (37.2±5.1 vs 16.8±1.5; p