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Background Inflammatory bowel disease (IBD), is a debilitating group of chronic diseases including Crohn’s Disease (CD) and ulcerative colitis (UC), which causes inflammation of the gut and affects millions of people worldwide. At different taxonomic levels, the structure of the gut microbiota is significantly altered in IBD patients compared to that of healthy individuals. However, it is unclear how these IBD-affected bacterial groups are related to other common bacteria in the gut, and how they are connected across different disease conditions at the global scale. Results In this study, using faecal samples from patients with IBD, we show through diversity analysis of the microbial community structure based on the 16S rRNA gene that the gut microbiome of IBD patients is less diverse compared to healthy individuals. Furthermore, we have identified which bacterial groups change in abundance in both CD and UC compared to healthy controls. A substantial imbalance was observed across four major bacterial phyla including Firmicutes, Bacteroidetes, Proteobacteria and Actinobacteria, which together constitute > 98% of the gut microbiota. Next, we reconstructed a bacterial family co-abundance network based on the correlation of abundance profiles obtained from the public gut microbiome data of > 22,000 samples of faecal and gut biopsies taken from both diseased and healthy individuals. The data was compiled using the EBI metagenomics database (Mitchell et al. in Nucleic Acids Res 46:D726–D735, 2018). By mapping IBD-altered bacterial families to the network, we show that the bacterial families which exhibit an increased abundance in IBD conditions are not well connected to other groups, implying that these families generally do not coexist together with common gut organisms. Whereas, the bacterial families whose abundance is reduced or did not change in IBD conditions compared to healthy conditions are very well connected to other bacterial groups, suggesting they are highly important groups of bacteria in the gut that can coexist with other bacteria across a range of conditions. Conclusions IBD patients exhibited a less diverse gut microbiome compared to healthy individuals. Bacterial groups which changed in IBD patients were found to be groups which do not co-exist well with common commensal gut bacteria, whereas bacterial groups which did not change in patients with IBD were found to commonly co-exist with commensal gut microbiota. This gives a potential insight into the dynamics of the gut microbiota in patients with IBD.

BACKGROUND AND AIMS: Over the last decade, clinical experiences and research studies raised concerns regarding use of proton pump inhibitors (PPIs) as part of the diagnostic strategy for eosinophilic esophagitis (EoE). We aimed to clarify the use of PPIs in the evaluation and treatment of children and adults with suspected EoE in order to develop updated international consensus criteria for EoE diagnosis. METHODS: A consensus conference was convened to address the issue of PPI use for esophageal eosinophilia using a process consistent with standards described in the Appraisal of Guidelines for Research and Evaluation II. Pediatric and adult physicians and researchers from gastroenterology, allergy, and pathology subspecialties representing 14 countries utilized on-line communications, teleconferences, and a face-to-face meeting to review the literature and clinical experiences. RESULTS: Substantial evidence documented that PPIs reduce esophageal eosinophilia in children, adolescents and adults, with several mechanisms potentially explaining the treatment effect. Based on these findings, an updated diagnostic algorithm for EoE was developed, with removal of the PPI trial requirement. CONCLUSIONS: EoE should be diagnosed when there are symptoms of esophageal dysfunction and at least 15 eosinophils per high-power field (or ~60 eosinophils per mm(2)) on esophageal biopsy, and after a comprehensive assessment of non-EoE disorders that could cause or potentially contribute to esophageal eosinophilia. The evidence suggests that PPIs are better classified as a treatment for esophageal eosinophilia that may be due to EoE than as a diagnostic criterion, and we have developed updated consensus criteria for EoE that reflect this change.

Johns Hopkins University School of Medicine, Baltimore, MD, {Erasmus Medical Center–Sophia Children’s Hospital, Rotterdam, The Netherlands, {Children, Youth, and Women’s Health Service, Adelaide, Australia, §Cedars-Sinai Medical Center, David Geffen School of Medicine at UCLA, Los Angeles, CA, Astrid Lindgren Children’s Hospital, Stockholm, Sweden, jjChild Institute–Hospital Journal of Pediatric Gastroenterology and Nutrition 47:266–272 # 2008 by European Society for Pediatric Gastroenterology, Hepatology, and Nutrition and North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition

Received April 24, 200 Authors’ affiliations ap Address correspondence MD, University of Penn Gastroenterology, Hepato of Philadelphia, 34th and The FIGER Symposiu Society of Pediatric Gast Academy of Allergy, Asth ship for Eosinophilic Di Institutes of Health; a phil and educational grants fro cia, TAP, Ception, Glaxo upta, James J. Lee, Simon P. Hoga Barry K. Wershil, Marc E. Rothenberg, Steven J. Ackerman, Ignatius Gomes, Simon Murch, Anil Mishra, and Glenn T. Furuta

Objective The aetiology of necrotising enterocolitis (NEC) is unknown, but luminal factors and epithelial leakiness appear critical triggers of an inflammatory cascade. A separate finding has been suggested in mouse models, in which disruption of glial cells in the myenteric plexus induced a severe NEC-like lesion. We have thus looked for evidence of neuroglial abnormality in NEC. Methods We studied full-thickness resected specimens from 20 preterm infants with acute NEC and from 13 control infants undergoing resection for other indications. Immunohistochemical analysis was performed for immunological (CD3, syndecan-1, human leucocyte antigen-DR), neural (glial fibrillary acidic protein [GFAP], nerve growth factor receptor, neurofilament protein, neuron-specific enolase), and functional markers (Ki67), and for potential inflammatory regulators (interleukin-12, transforming growth factor-β, CCL20, CCR6). Results Expression of the chemokine CCL20 and its receptor CCR6 was significantly upregulated in myenteric plexus in NEC, with CCL20 strongly expressed by glial cells. In 9 of 20 cases with NEC, myenteric plexus architecture and GFAP+ glial cells were normal, with preserved submucosal and mucosal innervation; however, 11 cases showed disrupted myenteric plexus architecture, reduced GFAP expression, and loss of submucosal and mucosal innervation. Persistent abnormalities were identified in the 2 infants who had ongoing inflammation at ileostomy closure. Conclusions Our findings identified heterogeneity among patients with NEC. Approximately half showed evidence of marked neural abnormality extending from the deeper layers of the intestine, associated with glial activation and myenteric plexus disruption. The factors that may activate enteric glia in this manner, potentially including bacterial products or viruses, remain to be determined.

Summary While Crohn's disease and ulcerative colitis are both conditions characterized by intestinal inflammation, with some overlap in their clinical and histological features, they are essentially different in pathogenesis. Crohn's disease appears to be primarily a condition of chronic T-lymphocyte activation, with tissue damage induced by secondary macrophage activation. What activates the T-cells is unknown. In this chapter we look at the evidence for and against cell-wall deficient mycobacteria species, viral infection of vascular endothelium and luminal contents as potential mechanisms of chronic activation. In ulcerative colitis, by contrast, there is no strong evidence for T-cell activation, and humoral mechanisms predominate. While the finding of atypical anti-neutrophil cytoplasmic antibodies (P-ANCAs) may be useful in screening, the only novel pathogenetic discovery is the co-localization of a 40 kD colonic autoantibody with immunoglobulins and complement on the apical enterocyte surface. Despite the fundamental differences in initiating mechanisms, the twoconditions have many ‘downstream’ inflammatory processes in common. We discuss the evidence for local production of cytokines, arachidonic acid metabolites and reactive oxygen and nitrogen radicals, highlighting the potential adverse consequences for intestinal vascular integrity.

Summary In the absence of a definitive cure for Crohn's disease and ulcerative colitis,the aim of therapy must be to induce and maintain clinical remission at acceptable cost to the patient in terms of adverse effects. Despite the differences in their pathogenesis, the first-line treatments for Crohn's disease and ulcerative colitis are still based upon combinations of aminosalicylic acid derivatives and corticosteroids, although the use of enteral nutrition regimes is becoming increasingly widespread in Crohn's disease. In this chapter we attempt to provide reasonably didactic guidance for the management of most cases of chronic inflammatory bowel disease. However, we have tried to go beyond this brief, motivated by the recent explosion in knowledge of inflammatory mechanisms, to suggest a rational approach to the choice of newer and less well tested therapeutic approaches in the affected child who is not responding effectively. The relative failure of cyclosporine therapy in Crohn's disease has been particularly disappointing in view of its ideal theoretical suitability. However, the encouraging early reports of treatment with anti-CD4 and anti-TNFα monoclonals suggest that the shift from broad spectrum immunomodulation to the targeting of critical components of the inflammatory cascade may yet field important dividends.

Background: Enterotoxin-producing Staphylococcus aureus may cause severe inflammatory intestinal disease, particularly in infants or immunodeficient or elderly patients. They are also recognized to be associated with sudden infant death syndrome. Little is known, however, about mucosal responses to staphylococci. Methods: The mucosal lesion in three infants with staphylococcal enterocolitis was assessed by immunohistochemistry and electron microscopy. The organisms underwent extensive molecular analysis. Their toxins were assessed for capacity to induce T-cell activation and host mucosal responses examined by in vitro organ culture. Epithelial responses were studied by coculture with HEp-2 and Caco-2 cells. Results: Intestinal biopsies from the patients showed marked epithelial damage with mucosal inflammation. The three staphylococci, representing two distinct clones, were methicillin-sensitive, producing SEG/I enterotoxins and Rho-inactivating EDIN toxins. Their enterotoxins potently activated T cells, but only whole organisms could induce in vitro enteropathy, characterized by remarkable epithelial desquamation uninhibited by tacrolimus. EDIN-producing staphylococci, but not their supernatants, induced striking cytopathy in HEp-2 epithelial cells but not in Caco-2 cells. Although HEp-2 and Caco-2 cells produced similar IL-8, CCL20, and cathelicidin LL37 responses upon bacterial exposure, only Caco-2 cells expressed mRNA for the β-defensins HBD2 and HBD3, while HEp-2 cells were unable to do so. Conclusions: Staphylococci induce enterocolitis by a combination of direct enterocyte cytopathy mediated by EDIN toxins, disrupting the epithelial barrier, and enterotoxin superantigen-induced mucosal T-cell activation. Gut epithelial production of β-defensins may contribute to host defense against invasive staphylococcal disease. (Inflamm Bowel Dis 2011;)

Background: T-cell hypersensitivity is implicated in the tissue damage in a variety of gastrointestinal diseases. The aim of this study was to characterize the spectrum of pathological changes induced by T-cell activation in explants of fetal human small intestine, to modulate the pathology by immunological intervention, and to define the cellular pathways in intestinal damage. Methods: Explants of fetal human small intestine were cultured with pokeweed mitogen in the presence or absence of dexamethasone or FK506. Pathological changes were then assessed by morphology or on frozen sections. Results: Activation of lamina propria T cells produced variable mucosal damage that could be classified into two main types. Some explants showed villous atrophy and crypt hyperplasia whereas in others there was almost total mucosal destruction. This latter appearance predominated in explants from older specimens. Explants that were destroyed contained high numbers of T cells and activated macrophages. FK506 could completely inhibit mucosal damage, especially in younger specimens, whereas dexamethasone, even at high doses, prevented mucosal destruction, but individual explants continued to show villus atrophy and crypt hyperplasia. Conclusions: These results indicate that there is a quantitative relationship between mucosal cell-mediated immunity and the pathological outcome and that large numbers of activated macrophages in the mucosa can result in tissue destruction.

Background: Human ex vivo evidence indicating that an inappropriate immune response(s) to nonpathogenic bacteria contributes to disease pathogenesis in pediatric Crohn's disease (CD) is limited. The aim of the present study was to compare and contrast the early innate immune response of pediatric "healthy'' versus CD mucosa to pathogenic, probiotic, and commensal bacteria. Methods: "Healthy control'' and CD pediatric mucosal biopsies (terminal ileum and transverse colon) were cocultured for 8 hours with E. coli O42, Lactobacillus GG (LGG), Bacteroides thetaiotaomicron (B. theta), or stimulated with interleukin (IL)-1 beta (positive control). Matched nonstimulated biopsies served as experimental controls. IL-8 was the immune marker of choice. IL-8 mRNA and protein levels were quantified by quantitative polymerase chain reaction and sandwich enzyme-linked immunosorbent assay, respectively. Results: IL-8 secretion was observed when control, ileal biopsies were exposed to pathogenic O42 and probiotic LGG, with no response noted to commensal B. theta. In comparison, Crohn's ileal biopsies showed impaired ability to induce IL-8 in response to O42 and LGG. Control colonic tissue showed a limited response to O42 or B. theta and LGG significantly reduced IL-8 secretion. Unlike control tissue, however, Crohn's ileal and colonic tissue did respond to B. theta, with more enhanced expression in the colon. Conclusions: We provide the first ex vivo data to support the notion that aberrant mucosal recognition of commensal bacteria may contribute to pediatric CD. While IL-8 responses to O42 and LGG varied with disease status and anatomical location, B. theta consistently induced significant IL-8 both in ileal and colonic CD tissue, which was not seen in control, healthy tissue. (Inflamm Bowel Dis 2011;17:1201-1208)

Mutations in alpha6 or beta4 integrins (ITGA6, ITGB4) are known to cause junctional epidermolysis bullosa with pyloric atresia (JEB-PA), often lethal in infancy through skin desquamation. There is 1 report of pyloric atresia associated with a desquamatory enteropathy but without skin disease, of unknown molecular basis.We report 2 Kuwaiti siblings with pyloric atresia and life-threatening intestinal desquamation without significant skin abnormality. The older sibling died of intractable diarrhoea, and the younger sibling suffered episodes of massive protein-losing enteropathy, triggered by viral infections, in addition to obstructive uropathy. Mutation analysis was performed for ITGA6 and ITGB4 and expression of ITGA6 and ITGB4 protein was examined in skin and intestinal biopsies. Her serum also was incubated with normal intestine.We identified a novel mutation in ITGB4, with homozygous deletion of a single residue (isoleucine 1314) within the intracellular plectin-binding domain. Expression of ITGA6 and ITGB4 within skin, duodenal, and colonic epithelium was normal or minimally reduced, in contrast to previous reports. Biopsies taken during relapse showed accumulation of immunoglobulin G and C1q within intestinal basement membrane, whereas immunoglobulin G from her serum bound to basement membrane of normal small intestine. Immunomodulatory therapy induced significant improvement following relapses.ITGB4 mutation may induce a desquamative enteropathy in infancy without significant skin disease. A history of pyloric atresia is important in infants with severe chronic diarrhoeal disease and should prompt investigation for JEB-PA associated mutations. Acquired immune responses may exacerbate primary genetic disorders of epithelial adhesion and immunomodulatory therapy may be beneficial.

Background and Aim: Small bowel disease in the paediatric population is varied and to date has relied on indirect l modalities such as small bowel follow-through with attendant radiation exposure. Wireless capsule endoscopy (WCE) has the potential to provide a safer and more effective means of investigating the paediatric small bowel. The aim of our study was to prospectively assess the diagnostic yield of WCE compared with standard investigation in children with suspected small bowel disease. Methods: Twenty-eight consecutive patients, median age 12.5 y (range, 9.4–15.9) with suspected small bowel disease were investigated with WCE. This included 16 patients with suspected small bowel Crohn disease (CD) (10 newly diagnosed; 6 known cases), 6 with obscure or occult gastrointestinal bleeding (GIB), 3 with Peutz-Jegher polyposis (PJP), 2 with protein-losing enteropathy and 1 with recurrent abdominal pain. All of the patients had preceding upper gastrointestinal endoscopy (OGD) and ileocolonoscopy, and 24 had a barium meal and follow-through (BMFT). Images were downloaded and analysed and results compared with the endoscopic and radiological findings. Results: Three patients were unable to swallow the capsule (1 CD, 1 PJP and 1 GIB). Two of these patients (1 GIB, 1 PJP) had the capsule placed in the stomach endoscopically and completed the WCE uneventfully thereafter. In 3 patients (CD group) the capsule remained in the stomach and/or proximal duodenum and no small bowel images were obtained. Hence, 24 patients had successful completion of the WCE through the small bowel, 23 of whom had clinically relevant findings identified. In all patients with CD who had successful WCE studies (12/16), small bowel disease was identified (11/12 active disease, 1/12 chronic disease). A possible small bowel bleeding source was identified in all 6 patients with GIB. Two patients with GIB also underwent push enteroscopy and 1 of these had a bleeding source identified. The 2 patients with protein-losing enteropathy had extensive patchy lymphangiectasia of the jejunum and ileum, not detected at OGD. The patient with abdominal pain had an intussusception of the upper jejunum. The 2 PJP patients had small bowel polyps identified, which were not detected at BMFT. WCE was more sensitive for small bowel pathology than both BMFT (19 vs 5; 26% sensitivity compared with WCE) and endoscopic investigations (23 vs 10; 43.4% sensitivity compared with WCE). Two patients with CD had delayed capsule transit. Conclusions: WCE led to a positive alteration in management in 18/24 (75%) of patients whose small bowel was examined by WCE and in 18/28 (64.3%) who were admitted to the study. WCE was safe, well tolerated, and more sensitive than radiological and standard endoscopic modalities in the detection of small bowel CD distribution, GIB source, and presence of polyps in children.

This study prospectively compared the rates of skill acquisition in ileocolonoscopy in 2 consecutive groups of trainees in paediatric gastroenterology, with 1 cohort exposed to virtual endoscopy. All paediatric gastroenterology trainees rotating through our department during a 7-year period between 1997 and 2004 were formally assessed while performing ileocolonoscopies using a trainer case-by-case method. Fourteen consecutive trainees with no previous experience of ileocolonoscopy were assessed. Comparison of rates of skill acquisition and lesion recognition using multiple linear regressions revealed a significant acceleration of achievement of endoscopic goals (P < 0.0001) in the group with prior exposure to virtual endoscopy.

Purpose of review The relationship between allergy and motility has been controversial. There is, however, accumulating evidence demonstrating that mucosal allergic responses may disrupt gut motility, and may also potentially alter nociceptive pathways to cause visceral hyperalgesia. Recent findings Experimental studies implicate T helper 2 cells and the cytokines interleukin-4 and -13 in antigen-induced dysmotility, and interleukin-5 in the pathogenesis of mucosal eosinophilia. Both mast cells and eosinophils play obligatory roles in different forms of experimental antigen-induced dysmotility. Overall clinical findings appear to implicate eosinophil infiltration in proximal and distal dysmotility syndromes (oesophageal, gastric and colorectal), and induced mast cell degranulation in mid-gut dysmotility. There is also evidence that mucosal allergic responses may induce long-term changes in visceral perception, including alteration of limbic response, leading to sustained abnormality in visceral sensation. Summary Clinical evidence implicating mucosal allergic responses in dysmotility has been extended to include disorders considered previously entirely functional, such as in some cases of irritable bowel syndrome. Only a proportion of cases are, however, caused by food allergy and a future challenge is to differentiate patients with similar symptoms, but induced by different mechanisms.

Protein-losing enteropathy (PLE), the excessive loss of plasma proteins through the intestine, often correlates with the episodic loss of heparan sulfate (HS) proteoglycans (HSPG) from the basolateral surface of intestinal epithelial cells. PLE onset is often associated with a proinflammatory state. We investigated whether loss of HS or treatment with the proinflammatory cytokine TNF-alpha directly causes protein leakage and whether a combination of both exacerbates this process. We established the first in vitro model of PLE and measured the flux of albumin/FITC through a monolayer of intestinal HT29 or Caco-2 cells grown on transwells and determined the integrity by transepithelial electrical resistance (TER). Loss of HS from the basolateral surface, either by heparanase digestion or by inhibition of HS synthesis, increased albumin flux 1.58 +/- 0.09-fold and reduced TER by 23.4 +/- 6.5%. TNF-alpha treatment increased albumin flux 4.04 +/- 0.03-fold and reduced TER by 75.7 +/- 4.7% but only slightly decreased HS content. The combined effects of HS loss and TNF-alpha treatment were not only additive, but synergistic, with a 7.00 +/- 0.11-fold increase in albumin flux and a 83.9 +/- 8.1% reduction of TER. Coincubation of TNF-alpha with soluble HS or heparin abolished these synergistic effects. Loss of basolateral HS directly causes protein leakage and amplifies the effects of the proinflammatory cytokine TNF-alpha. Our findings imply that loss of HSPGs renders patients more susceptible to PLE and offer a potential explanation for the favorable response some PLE patients have to heparin therapy.

BACKGROUND: Proteoglycans, consisting of glycosaminoglycan (GAG) side chains covalently linked to a protein core, are a major component of the extracellular matrix of the intestinal lamina propria. AIMS: This study investigated the effects of lamina propria T cell activation on the proteoglycan component of the matrix. METHODS: The high degree of sulphation of GAGs means that they are polyanionic and thus can be visualised in tissue sections by means of colloidal-gold labelled cationic probes. RESULTS: In human fetal small intestine there is a dense meshwork of anionic residues in the lamina propria and basement membrane. When explants of human fetal small intestine are cultured ex vivo, and resident lamina propria T cells are activated with pokeweed mitogen, mucosal destruction occurs within three days. This is associated with the rapid loss of anionic sites from the lamina propria. Dermatan sulphate proteoglycan is lost from the tissue and is present at increased concentrations in the organ culture supernatants, indicating that T cell activation has led to solubilisation of lamina propria proteoglycans. Tissue destruction and loss of anionic residues are inhibited in a dose dependent fashion by dexamethasone, and by the protease inhibitor, alpha 2 macroglobulin. CONCLUSIONS: Proteolytic degradation of the lamina propria may therefore be a mechanism by which T cell hypersensitivity injures the intestinal mucosa.

Background/Aims: Cytokines are thought to be important in mediating tissue damage in inflammatory bowel disease (IBD). Many of the in vivo activities of tumor necrosis factor α (TNF-α) match the changes found in IBD, but its importance is controversial. Methods: A sensitive, reverse hemolytic plaque assay was used to determine the frequency of TNF-α secreting cells isolated from mucosal biopsy specimens of children with Crohn's disease or ulcerative colitis (UC) and non-IBD controls before and after medical treatment. Results: Frequency of TNF-α secreting cells was significantly increased in biopsy specimens from children with mild, nonspecific inflammation compared with those with histologically normal intestine. Frequency did not increase in UC compared with children with nonspecific inflammation but was significantly greater in Crohn's disease than in UC. After treatment, the frequency of TNF-α secreting cells was reduced in patients receiving cyclosporin A, not reduced in patients with steroids or enteral nutrition, and not changed with treatment in UC. Conclusions: TNF-α secreting cells are increased in the mucosa of inflamed intestine, regardless of pathogenesis. In patients with IBD, higher levels are seen in Crohn's disease than in UC, probably reflecting the extensive T-cell activation in Crohn's disease. No relation existed between histological healing and the frequency of TNF-α-secreting cells.

Simple routine blood tests (full blood count, platelet count, erythrocyte sedimentation rate, C reactive protein, and serum albumin) may select children with chronic gastrointestinal symptoms who require endoscopic assessment and are thus an important aid in the prediction of endoscopic status. When all of the results are normal then chronic inflammatory bowel disease is an unlikely diagnosis.