[Show abstract][Hide abstract] ABSTRACT: Patients with colitis have an increased risk of colorectal cancer, compared to persons without colitis. Many studies have shown chromoendoscopy (CE) to be superior to standard methods of detecting dysplasia in patients with colitis at index examination. We performed a prospective, longitudinal study to compare standard colonoscopy vs CE in detecting dysplasia in patients with inflammatory bowel diseases in a surveillance program.
Full-text · Article · Dec 2015 · Clinical Gastroenterology and Hepatology
[Show abstract][Hide abstract] ABSTRACT: A single nucleotide polymorphism of PRDM1, the gene encoding Blimp-1, is strongly associated with inflammatory bowel disease. Here, we demonstrate that Blimp-1 in CD103+ dendritic cells (DCs) critically contributes to the regulation of macrophage homeostasis in the colon. Dextran sodium sulfate (DSS)-exposed Blimp-1(cko) mice with a deletion of Blimp-1 in CD103+ DCs and CD11c(hi) macrophages exhibited severe inflammatory symptoms, pronounced weight loss, high mortality, robust infiltration of neutrophils in epithelial regions of the colon, an increased expression of proinflammatory cytokines, and a significant decrease in CD103+ DCs in the colon compared to DSS exposed wild type (WT) mice. Purified colonic macrophages from Blimp-1(cko) mice expressed increased levels of matrix metalloproteinase 8, 9 and 12 mRNA. WT macrophages co-cultured with colonic DCs but not bone marrow derived DCs from Blimp-1(cko) produced increased MMPs in an IL-1β and IL-6 dependent manner. Treatment of Blimp-1(cko) mice with anti-IL-1β and anti-IL-6 abrogated the exaggerated clinical response. Overall, these data demonstrate that Blimp-1 expression in DCs can alter an innate inflammatory response by modulating the activation of myeloid cells. This is a novel mechanism of contribution of Blimp-1 for the pathogenesis of inflammatory bowel diseases, implicating another therapeutic target for the development of inflammatory bowel disease.
Full-text · Article · Dec 2014 · Molecular Medicine
[Show abstract][Hide abstract] ABSTRACT: Inflammation during inflammatory bowel disease may alter nutrient availability to adherent mucosal bacteria and impact their metabolic function. Microbial metabolites may regulate intestinal CD4 T-cell homeostasis. We investigated the relationship between inflammation and microbial function by inferred metagenomics of the mucosal microbiota from colonic pinch biopsies of patients with inflammatory bowel disease.
Paired pinch biopsy samples of known inflammation states were analyzed from ulcerative colitis (UC) (23), Crohn's disease (CD) (21), and control (24) subjects by 16S ribosomal sequencing, histopathologic assessment, and flow cytometry. PICRUSt was used to generate metagenomic data and derive relative Kyoto Encyclopedia of Genes and Genomes Pathway abundance information. Leukocytes were isolated from paired biopsy samples and analyzed by multicolor flow cytometry. Active inflammation was defined by neutrophil infiltration into the epithelium.
Carriage of metabolic pathways in the mucosal microbiota was relatively stable among patients with inflammatory bowel disease, despite large variations in individual bacterial community structures. However, microbial function was significantly altered in inflamed tissue of UC patients, with a reduction in carbohydrate and nucleotide metabolism in favor of increased lipid and amino acid metabolism. These differences were not observed in samples from CD patients. In CD, microbial lipid, carbohydrate, and amino acid metabolism tightly correlated with the frequency of CD4Foxp3 Tregs, whereas in UC, these pathways correlated with the frequency of CD4IL-22 (TH22) cells.
Metabolic pathways of the mucosal microbiota in CD do not vary as much as UC with inflammation state, indicating a more systemic perturbation of host-bacteria interactions in CD compared with more localized dysfunction in UC.
No preview · Article · Feb 2014 · Inflammatory Bowel Diseases
[Show abstract][Hide abstract] ABSTRACT: Recently, a study from the Consortium of Food Allergy Research (CoFAR) showed that allergen-induced IL-4 expression in CD25(+) mononuclear cells was increased in allergic patients. However, they did not find the expected increase in GATA-3 expression, suggesting that allergen-induced IL-4 might not be of T-cell origin. We sought to determine whether other cell types were responsible for the increased IL-4 expression in the CD25(+) cell population.
Comparing six allergic patients and six healthy controls, we analyzed the CD25(+) isolated population from PBMC for the presence of potential IL-4-expressing non-T cells. We also compared spontaneous expression levels of surface markers (CD203c, CD63, CD25, and HLA-DR) on basophils from whole blood of 42 peanut-allergic patients and from 12 non-atopic controls. Expression of these markers was also evaluated following basophil activation in eight peanut-allergic patients selected from the previous cohort.
In addition to CD4(+) T cells, a substantial proportion of non-T cells were found in the CD25(+) -isolated cell population: basophils, NK, and NK-T cells with a mean percentage ± s.e.m. of 5.24 ± 0.63%, 6.65 ± 1.01%, and 6.01 ± 1.04%, respectively. The majority of these cells exhibited positive intracytoplasmic staining for IL-4. Expression of CD63 and CD25 was significantly higher in allergic patients compared with controls (p < 0.05). Interestingly, we found a significantly higher proportion of activated basophils expressing HLA-DR, compared with non-activated basophils (p < 0.05).
Our results support the suggested key role of non-T cells secreting IL-4 in food allergy, particularly basophils, which may also play a central role in antigen presentation.
No preview · Article · Feb 2014 · Pediatric Allergy and Immunology
[Show abstract][Hide abstract] ABSTRACT: Inflammatory bowel disease (IBD) is comprised of Crohn's disease (CD), characterized by transmural inflammation involving the entire GI tract and ulcerative colitis (UC), which involves the mucosal lining of the colon. IBD results from a complex interplay between genetic factors and environmental triggers. It is believed that aberrant innate immune responses to commensal flora play an important role in IBD pathogenesis. Genome-wide association studies have identified 99 non-overlapping susceptibility loci for IBD. The analysis of these genetic loci underscores the importance of several pathways that are involved in the maintenance of gut homeostasis: epithelial restitution and barrier function, regulation of innate and adaptive immunity, microbial defense, and autophagy. Older treatment strategies are directed toward immune suppression, while newer therapies target members of the recently discovered molecular pathways, such as proinflammatory cytokines and adhesion molecules, providing a better prognosis and quality of life to patients with IBD.
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: The pathogenesis of inflammatory bowel disease (IBD) may be caused by abnormal interactions between the immune system and microbiome. Recent studies using 16S ribosomal sequencing have shown that IBD is associated with dysbiosis of the microbiota. Inflammation may alter nutrient availability to adherent mucosal bacteria and impact their metabolic function. Microbial metabolites may also regulate intestinal CD4+ T cell homeostasis. We investigated the relationship between inflammation and microbial function by inferred metagenomics of the mucosal microbiota from colonic pinch biopsies of IBD patients to characterize differences in microbial metabolic pathways between inflamed and non-inflamed biopsy sites.METHODS: Institutional review board approval was obtained before involving patients in the study. Paired pinch biopsy samples of known inflammation states were analyzed from UC (23), CD (21) and controls (24) by 16S ribosomal sequencing and inferred metagenomics with comparison to pathology results and flow cytometry data. The V4 region of the 16S rRNA gene was amplified and sequenced on a MiSeq sequencer. Sequences were assigned to operational taxonomic units (OTUs) and classified taxonomically according to the Ribosomal Database Project (RDP) for use in taxonomic analysis. PICRUSt was then used with the Greengenes OTU database to generate metagenomic data, and derive relative Kyoto Encyclopedia of Genes and Genomes (KEGG) Pathway abundance information. Leukocytes were isolated from paired biopsy samples and activated with PMA/Ionomycin for intracellular cytokine (IL-22, IL-17, IFN-g, IL-4 and TNFa) as well as nuclear antigen (Foxp3) analysis by multi-color flow cytometry on a BD LSRII. Active inflammation was defined by neutrophil infiltration into the epithelium, in the setting of epithelial cell damage.RESULTS: Carriage of metabolic pathways in the mucosal microbiota was relatively stable among IBD patients despite large variations in individual bacterial community structures (Fig. 1). However, microbial function was significantly altered in inflamed tissue of UC patients, with a reduction in carbohydrate and nucleotide metabolism in favor of increased lipid and amino acid metabolism (Fig. 2). These differences were not observed in samples from CD patients. In CD patients, microbial lipid and carbohydrate metabolism was tightly correlated with frequency of CD4+Foxp3+ Tregs, whereas in UC patients lipid and carbohydrate metabolism was correlated with frequency of CD4+IL-22+ (TH22) cells (Fig. 3).CONCLUSIONS: Metabolic pathways of the mucosal microbiota in CD do not vary as much as UC with inflammation state, indicating a more systemic perturbation of host-bacteria interactions in CD compared to more localized dysfunction in UC. The alterations in metabolic pathways correlate specifically with frequency of Tregs during CD, but with TH22 cells during UC. Alterations to metabolic pathways of the mucosal microbiota may affect the production of metabolites that can regulate intestinal CD4+ T cell populations and inflammatory responses of the gut.(C) Crohn's & Colitis Foundation of America, Inc.
No preview · Article · Dec 2013 · Inflammatory Bowel Diseases
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: CD8+ regulatory T (TrE) cells can be activated by intestinal epithelial cells (IECs) through the complex formed by the non-classical I molecule, CD1d and gp180 both expressed on IECs. mAb B9, an anti-gp180 Ab, can block this induction. We have previously demonstrated sequence homology between gp180 and CEACAM5. Furthermore, CEACAM5 has properties attributed to gp180, such as CD1d and CD8[alpha] binding and activation of CD8-associated LcK. Furthermore, the N70, 81A CEACAM5 mutant, which is devoid of the sugar bridge, appeared to lose the capacity to induce the phosphorylation of Lck when compared to CEACAM5. We suggest that unique set of interactions between CEACAM5, CD1d and CD8[alpha] can render CD1d more class I-like, facilitating antigen presentation by CD1d to T cells and the activation of CD8+ Tregs. Moreover, we showed that CEACAM5-activated CD8+ T cells acquire suppressive function. The aim of this study is to determine which amino acids of CEACAM5 are functional in the activation of CD8+ T cells in order to design synthetic peptides with immunoregulatory function.METHODS: We generated an overlapping peptide library of CEACAM5 N domain to identify the amino acids that contribute to its functionality (19 peptides with offset of 5). We tested the ability of the entire pool of peptides, 3 combinations of peptides and each single peptide to phosphorylate CD8 associated p56Lck kinase both in human isolated CD8+ T cells and in a murine T cell line transfected with human CD8[alpha], 3G8, using In-cell Western Blot assay. OKT8, an anti-CD8 antibody, was used as positive control.RESULTS: The entire collection of peptides induced phosphorylation of Lck compared to OKT8 and to the full length CEACAM5 in CD8+ T cells but not in CD4+ T cells. When pooled peptides were tested, pool 3>>2 (pool 2: G30-T55; pool 3: P66-Y107) resulted in the activation of CD8+ T cells. The peptides 12 (T56-N70) and P14 (I66-Q80) within pool 3 induces phosphorylation of LcK to the greatest extent.CONCLUSIONS: Our data suggest the crucial role of the N domain sugar bridge site in CEACAM5 binding to CD8[alpha] and in the activation of CD8+ Tregs. Furthermore, we identified the regions of the N domain where the functional amino acid residues are located. These finding are critical in designing peptides that can be used to induce CD8+ TrE cells as in Crohn's disease in which a defect of these cells have been described.(C) Crohn's & Colitis Foundation of America, Inc.
No preview · Article · Dec 2013 · Inflammatory Bowel Diseases
[Show abstract][Hide abstract] ABSTRACT: Crohn's disease (CD) is a chronic inflammatory disease with increasing incidence in children. Current medications have potentially serious side effects, hence increasing interest in alternative therapies. We previously developed an herbal formula, FAHF-2, based on a classical traditional Chinese herbal formula Wu Mei Wan that has long been used in China to treat colitis. We investigated FAHF-2's potential anti-inflammatory effects.
FAHF-2 efficacy was tested in vivo in the CD45RbRAG1 transfer colitis model. Weight loss, colonic histology, and cytokine production from mesenteric lymph nodes were assessed. Human peripheral blood mononuclear cells (PBMCs) and colonic biopsies were obtained from children newly diagnosed with CD and controls and cultured with or without FAHF-2. Cytokine levels were measured by multiplex immunoassay. The effect of FAHF-2 on TNF-α-producing cells was determined by flow cytometry. NF-κB signaling was investigated in human lamina propria mononuclear cells upon FAHF-2 treatment by In-Cell Western.
FAHF-2-treated mice had decreased weight loss, improved histology, and reduced TNF-α, IL-17, IL-6, and IFN-γ production. In vitro treated PBMCs produced less TNF-α, IFN-γ, and IL-12. FAHF-2 reduced the TNF-α-producing monocytes and T cells. Inflamed CD biopsies produced less TNF-α, IL-17, IL-6, and IL-1β. These effects are because of decreased NF-κB activation.
FAHF-2 inhibited both adaptive and innate immune proinflammatory cytokine responses in PBMCs and inflamed CD mucosa due in part to blockage of NF-κB activation. FAHF-2 was effective in halting progression of colitis in a murine model. This study shows that FAHF-2 has potential as a novel treatment of CD.
No preview · Article · Nov 2013 · Inflammatory Bowel Diseases
[Show abstract][Hide abstract] ABSTRACT: Normal intestinal epithelial cells (IECs) could act as non-professional antigen-presenting cells, selectively activating CD8(+)-suppressor T cells. An epithelial cell surface glycoprotein, gp180, recognized by monoclonal antibodies B9 and L12 was determined to be critical in this process. Purification and sequence analysis of mAb B9 reactive material revealed amino-acid sequence homology with CEACAM5. We demonstrate that CEACAM5 has properties attributed to gp180, such as CD8α binding and activation of CD8-associated Lck. CEACAM5 is the only CEACAM member interacting with CD1d through the B3 domain. Its N domain (recognized by B9) is required for CD8α binding. Removal of the N-domain glycosylated residues reduces B9 recognition, CD8α binding affinity, and activation of LcK. Therefore, conformational changes in CEACAM5 glycosylation site are critical for its interaction with CD8α. CEACAM5-activated CD8(+) T cells acquire the ability to suppress the proliferation of CD4(+) T cells in vitro in the presence of interleukin (IL)-15 or IL-7. We provide new insights into the role of CEACAM5 and define its specific immunoregulatory properties among the CEACAMs expressed on IECs. We suggest that unique set of interactions between CEACAM5, CD1d, and CD8 render CD1d more class I-like molecule, facilitating antigen presentation and activation of CD8(+)-suppressor regulatory T cells.Mucosal Immunology advance online publication, 9 October 2013; doi:10.1038/mi.2013.80.
Full-text · Article · Oct 2013 · Mucosal Immunology
[Show abstract][Hide abstract] ABSTRACT: A dense mucous layer in the large intestine prevents inflammation by shielding the underlying epithelium from luminal bacteria and food antigens. This mucous barrier is organized around the hyperglycosylated mucin MUC2. Here, we show that the small intestine has a porous mucous layer, which permitted the uptake of MUC2 by antigen-sampling dendritic cells (DCs). Glycans associated with MUC2 imprinted DCs with anti-inflammatory properties by assembling a galectin-3-Dectin-1-FcγRIIB receptor complex that activated β-catenin. This transcription factor interfered with DC expression of inflammatory but not tolerogenic cytokines by inhibiting gene transcription through nuclear factor-κB. MUC2 induced additional DC-conditioning signals via intestinal epithelial cells. Thus, mucus does not merely form a nonspecific physical barrier, but also constraints the immunogenicity of gut antigens by delivering tolerogenic signals.
[Show abstract][Hide abstract] ABSTRACT: Developing efficacious vaccines against enteric diseases is a global challenge that requires a better understanding of cellular recruitment dynamics at the mucosal surfaces. The current paradigm of T cell homing to the gastrointestinal (GI) tract involves the induction of α4β7 and CCR9 by Peyer's patch and mesenteric lymph node (MLN) dendritic cells (DCs) in a retinoic acid-dependent manner. This paradigm, however, cannot be reconciled with reports of GI T cell responses after intranasal (i.n.) delivery of antigens that do not directly target the GI lymphoid tissue. To explore alternative pathways of cellular migration, we have investigated the ability of DCs from mucosal and nonmucosal tissues to recruit lymphocytes to the GI tract. Unexpectedly, we found that lung DCs, like CD103(+) MLN DCs, up-regulate the gut-homing integrin α4β7 in vitro and in vivo, and induce T cell migration to the GI tract in vivo. Consistent with a role for this pathway in generating mucosal immune responses, lung DC targeting by i.n. immunization induced protective immunity against enteric challenge with a highly pathogenic strain of Salmonella. The present report demonstrates novel functional evidence of mucosal cross talk mediated by DCs, which has the potential to inform the design of novel vaccines against mucosal pathogens.
Full-text · Article · Aug 2013 · Journal of Experimental Medicine
[Show abstract][Hide abstract] ABSTRACT: Crohn's disease (CD) is an inflammatory pathology of the mucosal intestine that results from uncontrolled immune response towards commensal microbes. Clonal expansions of T cells have been found in patients with CD suggesting an antigen-specific stimulation of pathogenic T cells. Here we show, using T-cell receptor repertoire analysis by real-time PCR, that oligoclonal expansions are found in both CD8+ and CD4+ T cells in the blood and intestinal mucosa of CD patients. The majority of CD4+ T-cell-expanded clones are CD4+NKG2D+ T cells. These clonal expansions were found in both inflamed and neighboring healthy tissue and were persisting during the course of the disease. The presence of these CD4+NKG2D+ T-cell clones at the macroscopically normal edge of the surgical resection might be predictive of inflammation relapse post surgery.Mucosal Immunology advance online publication, 14 August 2013; doi:10.1038/mi.2013.51.
Full-text · Article · Aug 2013 · Mucosal Immunology
[Show abstract][Hide abstract] ABSTRACT: T helper type (Th17) cytokines such as interleukin (IL)-17A and IL-22 are important in maintaining mucosal barrier function and may be important in the pathogenesis of inflammatory bowel diseases (IBDs). Here, we analyzed cells from the colon of IBD patients and show that Crohn's disease (CD) patients had significantly elevated numbers of IL-17+, CD4+ cells compared with healthy controls and ulcerative colitis (UC) patients, but these numbers did not vary based on the inflammatory status of the mucosa. By contrast, UC patients had significantly reduced numbers of IL-22+ cells in actively inflamed tissues compared with both normal tissue and healthy controls. There was a selective increase in mono-IL-17-producing cells from the mucosa of UC patients with active inflammation together with increased expression of transforming growth factor (TGF)-β and c-Maf. Increasing concentrations of TGF-β in lamina propria mononuclear cell cultures significantly depleted Th22 cells, whereas anti-TGF-β antibodies increased IL-22 production. When mucosal microbiota was examined, depletion of Th22 cells in actively inflamed tissue was associated with reduced populations of Clostridiales and increased populations of Proteobacteria. These results suggest that increased TGF-β during active inflammation in UC may lead to the loss of Th22 cells in the human intestinal mucosa.Mucosal Immunology advance online publication 22 May 2013; doi:10.1038/mi.2013.31.
Full-text · Article · May 2013 · Mucosal Immunology
[Show abstract][Hide abstract] ABSTRACT: The Ashkenazi Jewish population has a several-fold higher prevalence of Crohn's disease (CD) compared with non-Jewish European ancestry populations and has a unique genetic history. Haplotype association is critical to CD etiology in this population, most notably at NOD2, in which three causal, uncommon and conditionally independent NOD2 variants reside on a shared background haplotype. We present an analysis of extended haplotypes that showed significantly greater association to CD in the Ashkenazi Jewish population compared with a non-Jewish population (145 haplotypes and no haplotypes with P-value <10(-3), respectively). Two haplotype regions, one each on chromosomes 16 and 21, conferred increased disease risk within established CD loci. We performed exome sequencing of 55 Ashkenazi Jewish individuals and follow-up genotyping focused on variants in these two regions. We observed Ashkenazi Jewish-specific nominal association at R755C in TRPM2 on chromosome 21. Within the chromosome 16 region, R642S of HEATR3 and rs9922362 of BRD7 showed genome-wide significance. Expression studies of HEATR3 demonstrated a positive role in NOD2-mediated NF-κB signaling. The BRD7 signal showed conditional dependence with only the downstream rare CD-causal variants in NOD2, but not with the background haplotype; this elaborates NOD2 as a key illustration of synthetic association.Genes and Immunity advance online publication, 25 April 2013; doi:10.1038/gene.2013.19.
Full-text · Article · Apr 2013 · Genes and immunity
[Show abstract][Hide abstract] ABSTRACT: Gastrointestinal disorders such as chronic or acute diarrhea, malabsorption, abdominal pain, and inflammatory bowel diseases can indicate immune deficiency. The gastrointestinal tract is the largest lymphoid organ in the body, so it is not surprising that intestinal diseases are common among immunodeficient patients. Gastroenterologists must therefore be able to diagnose and treat patients with primary immunodeficiency. Immune-related gastrointestinal diseases can be classified as those that primarily develop via autoimmunity, infection, an inflammatory response, or malignancy. Immunodeficient and immunocompetent patients with gastrointestinal diseases present with similar symptoms. However, intestinal biopsies from immunodeficient patients often have distinct histologic features, and these patients often fail to respond to conventional therapies. Therefore, early recognition of symptoms and referral to an immunologist for a basic immune evaluation is required to select appropriate treatments. Treatment of immunodeficient patients with concomitant gastrointestinal disease can be challenging. Therapies for primary immunodeficiency comprise immunoglobulin replacement, antibiotics, immunomodulators, and in severe cases, bone-marrow transplantation. Treatment of immunodeficient patients with concomitant gastrointestinal disease can be challenging, and therapy with immunomodulators is often required for severe disease. This review aims to guide gastroenterologists in diagnosis and treatment of patients with primary immunodeficiency.
No preview · Article · Mar 2013 · Clinical gastroenterology and hepatology: the official clinical practice journal of the American Gastroenterological Association
[Show abstract][Hide abstract] ABSTRACT: OBJECTIVE: Interferon-γ-inducible protein-10 (IP-10 or CXCL10) plays a role in inflammatory cell migration and epithelial cell survival and migration. It is expressed in higher levels in the colonic tissue and plasma of patients with ulcerative colitis (UC). This phase II study assessed the efficacy and safety of BMS-936557, a fully human, monoclonal antibody to IP-10, in the treatment of moderately-to-severely active UC. DESIGN: In this 8-week, phase II, double-blind, multicentre, randomised study, patients with active UC received placebo or BMS-936557 (10 mg/kg) intravenously every other week. The primary endpoint was the rate of clinical response at Day 57; clinical remission and mucosal healing rates were secondary endpoints. Post hoc analyses evaluated the drug exposure-response relationship and histological improvement. RESULTS: 109 patients were included (BMS-936557: n=55; placebo: n=54). Prespecified primary and secondary endpoints were not met; clinical response rate at Day 57 was 52.7% versus 35.2% for BMS-936557 versus placebo (p=0.083), and clinical remission and mucosal healing rates were 18.2% versus 16.7% (p=1.00) and 41.8% versus 35.2% (p=0.556), respectively. However, higher BMS-936557 steady-state trough concentration (Cminss) was associated with increased clinical response (87.5% vs 37.0% (p<0.001) for patients with Cminss 108-235 μg/ml vs placebo) and histological improvements (73.0% vs 41.0%; p=0.004). Infections occurred in 7 (12.7%) BMS-936557-treated patients and 3 (5.8%) placebo-treated patients. 2 (3.6%) BMS-936557 patients discontinued due to adverse events. CONCLUSIONS: Anti-IP-10 antibody, BMS-936557, is a potentially effective therapy for moderately-to-severely active UC. Higher drug exposure correlated with increasing clinical response and histological improvement. Further dose-response studies are warranted. CLINICAL TRIAL REGISTRATION NUMBER:: ClinicalTrials.gov NCT00656890.