Ulrike Strauch

The role of the sympathetic nervous system in intestinal inflammation

The nervous system in the intestine controls motility, secretion, sensory perception, and immune function. Peptidergic neurones with neurotransmitters such as substance P and nerve growth factors have been the main focus of neuroimmunomodulation research in the gut. This review summarises the present knowledge concerning the role of the sympathetic nervous system (SNS) in modulating intestinal inflammation. The role of the SNS for gut inflammation is compared with its role in rheumatoid arthritis which demonstrates notable similarities. Nerve fibres of the SNS not only enter the enteric plexuses but also innervate the mucosa and gut associated lymphoid tissue (GALT). The SNS has pro- and anti-inflammatory functions. Neurotransmitters such as norepinephrine, adenosine, and others can evoke remarkably different opposing effects depending on concentration (presence of sympathetic nerve fibres and extent of neurotransmitter release), receptor affinity at different receptor subtypes, expression of adrenoceptors, availability of cotransmitters, and timing of SNS activity in relation to the inflammatory course. This review attempts to integrate the different perspectives of the pro- and anti-inflammatory effects of the SNS on inflammatory disease of the gut.

Influence of intestinal bacteria on induction of regulatory T cells: lessons from a transfer model of colitis

Background: The resident flora plays a critical role in initiation and perpetuation of intestinal inflammation, as demonstrated in experimental models of colitis where animals fail to develop disease under germ free conditions. However, the importance of exposure to commensal bacteria before the onset of colitis is unclear. Our aim was to investigate the influence of previous exposure of donor animals to bacterial antigens on colitis development using a transfer model. Methods: Clinical course and histology were evaluated after transfer of CD4+CD62L+ lymphocytes from germ free and conventionally housed donor mice into SCID recipients. Cotransfer of CD4+CD62L+ cells with CD4+CD62L− lymphocytes from both groups of mice was initiated. Lymphocytes were analysed by FACS, polarisation potential of cells determined, and cytokines measured within the supernatant by enzyme linked immunosorbent assay. Results: Animals that received cells from germ free donors developed an earlier onset of colitis compared with mice reconstituted with lymphocytes from conventionally housed animals. Additionally, CD4+CD62L− cells from germ free mice were not able to abrogate colitis induced by cotransfer with CD4+CD62L+ lymphocytes whereas CD4+CD62L− T cells from normal mice ameliorated disease. The higher percentage of CD4+GITR+ expressing lymphocytes and the production of interleukin 10 after priming by dendritic cells suggests the presence of Treg cells within the CD4+CD62L+ lymphocyte subset derived from conventional housed mice and assumes a lack of Treg cells within germ free mice. Conclusion: The results indicate that bacterial antigens are crucial for the generation and/or expansion of Treg cells in a healthy individual. Therefore, bacterial colonisation is of great importance in maintaining the immunological balance.

Development of the Lémann Index to Assess Digestive Tract Damage in Patients With Crohn's Disease

BACKGROUND & AIMS There is a need for a scoring system that provides a comprehensive assessment of structural bowel damage, including stricturing lesions, penetrating lesions, and surgical resection, for measuring disease progression. We developed the Lémann Index and assessed its ability to measure cumulative structural bowel damage in patients with Crohns disease (CD). METHODS We performed a prospective, multicenter, international, cross-sectional study of patients with CD evaluated at 24 centers in 15 countries. Inclusions were stratified based on CD location and duration. All patients underwent clinical examination and abdominal magnetic resonance imaging analyses. Upper endoscopy, colonoscopy, and pelvic magnetic resonance imaging analyses were performed according to suspected disease locations. The digestive tract was divided into 4 organs and subsequently into segments. For each segment, investigators collected information on previous operations, predefined strictures, and/or penetrating lesions of maximal severity (grades 1-3), and then provided damage evaluations ranging from 0.0 (no lesion) to 10.0 (complete resection). Overall level of organ damage was calculated from the average of segmental damage. Investigators provided a global damage evaluation (from 0.0 to 10.0) using calculated organ damage evaluations. Predicted organ indexes and Lémann Index were constructed using a multiple linear mixed model, showing the best fit with investigator organ and global damage evaluations, respectively. An internal cross-validation was performed using bootstrap methods. RESULTS Data from 138 patients (24, 115, 92, and 59 with upper tract, small bowel, colon/rectum, and anus CD location, respectively) were analyzed. According to validation, the unbiased correlation coefficients between predicted indexes and investigator damage evaluations were 0.85, 0.98, 0.90, 0.82 for upper tract, small bowel, colon/rectum, anus, respectively, and 0.84 overall. CONCLUSIONS In a cross-sectional study, we assessed the ability of the Lémann Index to measure cumulative structural bowel damage in patients with CD. Provided further successful validation and good sensitivity to change, the index should be used to evaluate progression of CD and efficacy of treatment.

The mucosal addressin cell adhesion molecule antibody PF-00547,659 in ulcerative colitis: a randomised study

Background and aims Leucocyte migration to gut mucosa, mediated by integrin binding to mucosal addressin cell adhesion molecule (MAdCAM), is a promising target for therapeutic intervention in inflammatory bowel disease. This first-in-human study of a monoclonal antibody to MAdCAM, PF-00547,659, aimed to explore the safety and preliminary efficacy of this gut-specific mechanism in ulcerative colitis. Methods In this randomised, double-blind placebo-controlled study, 80 patients with active ulcerative colitis received single or multiple (three doses, 4-week intervals) doses of PF-00547,659 0.03–10 mg/kg IV/SC, or placebo. Safety was assessed by adverse events, laboratory tests, and immunogenicity. Exploratory efficacy analyses were based on Mayo score and endoscopic responder rates at weeks 4 and 12. Faecal calprotectin was quantified as a measure of disease activity, and the number of α4β7+ lymphocytes was measured to demonstrate drug activity. Results No obvious drug-related side effects were observed in the PF-00547,659 group, while patient numbers, especially those fully exposed, were small. Overall responder/remission rates at 4 and 12 weeks were 52%/13% and 42%/22%, respectively with combined PF-00547,659 doses compared with 32%/11% and 21%/0%, respectively with placebo. Equivalent endoscopic responder rates were 50% and 42% versus 26% and 29%, respectively. Faecal calprotectin levels decreased to a greater extent with PF-00547,659 than placebo (week 4: 63% vs 18%). Despite variability, there was a trend for an increase in α4β7+ lymphocytes in patients receiving PF-00547,659. Conclusions The favourable short-term safety profile and preliminary efficacy findings for PF-00547,659 in this first-in-human study pave the way for further investigation in larger trials, to establish the role of PF-00547,659 in ulcerative colitis treatment. Trial Register No: NCT00928681.

Preventive effects of Escherichia coli strain Nissle 1917 on acute and chronic intestinal inflammation in two different murine models of colitis.

ABSTRACT Escherichia coli strain Nissle 1917 (EcN) is as effective in maintaining remission in ulcerative colitis as is treatment with mesalazine. This study aims to evaluate murine models of acute and chronic intestinal inflammation to study the antiinflammatory effect of EcN in vivo. Acute colitis was induced in mice with 2% dextran-sodium sulfate (DSS) in drinking water. EcN was administered from day −2 to day +7. Chronic colitis was induced by transfer of CD4+ CD62L+ T lymphocytes from BALB/c mice in SCID mice. EcN was administered three times/week from week 1 to week 8 after cell transfer. Mesenteric lymph node (MLN) cytokine secretion (of gamma interferon [IFN-γ], interleukin 5 [IL-5], IL-6, and IL-10) was measured by enzyme-linked immunosorbent assay. Histologic sections of the colon were analyzed by using a score system ranging from 0 to 4. Intestinal contents and homogenized MLN were cultured, and the number of E. coli-like colonies was determined. EcN was identified by repetitive extragenic palindromic (REP) PCR. EcN administration to DSS-treated mice reduced the secretion of proinflammatory cytokines (IFN-γ, 32,477 ± 6,377 versus 9,734 ± 1,717 [P = 0.004]; IL-6, 231 ± 35 versus 121 ± 17 [P = 0.02]) but had no effect on the mucosal inflammation. In the chronic experimental colitis of the transfer model, EcN ameliorated the intestinal inflammation (histology score, 2.7 ± 0.2 versus 1.9 ± 0.3 [P = 0.02]) and reduced the secretion of proinflammatory cytokines. Translocation of EcN and resident E. coli into MLN was observed in the chronic colitis model but not in healthy controls. Administration of EcN ameliorated acute and chronic experimental colitis by modifying proinflammatory cytokine secretion but had no influence on the acute DSS-induced colitis. In this model, preexisting colitis was necessary for translocation of EcN and resident E. coli into MLN.

Contrasting activity of cytosin-guanosin dinucleotide oligonucleotides in mice with experimental colitis

Intestinal inflammation in inflammatory bowel disease (IBD) and experimental models of colitis is characterized by a dysregulated intestinal immune response with elevated levels of Th1 cytokines. The luminal flora has been implicated as a major factor contributing to the initiation and perpetuation of inflammation in experimental colitis by mechanisms not known. Bacterial DNA contains unmethylated cytosin–guanosin dinucleotides (CpG) which strongly activate Th1‐mediated immune responses. To test whether these CpG‐motifs modulate intestinal inflammation we treated mice with dextran sulphate sodium (DSS)‐induced colitis with CpG‐containing oligodeoxynucleotides (CpG‐ODN). CpG‐ODN given after the onset of DSS colitis aggravated the disease, as indicated by a significantly increased loss of body weight and a 30% increase of the histological score. Further, we found a severe increase of proinflammatory cytokines (interleukin (IL)‐6: 40‐fold; interferon (IFN)‐γ : 11‐fold). In a pretreatment setting CpG‐ODN reduced weight loss significantly and reduced intestinal inflammation by 45%. Colonic IFN‐γ and IL‐6 mRNA levels were reduced by 75%, and IL‐10 was elevated by 400% compared to controls. The prophylactic CpG‐effect was not imitated by IL‐12 because IL‐12 pretreatment was not protective. In time‐course experiments, CpG‐ODN pretreatment over 5 days resulted in a tolerance effect concerning its IFN‐γ‐inducing quality, and during the following days of colitis induction IL‐10 secretion from mesenterial lymph node cells was elevated compared to controls. Therefore, the prophylactic effect of CpG‐ODN might be explained by its tolerizing effect and/or the increased ability for IL‐10 production during the consecutive intestinal inflammation.

Anti-Inflammatory Role of Sympathetic Nerves in Chronic Intestinal Inflammation

Background: Substance P (SP) is a pro-inflammatory neuropeptide in colitis, whereas sympathetic neurotransmitters are anti-inflammatory at high concentrations. Aim and methods: In all layers of the colon, nerve fibre densities of SP+ and sympathetic nerve fibres were investigated (22 Crohn’s disease, six diverticulitis, and 22 controls). In addition, the nerve fibre repellent factor semaphorin 3C (SEMA3C) was studied. The functional role of the sympathetic nervous system was tested in dextran sodium sulfate (DSS) and Il10−/− colitis. Results: In all layers, Crohn’s disease patients demonstrated a loss of sympathetic nerve fibres. Sprouting of SP+ nerve fibres was particularly observed in the mucosa and muscular layer in Crohn’s disease. SEMA3C was detected in epithelial cells, and there was a marked increase of SEMA3C-positive crypts in the mucosa of Crohn’s disease patients compared to controls. In Crohn’s disease, the number of SEMA3C-positive crypts was negatively related to the density of mucosal sympathetic nerve fibres. Sympathectomy reduced acute DSS colitis but increased chronic DSS colitis. Sympathectomy also increased chronic colitis in Il10−/− mice. Conclusions: This study demonstrated a loss of sympathetic and an increase of SP+ nerve fibres in Crohn’s disease. SEMA3C, a sympathetic nerve repellent factor, is highly expressed in the epithelium of Crohn’s disease patients. In chronic experimental colitis, the sympathetic nervous system confers an anti-inflammatory influence. Thus, the loss of sympathetic nerve fibres in the chronic phase of the disease is most probably a pro-inflammatory signal, which might be related to repulsion of these fibres by SEMA3C and other repellents.

Analysis of intestinal haem-oxygenase-1 (HO-1) in clinical and experimental colitis.

Haem‐oxygenase‐1 (HO‐1) has been shown to exert anti‐inflammatory, anti‐apoptotic and anti‐proliferative effects. We investigated HO‐1 expression in patients with inflammatory bowel disease (IBD) and could demonstrate a scattered expression of HO‐1 in the intestinal epithelium of severely inflamed colonic mucosa of patients with IBD compared to control specimens such as diverticulitis, suggesting dysregulated expression in IBD. To further analyse potential mechanisms of HO‐1 induction in the intestine we employed an in vitro epithelial cell apoptosis model and an experimental colitis model. In vitro induction of HO‐1 by the HO‐1 inducer cobalt protoporphyrin (CoPP) resulted in a dose‐dependent down‐regulation of caspase‐3 activation in HT‐29 cells, indicating an anti‐apoptotic function of HO‐1 in the intestine. In vivo, preventive HO‐1 induction by CoPP in acute dextran sodium sulphate (DSS)‐induced colitis led to a significant down‐regulation of colonic inflammation (P < 0·01) with a concomitant reduction in interferon (IFN)‐γ − but unaffected interleukin (IL)‐10‐secretion by isolated mesenteric lymph nodes (P < 0·01). Additionally, TUNEL staining of colonic sections demonstrated fewer apoptotic epithelial cells in the colon of CoPP treated animals. No beneficial effects were observed if HO‐1 was induced by CoPP after the onset of acute colitis or in chronic DSS‐induced colitis. In conclusion, the data suggest a protective role of HO‐1 if it is induced before the onset of inflammation. However, as shown by the lack of effects in established acute or in chronic colitis, the induction of HO‐1 may not be a promising approach for the treatment of IBD.

Evidence for a role of epithelial mesenchymal transition during pathogenesis of fistulae in Crohn's disease

Background: The pathogenesis of fistulae in Crohns disease (CD) patients is barely understood. We recently showed that more than two‐thirds of CD fistulae are covered with flat, mesenchymal‐like cells (transitional cells [TC]) forming a patchy basement membrane. Epithelial‐to‐mesenchymal transition (EMT) is a process of reprogramming epithelial cells, allowing them to migrate more effectively and giving epithelial cells an “invasive” potential. EMT has been suggested to be crucial in fibrosis found in different tissues and diseases. We therefore investigated whether EMT could be involved in the pathogenesis of fistulae formation in CD. Methods: In all, 18 perianal fistulae, 2 enteroenteric, and 1 enterovesical fistulae from 17 CD patients were analyzed. In addition 2 perianal fistulae of non‐CD patients were studied. Hematoxylin and eosin staining, immunohistochemistry for the expression of cytokeratins 8 and 20, &bgr;6‐integrin, E‐cadherin, &bgr;‐catenin, vimentin, and TGF‐&bgr;1 and 2 were performed according to standard techniques. Results: The TC covering perianal or enteroenteric fistulae were strongly positive for cytokeratins 8 and 20 but negative for vimentin, indicating their epithelial origin. &bgr;6‐Integrin and TGF‐&bgr; had the highest staining intensities in the transitional zone between the epithelium and the TC. Expression of junctional proteins such as E‐cadherin was reduced in TC as compared to regular fistulae epithelium. In addition, a translocation of &bgr;‐catenin from the membrane to the cytoplasm was observed. Conclusions: Our data for the first time indicate an expression pattern of epithelial and mesenchymal markers in TC associated with fistulae formation that is characteristic for EMT. Studying the pathways of EMT during intestinal fistulae formation may help to develop new therapeutic strategies.

Association of the novel serologic anti-glycan antibodies anti-laminarin and anti-chitin with complicated Crohn's disease behavior.

Background: We tested a panel of novel serological anti‐glycan antibodies including the previously unpublished anti‐laminarin IgA (Anti‐L) and anti‐chitin IgA (Anti‐C) carbohydrate antibodies for the presence in Crohns disease (CD) patients, diagnosis and differentiation of CD, association with complicated disease behavior, and marker stability over time. Methods: The presence of Anti‐L, Anti‐C, anti‐chitobioside IgA (ACCA), anti‐laminaribioside IgG (ALCA), anti‐mannobioside IgG (AMCA), and anti‐Saccaromyces cervisiae IgG (gASCA) carbohydrate antibodies were tested in serum samples from 824 participants (363 CD, 130 ulcerative colitis [UC], 74 other gastrointestinal diseases, and 257 noninflammatory bowel/gastrointestinal disease controls) of the German IBD‐network by enzyme‐linked immunosorbent assay (ELISA; Glycominds, Lod, Israel) and for perinuclear antineutrophil cytoplasmic antibody (pANCA) by immunofluorescence. Results: In all, 77.4% of the CD patients were positive for at least 1 of the anti‐glycan antibodies. gASCA or the combination of gASCA/pANCA remained most accurate for the diagnosis of CD, but the combined use of the antibodies improved differentiation of CD from UC. Several single markers as well as an increasing antibody response were independently linked to a severe disease phenotype, as shown for the occurrence of complications, CD‐related surgery, early disease onset, and ileal disease location. This was observed for both quantitative and qualitative antibody responses. The antibody status remained stable over time in most IBD patients. Conclusions: A panel of anti‐glycan antibodies including the novel Anti‐L and Anti‐C may aid in differentiation of CD from UC, is associated with complicated CD behavior and IBD‐related surgery, and is stable over time in a large patient cohort. Inflamm Bowel Dis 2009