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Recently we demonstrated that in inflammatory bowel disease (IBD) macrophage-oxidative burst activity is increased and NADPH oxidase mRNA is induced. The herbal phenylethanoid acteoside isolated from Plantago lanceolata L. was shown to exhibit anti-oxidative potential. Using the dextran sulphate sodium (DSS)-induced colitis model, in this study we have assessed whether systemic application of acteoside affects colitis. Colitis was induced by DSS in Balb/c mice. Treatment with acteoside (120, 600 µg/mouse/day) was performed intraperitoneally. The colon lengths were determined. Colonic tissue was scored histologically (max. score 8) by a blinded investigator. T cells isolated from mesenteric lymph nodes (MLN) were stimulated with anti-CD3 antibody in the presence of interleukin (IL)-2 (final concentration 10 U/ml). After incubation for 24 h, IL-1β, IL-6, IL-12 tumour necrosis factor (TNF)-α and interferon (IFN)-γ levels in supernatants were analysed by the beadlyte® cytokine detection system. Histological scoring of colonic tissue revealed that application of acteoside was followed by a significantly improved histological score. In acute colitis the histological score was 3·2 with acteoside versus 5·2 with phosphate-buffered saline (PBS) ( P < 0·02). In chronic colitis both 120 µg (3·3 versus 5·2) or 600 µg acteoside (3·0 versus 5·2) significantly ameliorated colitis (both P < 0·02). Stimulated MLN from mice with chronic DSS-induced colitis treated with acteoside showed a significant down-regulation of IFN-γ secretion (195 pg/ml with 600 µg acteoside versus 612 pg/ml with PBS, P < 0·02). Inhibition of oxidative burst activity with acteoside reduced mucosal tissue damage in DSS colitis and could be a therapeutic alternative for IBD treatment. Further studies of this agent are warranted. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

The lymphotoxin- β receptor (LT βR) pathway is critical for maintenance of organized lymphoid structures and is involved in the development of colitis. To investigate the mechanisms by which LT βR activation contributes to the pathology of chronic inflammation we used a soluble LT βR-Ig fusion protein as a competitive inhibitor of LT βR activation in the mouse model of chronic colitis induced by oral administration of dextran sulphate sodium. Strong expression of LT β which constitutes part of the LT α1 β2 ligand complex was detected in colonic tissue of mice with chronic colitis. Treatment with LT βR-Ig significantly attenuated the development and histological manifestations of the chronic inflammation and reduced the production of inflammatory cytokines such as TNF, IL-1 β, and IL-6. Moreover, LT βR-Ig treatment significantly down-regulated mucosal addressin cell adhesion molecule-1 (MAdCAM-1) expression, leading to reduced leucocyte rolling and sticking in postcapillary and collecting venules and reduced extravasation into the intestinal mucosa as quantified by in vivo fluorescence microscopy. Thus, LT βR pathway inhibition ameliorates DSS-induced experimental chronic colitis in mice by MAdCAM-1 down-regulation entailing reduced lymphocyte margination and extravasation into the inflamed mucosa. Therefore, a combined treatment with reagents blocking T cell-mediated perpetuation of chronic inflammation such as LT βR-Ig together with direct anti-inflammatory reagents such as TNF inhibitors could constitute a promising treatment strategy for chronic colitis. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

The cathepsins D (CTSD), B (CTSB) and L (CTSL) are important for the intracellular degradation of proteins. Increased cathepsin expression is associated with inflammatory diseases. We have shown previously an induction of CTSD expression in intestinal macrophages (IMAC) in inflamed mucosa of patients with inflammatory bowel disease (IBD). Here we investigated the regulation of CTSB and CTSL in IMAC during IBD and effects of CTSD and CTSB/CTSL inhibition in vivo. Human IMAC were isolated from normal and inflamed mucosa. Reverse transcription–polymerase chain reaction (RT–PCR) was performed for CTSB and CTSL mRNA. Immunostaining was used to confirm PCR results. Cathepsin inhibition was investigated in the dextran–sulphate–sodium (DSS) colitis model in mice with application of pepstatin A (CTSD inhibitor), CA-074 (CTSB inhibitor) and Z-Phe-Tyr-aldehyde (CTSL inhibitor). CTSL mRNA was significantly up-regulated in IMAC isolated from IBD mucosa. Up-regulated protein expression was found mainly in areas of mucosal damage by immunostaining. Inhibition of CTSD in mouse DSS colitis was followed by an amelioration of the disease. Inhibitor-treated mice showed a significant lower histological score (HS) and less colon reduction in comparison to controls. Similarly, simultaneous inhibition of CTSB/CTSL was followed by a significant amelioration of colitis. Expression of tissue-degrading cathepsins is increased in IMAC in IBD. Inhibition of CTSD as well as CTSB/CTSL is followed by an amelioration of experimental colitis. The prevention of mucosal damage by cathepsin inhibition could represent a new approach for the therapy of IBD. [ABSTRACT FROM AUTHOR]

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 anin vitroepithelial cell apoptosis model and an experimental colitis model.In vitroinduction 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. [ABSTRACT FROM AUTHOR]

Background: Studies postulate an involvement of adipokines in inflammatory gastrointestinal diseases. Leptin-deficient ob/ob mice as well as TLR9-deficient mice have a more moderate course of chronic DSS-induced colitis (DSS-CC) and adipocytes do express functional TLR9 molecules. Material and Methods: Adipokine mRNA expression in visceral adipose tissue of mice before and after the induction of DSS-CC was investigated. Experiments were performed in both TLR9wt/wt and TLR9-/- mice. In vitro, the effect of TLR9 blocking peptide on leptin and visfatin protein secretion was studied in 3T3-L1 adipocytes. Results: Induction of DSS-CC led to an upregulation of leptin mRNA expression in TLR9wt/wt mice, while TLR9-/- animals showed a significant reduction of leptin expression even below baseline. While visfatin expression remained unchanged in TLR9wt/wt animals, TLR9-/- mice exhibited a significant induction during DSSCC. CTRP-3 expression was reduced after colitis induction only in TLR9-/- animals. Of note, IL-6 expression levels remained unchanged, while CXCL1/KC and cyclophilin A expression was reduced in DSS-CC. Inhibition of TLR9 signaling by using TLR9 blocking peptide led to reduced leptin protein secretion into cell culture supernatants in 3T3-L1 adipocytes, while visfatin protein secretion was enhanced. Conclusions: DSS-CC leads to differential adipokine expression profiles in the visceral fat pad in TLR9wt/wt vs. TLR9-/- mice. In vitro, inhibition of TLR9 signaling induces visfatin secretion while inhibiting leptin secretion in adipocytes. Thus, visceral adipokines are regulated by intact TLR9 signaling pathway and a specific interplay between the leptin- and the TLR9-pathways might be of pathophysiological importance in chronic intestinal inflammation. [ABSTRACT FROM AUTHOR]

Background: Clinical and experimental studies suggest that appendectomy can protect against development of ulcerative colitis and Crohn’s disease. However, how T cells in the appendix affect the development of colitis has not been clarified. Aim: To investigate the in vivo migration and activation of colitis-inducing CD62L+ cells during development of chronic colitis. Methods: CD62L+CD4+ cells were fluorescently labeled and transferred to severe combined immunodeficient (SCID) mice to induce colitis. In vivo migration of T cells into the mucosa of the appendix and colon was quantified by in vivo microscopy after 7 weeks. In a second experiment, unlabeled CD62L+CD4+ cells were transferred, reisolated after 7 weeks, and adhesion molecule (integrin α4β7) and costimulatory molecule (CD154) expression was analyzed. Results: Six to eight weeks after CD62L+CD4+ cell transfer, SCID mice developed chronic colitis. In vivo microscopic analysis demonstrated a preferential migration of fluorescence-labeled CD62L+CD4+ cells into the mucosa of the appendix versus the colon. Re-isolation of lamina propria cells from mice with colitis confirmed that CD62L+CD4+ cell migration was significantly enhanced in the appendix, compared to the colon (3.5-fold). Furthermore, a higher proportion of CD62L+CD4+ cells re-isolated from the appendix expressed integrin α4β7 and CD154 than from the colon. Conclusion: This study demonstrates the preferential migration of CD62L+CD4+ cells into the appendix as compared to the colon. This migration pattern correlated with upregulation of integrin α4β7 and CD154 (CD40 ligand) on T cells. Our results suggest an important role of the appendix in the pathogenesis of colitis. Copyright © 2005 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]