Chronic Dextran Sulfate Sodium Colitis Research Articles

PTPN2 controls differentiation of CD4⁺ T cells and limits intestinal inflammation and intestinal dysbiosis.

Loss-of-function variants within the gene locus encoding protein tyrosine phosphatase non-receptor type 2 (PTPN2) are associated with increased risk for Crohn's disease (CD). A disturbed regulation of T helper (Th) cell responses causing loss of tolerance against self- or commensal-derived antigens and an altered intestinal microbiota plays a pivotal role in CD pathogenesis. Loss of PTPN2 in the T-cell compartment causes enhanced induction of Th1 and Th17 cells, but impaired induction of regulatory T cells (Tregs) in several mouse colitis models, namely acute and chronic dextran sodium sulfate colitis, and T-cell transfer colitis models. This results in increased susceptibility to intestinal inflammation and intestinal dysbiosis which is comparable with that observed in CD patients. We detected inflammatory infiltrates in liver, kidney, and skin and elevated autoantibody levels indicating systemic loss of tolerance in PTPN2-deficient animals. CD patients featuring a loss-of-function PTPN2 variant exhibit enhanced Th1 and Th17 cell, but reduced Treg markers when compared with PTPN2 wild-type patients in serum and intestinal tissue samples. Our data demonstrate that dysfunction of PTPN2 results in aberrant T-cell differentiation and intestinal dysbiosis similar to those observed in human CD. Our findings indicate a novel and crucial role for PTPN2 in chronic intestinal inflammation.

Maresin 1, a Proresolving Lipid Mediator Derived from Omega-3 Polyunsaturated Fatty Acids, Exerts Protective Actions in Murine Models of Colitis

It has been previously reported that dietary fish oils, which are rich in the polyunsaturated fatty acids eicosapentaenoic acid and docosahexaenoic acid, can exert beneficial effects in inflammatory bowel disease. In this study, we investigated the effects of docosahexaenoic acid-derived lipid mediator maresin 1 (MaR1) in dextran sulfate sodium (DSS)- and 2,4,6-trinitrobenzenesulfonic acid-induced colitis in mice. Systemic treatment with MaR1 significantly attenuated both DSS- and 2,4,6-trinitrobenzene sulfonic acid-induced colonic inflammation by improving the disease activity index and reducing body weight loss and colonic tissue damage. MaR1 treatment also induced a significant decrease in levels of inflammatory mediators, such as IL-1β, TNF-α, IL-6, and IFN-γ, in the acute protocol, as well as IL-1β and IL-6, but not TNF-α and INF-γ, in the chronic DSS colitis protocol. Additionally, MaR1 decreased ICAM-1 mRNA expression in both the acute and chronic protocols of DSS-induced colitis. Furthermore, the beneficial effects of MaR1 seem to be associated with inhibition of the NF-κB pathway. Moreover, incubation of LPS-stimulated bone marrow-derived macrophage cultures with MaR1 reduced neutrophil migration and reactive oxygen species production, besides decreasing IL-1β, TNF-α, IL-6, and INF-γ production. Interestingly, macrophages incubated only with MaR1 showed a significant upregulation of mannose receptor C, type 1 mRNA expression, an M2 macrophage phenotype marker. These results indicate that MaR1 consistently protects mice against different models of experimental colitis, possibly by inhibiting the NF-κB pathway and consequently multiple inflammatory mediators, as well as by enhancing the macrophage M2 phenotype.

The impact of JNK inhibitor D-JNKI-1 in a murine model of chronic colitis induced by dextran sulfate sodium

Purpose:The c-Jun N-terminal kinases (JNK) are involved in the activation of T cells and the synthesis of proinflammatory cytokines. Several studies have established the relevance of the JNK pathway in inflammatory bowel diseases. The present study analyzed the therapeutic effect of D-JNKI-1, a specific JNK-inhibiting peptide, in a low-dose dextran sulfate sodium (DSS) model of chronic colitis.Methods:DSS colitis was induced in female C57/BL6 mice by cyclic administration using different concentrations of DSS (1.0% and 1.5%). Mice in the intervention groups received subcutaneous administration of 1 μg/kg D-JNKI-1 on days 2, 12, and 22. They were monitored daily to assess the severity of colitis, body weight, stool consistency, and the occurrence of occult blood or gross rectal bleeding using evaluation of the disease activity index. The animals were sacrificed after 30 days, and the inflamed intestine was histologically evaluated using a crypt damage score. Immunohistochemical quantification of CD4+ and CD8+ cells was also carried out.Results:Administration of 1 μg/kg D-JNKI-1 resulted in a significant decrease in the disease activity index (P = 0.013 for 1.0% DSS; P = 0.007 for 1.5% DSS). As a mild form of colitis was induced, histological examination did not show any distinct damage to the mucosa and crypts. However, expression of CD4+ and CD8+ cells was reduced in mice treated with D-JNKI-1 (not significant).Conclusion:Administration of D-JNKI-1 resulted in a clinical attenuation of chronic DSS colitis, and a therapeutic effect of D-JNKI-1 must therefore be assumed. The decrease in CD4+ and CD8+ cells may reflect the influence of D-JNKI-1 on T-cell activation, differentiation, and migration.

Knockout of the c-Jun N-terminal Kinase 2 aggravates the development of mild chronic dextran sulfate sodium colitis independently of expression of intestinal cytokines TNFα, TGFB1, and IL-6

IntroductionThe c-Jun N-terminal kinases (JNKs) are involved in signal transduction of inflammatory bowel diseases. The aim of this study was to examine the function of JNKs by using a low-dose dextran sulfate sodium (DSS) model in JNK1 knockout mice (Mapk8−/−), JNK2 knockout mice (Mapk9−/−), and wild-type controls (WT1, WT2).MethodsThe animals were evaluated daily using a disease activity index. After 30 days, the intestine was evaluated histologically with a crypt damage score. CD4+ and CD8+ cells were quantified using immunofluorescence. Analysis of tumor necrosis factor-α (TNFα), interleukin-6 (IL-6), and transforming growth factor β1 (TGFB1) expression was carried out using LightCycler® real-time polymerase chain reaction.ResultsCyclic administration of low-dose DSS (1%) was not able to induce features of chronic colitis in Mapk8−/− WT2 mice. By contrast, DSS administration significantly increased the disease activity index in WT1 and Mapk9−/− mice. In Mapk9−/− mice, the crypt damage score and the number of CD4+ and CD8+ cells as features of chronic colitis/inflammation were also significantly elevated. Expression of TNFα, IL-6, and TGFB1 was not altered by the JNK knockout.ConclusionAdministering DSS at a defined low concentration that is unable to induce colitis in WT animals leads to clinically and histologically detectable chronic colitis in Mapk9−/− mice. The reason for this disease-inducing effect resulting from the loss of JNK2 remains to be elucidated. Expression of TNFα, IL-6, and TGFB1 does not appear to be involved; proapoptotic JNK2 may prolong the activity of proinflammatory immune cells, leading to perpetuation of the inflammation.

5-Hydroxytryptophan Alleviates Inflammation-Associated Mood Disturbance in Two Mouse Models of Colitis

Anxiety and depression states are common in patients with inflammatory bowel disease (IBD) and correlate with disease activity suggesting a biologic link to inflammation. Experimentally lowered serum levels of tryptophan, the amino acid precursor of serotonin, can induce mood disturbance. We previously reported that in patients with active Crohn's disease serum tryptophan levels can be severely depressed while kynurenine levels are elevated. This elevated kynurenine/tryptophan ratio reflects increased expression of the immunomodulatory enzyme indoleamine 2,3 dioxygenase (IDO) present in active IBD and colitis models. In the current study we aimed to determine if enteral delivery of 5-hydroxytryptophan (5-HTP), the tryptophan-based serotonin precursor, improved mood disturbance or affected colitis severity in two mouse models. Dextran sodium sulfate (DSS) colitis was induced in Bl6 mice with 3% DSS given over 3 5-day courses, with 10-day recovery periods. 2,4,6 trinitrobenzene sulfonic acid (TNBS) colitis was induced in BALB/c mice using 4 escalating doses of TNBS (0.35 mg to 1.2 mg). In each experiment there were 3 groups of 7-10 mice: 1) control group (PBS gavage), 2) colitis control group (PBS gavage), and 3) colitis group with 5-HTP (0.5 mg gavage before testing). Behavior was filmed and scored at three time points: pre-colitis, after acute colitis, and after chronic colitis. Anxiety-like behavior was tested using the light-dark preference test (LDT). Depressive behavior was tested using the tail suspension test (TST). Locomotor activity (LMA) was tested at each time-point to rule out potential physical impairment prior to behavioral testing. Colitis activity was followed clinically (diarrhea, hematochezia, and weight loss) and assessed by histology and myeloperoxidase (MPO) activity. No differences in clinical or histology parameters were identified between colitis groups. Similarly, MPO activity did not differ with or without 5-HTP supplementation in both DSS and TNBS colitis. Locomotor abilities were similar in all 3 groups. No differences in mouse behavior were present at baseline or after acute colitis. Mice with chronic colitis from DSS spent significantly less time in light than controls, indicating anxiety-like behavior. This anxiety behavior was normalized by 5-HTP supplementation. Depressive behavior as measured by TST immobility time was longer in chronic DSS colitis; however, a therapeutic effect of 5-HTP was not observed. In TNBS colitis, mice had fewer entries to the light compared to controls in the LDT. This anxiety-like behavior was normalized by 5-HTP supplementation. There was no effect of TNBS colitis or 5-HTP on immobility as measured by the TST. Table 1 summarizes these findings. Anxiety- and depressive-like behavior is heightened in 2 models of chronic colitis. While anxiety behavior patterns vary by mouse strain, they are normalized by administration of the tryptophan-derived serotonin precursor 5-HTP. Importantly, colitis is not worsened by enteral supplementation with physiologic doses of 5-HTP (i.e. proportional to human dosing). With translational validation, 5-HTP may be considered an intervention for colitis-associated anxiety in IBD patients.

P53 Mediates TNF-Induced Epithelial Cell Apoptosis in IBD

Chronic ulcerative colitis (CUC) is characterized by increased intestinal epithelial cell (IEC) apoptosis associated with elevated tumor necrosis factor (TNF), inducible nitric oxide synthase (iNOS), and p53. We previously showed that p53 is increased in crypt IECs in human colitis and is needed for IEC apoptosis in chronic dextran sulfate sodium-colitis. Herein, we examined the roles of TNF and iNOS in regulating p53-induced IEC apoptosis in CUC. The IEC TUNEL staining, caspases 3, 8, and 9, and p53 protein levels, induced by anti-CD3 monoclonal antibody (mAb) activation of T cells, were markedly reduced in TNF receptor 1 and 2 gene knockout mice. Induction of IEC apoptosis correlated with increased p53, which was attenuated in iNOS(-/-) mice. IEC p53 levels and apoptosis were reduced in IL-10(-/-) colitic mice treated with neutralizing TNF mAb and the iNOS inhibitor, aminoguanidine, further suggesting that TNF and iNOS are upstream of p53 during colitis-induced IEC apoptosis. IEC apoptosis and p53 levels were assessed in control versus untreated or anti-TNF-treated CUC patients with equivalent levels of inflammation. Data indicated that IEC apoptosis and p53 levels were clearly higher in untreated CUC but markedly reduced in patients treated with anti-TNF mAb. Therefore, TNF-induced iNOS activates a p53-dependent pathway of IEC apoptosis in CUC. The inhibition of IEC apoptosis may be an important mechanism for mucosal healing in anti-TNF-treated CUC patients.

IL-33 attenuates development and perpetuation of chronic intestinal inflammation

Interleukin-33 (IL-33) is a member of the IL-1 family. Recent evidence shows the importance of IL-33 in autoimmune and inflammatory diseases. To elucidate its impact on inflammatory bowel disease we studied the effects of exogenous IL-33 during the induction of acute dextran sodium sulfate (DSS)-induced colitis, the induction period of chronic DSS colitis, and after establishment of chronic inflammation. For induction of acute colitis mice received DSS in their drinking water for 7 days and were killed at day 8 or 14 after first DSS administration. Chronic colitis was induced by four cycles of DSS. Animals were treated with IL-33 between the DSS cycles (intermediate treatment) or after onset of chronic disease (posttreatment). Colons and mesenteric lymph nodes were isolated for histology and cytokine secretion, flow cytometric analysis, determination of myeloperoxidase, and transcription factor activity. While IL-33 in acute colitis led to slight aggravation of inflammation, both chronic colitis approaches resulted in a significant reduction of inflammatory colon contraction, amelioration of disease scores, suppression of interferon-gamma (IFN-γ), and a shift to T helper (Th)2-associated cytokines. Examination of colon tissue revealed increased Ly6g-mRNA levels and myeloperoxidase (MPO) activity in IL-33-treated animals. Evaluation of bacterial translocation revealed decreased translocation incidence in IL-33-treated mice. In summary, IL-33 has extenuating effects in chronic DSS-induced colitis: Excessive Th1-directed cytokine responses are shifted toward Th2-like immune reactions and general inflammation parameters are reduced. IL-33-induced neutrophil influx during chronic inflammation reduced translocation of pathogenic bacteria across damaged epithelium.

Extracellular cathepsin K exerts antimicrobial activity and is protective against chronic intestinal inflammation in mice

ObjectiveCathepsin K is a lysosomal cysteine protease that has pleiotropic roles in bone resorption, arthritis, atherosclerosis, blood pressure regulation, obesity and cancer. Recently, it was demonstrated that cathepsin K-deficient (Ctsk−/−)...

In-Vivo Effects of GM-CSF Treated Monocytes in Chronic DSS Colitis

Background. Granulocyte Macrophage-Colony Stimulating Factor (GM-CSF) has been shown to induce clinical response and remission in patients with active Crohn’s disease (CD). Moreover the beneficial role of GM-CSF was shown in a mouse model of acute colitis induced by Dextran Sodium Sulfate (DSS). Since it is known that GM-CSF has pleiotropic effects on monocytes and since those cells represent the exclusive source of macrophages in inflamed intestinal mucosa we characterized GM-CSF treated mouse monocytes in vitro and analyzed their function in vivo in a model of chronic DSS colitis. Methods. Mouse bone marrow-derived monocyte precursors were treated for 48 hours with GM-CSF in vitro and phenotypical changes were assessed by real-time polymerase chain reaction (qRT-PCR) and flow cytometry. Functional properties were evaluated in mixed lymphocyte reactions (MLR), by phagocytosis assays, adherence to plastic surface, cytokine production upon lipopolysaccharide (LPS) stimulation and measurement of reactive oxygen species (ROS) production. Therapeutic effects of GM-CSF-treated monocytes were evaluated in a model of chronic colitis induced by repeated oral administration of DSS (2%) on three days per week. GM-CSF treated monocytes were administered i.v. one day prior start of the third DSS treatment cycle and the severity of developed colitis was evaluated by body weight change, colon length, histology and expression of inflammatory markers. The distribution of injected GM-CSF-monocytes across intestine was evaluated by in vivo near infrared imaging. Results. We found that GM-CSF-treated monocytes express higher levels of anti-inflammatory molecules on mRNA and protein levels (e.g. IL1-Ra, IL4-R and ARG1). Functionally, GM-CSF promotes ROS production of the cells but reduces phagocytosis and adherence. In MLR we observed an induction of Foxp3 in naive T cells. Clinically, in chronic DSS-induced colitis, treatment of mice with GM-CSF-monocytes showed an overall better outcome compared to mice treated with control monocytes. This encompasses body weight, colon length, mRNA expression of inflammatory marker molecules (e.g. IL6 and TNF) and histological observations. Using in vivo imaging technology we found that a higher percentage of GM-CSF treated monocytes enter the inflamed intestine compared to control monocytes. In addition, GM-CSF-monocytes persisted longer at sides of inflammation. Conclusion. We showed that the beneficial effects of GM-CSF in CD are probably due to its modifying action especially on monocytes and concomitant anti-inflammatory alterations in phenotype and function of macrophages developed from these cells in an experimental model of colitis.

Analysis of intestinal fibrosis in chronic colitis in mice induced by dextran sulfate sodium

Fibrogenic mesenchymal cells including fibroblasts and myofibroblasts play a key role in intestinal fibrosis, however, their precise role is largely unknown. To investigate their role in intestinal fibrosis, we analyzed the lesions of chronic colitis in C57BL/6 (B6) mice induced by dextran sulfate sodium (DSS). B6 mice exposed to single cycle administration of DSS for 5 days developed acute colitis that progressed to severe chronic inflammation with dense infiltrates of mononuclear cells, irregular epithelial structure, thickening of colonic wall, and persistent deposits of collagen. Increased mRNA expressions of proinflammatory cytokines are correlated with extensive cellular infiltration, and the mRNA expressions of collagen 1, transforming growth factor (TGF)-β, and matrix metalloproteinases were also enhanced in the colon. In the colon of chronic DSS colitis, fibroblasts (vimentin(+), α-smooth muscle actin (α-SMA)(-)) were increased in both mucosal and submucosal layers, while myofibroblasts (vimentin(+), α-SMA(+)) were increased in mucosal but not in submucosal layers. Primary mouse subcutaneous fibroblast cultures experiments revealed that exogenously added TGF-β 1 substantially augmented the expressions of both vimentin and α-SMA proteins with increased production of collagen. In conclusion, profibrogenic mesenchymal cells play an important role in the development of intestinal fibrosis in this chronic DSS-induced colitis model.

4SC-101, a novel immunosuppressive drug, inhibits IL-17 and attenuates colitis in two murine models of inflammatory bowel disease†

Dihydroorotate dehydrogenase (DHODH) is a key enzyme involved in pyrimidine biosynthesis. DHODH is a known target for the treatment of autoimmune diseases. 4SC-101 is a novel immunosuppressive drug that inhibits DHODH. A goal of our study was to examine the in vitro effects of 4SC-101 on IL-17 production by mononuclear cells. In addition, we evaluated the efficacy of 4SC-101 against acute TNBS (2,4,6-tritrobenzene sulfonic acid) and chronic dextran sodium sulfate (DSS)-induced colitis in mice. Peripheral blood mononuclear cells (PBMCs) from healthy human donors were used to evaluate cellular proliferation and cytokine (IL-17, TNF-α) production. The oral effects of 4SC-101 (100 or 200 mg/kg) were examined following induction of chronic colitis by the administration of 3% DSS (4 cycles) to Balb/c mice. Morphometric and histological indices of colitis were evaluated as indicators of drug efficacy. 4SC-101 was also administered for 6 days after the intracolonic administration of TNBS (20 mg in 50% ethanol) to female Balb/c mice. The colons were analyzed for overall macroscopic damage, ulceration, total length, distal segment weight, MPO activity, and histological pathology as indicators for the effectiveness of 4SC-101. In vitro, 4SC-101 is a potent inhibitor of human DHODH, inhibits lymphocyte proliferation, and uniquely blocks phytohemagglutinin-stimulated IL-17 production by lymphocytes. In vivo, oral administration of 4SC-101 effectively improved both chronic DSS and acute TNBS colitis in mice. In these colitis models the overall efficacy profile of 4SC-101 was similar to that of dexamethasone. 4SC-101 is a novel immunosuppressive drug with excellent potential for the treatment of intestinal inflammation.

Loss of CD103+ intestinal dendritic cells during colonic inflammation.

To investigate possible differences in dendritic cells (DC) within intestinal tissue of mice before and after induction of colitis. Mucosal DC derived from intestinal tissue, as well as from mesenteric lymph nodes and spleen, were analyzed by fluorescence activated cell sorting (FACS) analysis. Supernatants of these cells were analyzed for secretion of different pro- and anti-inflammatory cytokines. Immunohistochemistry and immunofluorescence were performed on cryosections of mucosal tissue derived from animals with colitis as well as from healthy mice. It was shown that DC derived from healthy intestinal lamina propria (LP) represented an immature phenotype as characterized by low-level expression of costimulatory cytokines. In contrast to DC from spleen and mesenteric lymph nodes (MLN) that secreted proinflammatory cytokines, LP-DC produced high levels of the anti-inflammatory cytokine IL-10. After induction of murine colitis in a CD4(+)CD62L(+) transfer model or in chronic dextran sulfate sodium-colitis, a marked increase of activated CD80(+) DC could be observed within the inflamed colonic tissue. Interestingly, in contrast to splenic DC, a significant population of DC within MLN and colonic LP expressed the mucosal integrin CD103 which was lost during colitis. The constitutive secretion of anti-inflammatory cytokines by immature DC within the intestinal LP might regulate the homeostatic balance between mucosal immunity and tolerance. CD103(+) DC could mediate this important function.

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...

Treatment of experimental colitis in mice with LMP-420, an inhibitor of TNF transcription

BackgroundLMP-420 is a boronic acid-containing purine nucleoside analogue that transcriptionally inhibits TNF production but is non-cytotoxic to TNF-producing cells.MethodsThis study investigated the efficacy of LMP-420 as an anti-inflammatory agent in acute and chronic colitis induced by oral administration of dextran sulfate sodium (DSS) to mice and in chronic colitis following piroxicam administration to IL-10-deficient mice. The severity of colon inflammation was assessed histologically. TNF levels were measured by enzyme immunoassay.ResultsAdministration of DSS for 7 days resulted in severe acute colitis that was associated with a marked increase in stool and colon tissue TNF levels. Initiation of therapy with intraperitoneal (i.p.) LMP-420 on day 4 of DSS exposure decreased colonic TNF to near normal levels on day 7. However, neither i.p. nor oral treatment with LMP-420 affected the development or severity of acute DSS colitis. Initiation of LMP-420 therapy after 3 cycles of DSS administration to establish chronic colitis also had no effect on the severity of chronic colitis. Analysis of colonic TNF combined with longitudinal analysis of TNF and TNF receptor (TNF-RII) levels in stool during the development of chronic DSS colitis demonstrated that the initially elevated colonic TNF levels returned to normal despite intense on-going inflammation in mice with chronic colitis. RAG-2-/- mice deficient in T and B cells also developed severe ongoing colitis in response to 3 cycles of DSS, but showed marked differences vs. wild type mice in stool TNF and TNF-RII in response to DSS exposure. Systemic and oral LMP-420 treatment for 16 days decreased colonic TNF levels in IL-10-deficient mice with chronic colitis, with a trend to decreased histologic inflammation for oral LMP-420.ConclusionThese studies demonstrate that short-term treatment with a transcriptional inhibitor of TNF production can decrease systemic and local colonic levels of TNF but may not decrease the histologic severity of colitis. Longer term studies using colitis models that are more dependent on TNF elevation should be performed to more accurately assess the potential of LMP-420 for therapy of inflammatory bowel disease.

PECAM-1 (CD 31) mediates transendothelial leukocyte migration in experimental colitis

Transendothelial migration of circulating leukocytes into the colonic wall is a key step in the development of the inflammatory infiltrate in inflammatory bowel disease (IBD). The platelet-endothelial cell adhesion molecule-1 PECAM-1 (CD31) is expressed in the tight junction area of endothelial cells, where it is supposed to support the transmigration process. The aim of this study was to determine the role of PECAM-1 in experimental IBD and to show whether blockade of PECAM-1 has therapeutic effects. Chronic colitis was induced in female BALB/c mice by cyclic oral administration of dextran sodium sulfate (DSS) 3% (wt/vol). Expression of PECAM-1 was visualized by immunohistochemistry. In the treatment group animals received 1 mg/kg anti-PECAM-1 (2H8) ip daily starting on day 26. On day 30 leukocyte adhesion and migration was measured during N(2)O-isoflurane anesthesia in the distal colon by intravital microscopy. Disease activity index (DAI), histology, and MPO levels were compared with healthy and diseased controls. PECAM-1 was expressed in colitic mice. Chronic DSS colitis was characterized by a marked increase in rolling, adherent, and transmigrated leukocytes compared with healthy controls. Immunoblockade of PECAM-1 reduced leukocyte transmigration significantly and also diminished leukocyte rolling and sticking in an indirect manner. It also resulted in a significantly diminished DAI and MPO levels, as well as an amelioration of the histological inflammation score. PECAM-1 plays an important role in transendothelial leukocyte migration in DSS colitis. PECAM-1 could be a novel target for antibody-based treatment in IBD.

Chemically induced mouse models of intestinal inflammation

Animal models of intestinal inflammation are indispensable for our understanding of the pathogenesis of Crohn disease and ulcerative colitis, the two major forms of inflammatory bowel disease in humans. Here, we provide protocols for establishing murine 2,4,6-trinitro benzene sulfonic acid (TNBS)-, oxazolone- and both acute and chronic dextran sodium sulfate (DSS) colitis, the most widely used chemically induced models of intestinal inflammation. In the former two models, colitis is induced by intrarectal administration of the covalently reactive reagents TNBS/oxazolone, which are believed to induce a T-cell-mediated response against hapten-modified autologous proteins/luminal antigens. In the DSS model, mice are subjected several days to drinking water supplemented with DSS, which seems to be directly toxic to colonic epithelial cells of the basal crypts. The procedures for the hapten models of colitis and acute DSS colitis can be accomplished in about 2 weeks but the protocol for chronic DSS colitis takes about 2 months.

Oral Administration of Taurolidine Ameliorates Chronic DSS Colitis in Mice

Taurolidine (TRD) has antimicrobial and anti-inflammatory properties. However, the anti-inflammatory effects of TRD in inflammatory bowel diseases (IBD) have not been investigated. Here, we have analyzed the toxicity of TRD after oral long-term application in mice and examined the impact of oral TRD in a dextran sulfate sodium (DSS) model of experimental colitis. Female C57/BL6 mice received TRD in various concentrations (0.1% to 0.4%) for 60 days. Toxicity was evaluated by use of a disease activity index (DAI) and histological examination of major metabolic organs. Furthermore, the impact of 0.2% TRD on a chronic DSS colitis was examined by daily DAI, histological crypt damage score (CDS), bacterial translocation into mesenteric lymph nodes (MLN), and colonic expression of tumor necrosis factor (TNF) α, transforming growth factor (TGF) β, interleukin (IL)-1β, IL-6, cytochrome oxidase (COX)-2, and monocyte chemotactic protein (MCP)-1 by real-time polymerase chain reaction (PCR). Oral TRD administration for 60 days was well tolerated by the animals and did not show any toxic effects in terms of DAI and histological changes. TRD treatment of DSS colitis led to increased survival of 100%, compared to 33% in the untreated colitis group (p ≤. 005). Clinical amelioration was mirrored by significantly reduced DAI and CDS in the TRD treated colitis. Colonic cytokine expression and bacterial translocation into MLN showed no differences between both groups.We thus report for the first time that oral application of TRD results in amelioration of an experimental IBD model. We hypothesize direct intraluminal antimicrobial effects of TRD as well as anti-inflammatory effects during the acute phase of DSS colitis.

Epidermal Growth Factor Partially Restores Colonic Ion Transport Responses in Mouse Models of Chronic Colitis

Epidermal growth factor receptor (EGFR) activation, which plays an important role in regulating intestinal ion transport, can alleviate clinical symptoms such as diarrhea in patients with ulcerative colitis and promote mucosal restitution in animal models of colitis. Here, we investigate whether EGFR can regulate colonic ion transport in the setting of colitis. Distal colon from control mice and mice with colitis was retained for immunohistochemistry or mounted in Ussing chambers. Ion transport responses across the tissues to the calcium agonist carbachol and the adenosine 3',5'-cyclic monophosphate agonist forskolin were measured with or without epidermal growth factor (EGF) pretreatment. EGF pretreatment of normal colonic mucosa inhibited ion transport responses to carbachol and forskolin but potentiated the reduced ion transport responses seen in dextran sulfate sodium (DSS)-treated and mdr1a knockout mouse colon. Ion substitution studies and the sodium transport inhibitor amiloride showed that sodium movement primarily accounted for the potentiating effect of EGF in DSS-treated tissues, despite decreased sodium channel expression. EGF potentiation of transport responses in DSS-treated colon was completely blocked by the cytoskeletal disruptor cytochalasin D and the phosphatidylinositol 3-kinase inhibitor wortmannin, whereas the novel and conventional protein kinase C isoform inhibitor Gö6850 and the extracellular signal-regulated kinase inhibitor PD98059 partially reduced EGF effects. EGFR epithelial distribution and transforming growth factor alpha expression were also altered in DSS-treated tissues. Chronic inflammation uncovers a potentiating effect of EGFR activation on epithelial electrogenic sodium absorption that would be expected to ameliorate diarrheal symptoms associated with colitis.

Intracellular polyamine levels of intestinal epithelial cells in inflammatory bowel disease.

Polyamines and their acetylated derivatives are a prerequisite for cellular metabolism and considered to be essential for proliferation and differentiation of the rapidly renewing intestinal mucosa. However, their role during mucosal inflammation is less clear. Polyamine concentrations were determined in isolated colonic epithelial cells (CECs) from endoscopic biopsies from 26 patients with inflammatory bowel disease (IBD) and 40 controls as well as colon samples from mice with and without acute or chronic dextran sodium sulfate (DSS)-induced colitis. In patients with ulcerative colitis, CEC spermidine and N8-acetylspermidine levels were significantly enhanced and spermine levels were reduced compared with healthy controls. A correlation of polyamine levels of patients with IBD with their corresponding inflammatory index revealed that increased concentrations of spermidine, N8-acetylspermidine, and N1-acetylspermine were found in CECs from the most severe inflamed mucosal areas. Using acute and chronic DSS colitis as a model of mucosal inflammation, we found enhanced levels of spermidine and spermine in acute colitis, whereas in chronic inflammation, CEC spermine concentrations were decreased. Our data indicate a lack of the anti-inflammatory polyamine spermine in severe ulcerative colitis and chronic DSS colitis, which may aggravate the disease. Increased spermidine and N8-acetylspermidine levels reflect increased uptake and metabolism likely due to accelerated proliferation and regeneration of CECs.

Ameliorative effect of IDS 30, a stinging nettle leaf extract, on chronic colitis.

Anti-TNF-alpha antibodies are very effective in the treatment of acute Crohn's disease, but are limited by the decline of their effectiveness after repeated applications. The stinging nettle leaf extract, IDS 30, is an adjuvant remedy in rheumatic diseases dependent on a cytokine suppressive effect. We investigated the effect of IDS 30 on disease activity of murine colitis in different models. C3H.IL-10-/- and BALB/c mice with colitis induced by dextran sodium sulphate (DSS) were treated with either IDS 30 or water. Mice were monitored for clinical signs of colitis. Inflammation was scored histologically, and faecal IL-1beta and mucosal cytokines were measured by ELISA. Mononuclear cell proliferation of spleen and Peyer's patches were quantified by 3H-thymidine. Mice with chronic DSS colitis or IL-10-/- mice treated with IDS 30 clinically and histologically revealed significantly (p < 0.05) fewer signs of colitis than untreated animals. Furthermore, faecal IL-1beta and mucosal TNF-alpha concentrations were significantly lower (p < 0.05) in treated mice. Mononuclear cell proliferation after stimulation with lipopolysaccharide was significantly (p < 0.001) reduced in mice treated with IDS 30. The long-term use of IDS 30 is effective in the prevention of chronic murine colitis. This effect seems to be due to a decrease in the Th1 response and may be a new therapeutic option for prolonging remission in inflammatory bowel disease.