Development Of Acute Colitis Research Articles

Bacteroides dorei BDX-01 alleviates DSS-induced experimental colitis in mice by regulating intestinal bile salt hydrolase activity and the FXR-NLRP3 signaling pathway.

Background: The relationships among intestinal dysbiosis, bile acid (BA) metabolism disorders, and ulcerative colitis pathogenesis are now recognized. However, how specific strains regulate BA metabolism to alleviate colitis is still unclear. This study investigated the effects of Bacteroides dorei on the development of acute colitis and elucidated the underlying mechanisms. Methods: The safety of BDX-01 was evaluated in vitro and in vivo. 2.5% dextran sulfate sodium (DSS) induced colitis in C57BL/6 mice, Caco-2, and J774A.1 cells were used to evaluate the anti-inflammatory effect of BDX-01. qPCR and Western blotting were used to detect the expression of inflammatory pathways. Microbiota composition was analyzed by 16S rRNA gene sequencing. Enzyme activity analysis and targeted metabolomics were used to analyze fecal bile salt hydrolase (BSH) and BA levels. Antibiotic-induced pseudo-germ-free mice were used to investigate the role of gut microbiota in the alleviation of colitis by BDX-01. Results: We confirmed the safety of novel strain Bacteroides dorei BDX-01 in vitro and in vivo. Oral BDX-01 administration significantly ameliorated the symptoms and pathological damage of DSS-induced acute colitis. Moreoever, 16S rRNA sequencing and enzyme activity analysis showed that BDX-01 treatment increased intestinal BSH activity and the abundance of bacteria harboring this enzyme. Targeted metabolomics revealed that BDX-01 significantly increased intestinal BA excretion and deconjugation. Certain BAs act as FXR agonists. The β-muricholic acid (βMCA): taurine β-muricholic acid (T-βMCA) and cholic acid (CA): taurocholic acid (TCA) ratios and the deoxycholic acid (DCA) level decreased markedly in the colitis models but increased substantially in BDX-01-treated mice. The colonic farnesoid X receptor (FXR) and fibroblast growth factor 15 (FGF15) were upregulated in mice treated with BDX-01. BDX-01 downregulated the expression of colonic proinflammatory cytokines pyrin domain-containing 3 (NLRP3), ASC, cleaved caspase-1, and IL-1β. Antibiotic treatment didn't abolish the protective effect of BDX-01 on colitis. In vitro studies showed TβMCA abolished the effects of BDX-01 on FXR activation and inhibition of the NLRP3 inflammasome activation. Conclusion: BDX-01 improved DSS-induced acute colitis by regulating intestinal BSH activity and the FXR-NLRP3 signaling pathway. Our findings indicate that BDX-01 is a promising probiotic to improve the management of ulcerative colitis.

A54 THE IMPACT OF NON-INVASIVE TRANSCUTANEOUS AURICULAR VAGUS NERVE STIMULATION (NITAVNS) ON THE DEVELOPMENT OF EXPERIMENTAL COLITIS IN MICE

Abstract Background The anti-inflammatory impacts of stimulating the vagus nerve as a part of the gut-brain axis on intestinal macrophage activity and inflammatory cytokine production have been documented. While vagus nerve stimulation until now has been chiefly invasive, non-invasive auricular vagus nerve stimulation (NitaVNS) has no adverse effects associated with invasive stimulation. The NitaVNS-mediated activation of brainstem nuclei can exert anti-inflammatory effects directly on the colon or indirectly through the spleen via a macrophage-acetylcholine-dependent pathway. Purpose To examine the effects of NitaVNS on the development of acute colitis in mice using a preclinical model. Method Thirty C57BL/6 mice aged 11 to 12 weeks were divided into four groups: control (CTRL)-NitaVNS, CTRL-Sham, dextran sulphate sodium (DSS)-NitaVNS and DSS-Sham. Pre-set stimulation parameters (10 minutes duration, 10 V voltage, 500 s pulse width, and 20 Hz frequency) were applied for NitaVNS or sham-paw stimulation. 5% DSS was administered to induce colitis for five consecutive days, whereas CTRL mice received regular water. Weight loss, stool consistency, blood in the feces, and disease activity index were evaluated daily. Rectal bleeding, colon bleeding, and fecal consistency were determined as a composite macroscopic score upon sacrifice. Collected colon and blood samples were assessed for pro-inflammatory parameters such as interleukin (IL)-1β, tumor necrosis factor (TNF)-α, IL-6, C-reactive protein (CRP), matrix metalloproteinase (MMP)-2, macrophage inflammatory protein (Mip)-1α and β, and anti-inflammatory cytokine tumor growth factor (TGF)-β using qRT-PCR and ELISA. Result(s) Compared to colitic-sham mice, colitic NitaVNS mice demonstrated a delayed onset and a significantly lower clinical disease severity. Colitis induced significant weight loss, rectal bleeding, and stool consistency loss, and NitaVNS significantly abolished these effects. In colitic conditions, NitaVNS decreased the disease activity index significantly compared to the colitic sham group. Colitis increased the macroscopic scores, and NitaVNS decreased them significantly. Conversely, colon length was significantly reduced in colitic conditions, but NitaVNS abrogated that deleterious effect. mRNA expression levels of IL-1β and TNF-α were significantly increased in colitic mice and NitaVNS decreased them significantly. Whereas Mip-1α and 1β were not modified in colitic conditions, NitaVNS significantly decreased these markers. MMP2 mRNA expression and colonic IL-6 and IL-1β protein levels were not altered in all conditions. However, colitis induced a significant decrease in colonic TGF-β levels, but NitaVNS did not amplify that effect. Interestingly, in the serum of colitic mice, NitaVNS significantly increased and decreased TGF-β and CRP levels, respectively. Conclusion(s) These findings suggest that NitaVNS possess some spatial anti-inflammatory effects in acute colitis through the up or downregulation of anti- or pro-inflammatory markers. Disclosure of Interest F. Hesampour: None Declared, D. Tshikudi: None Declared, A. Veysel Özden Employee of: Company providing the stimulator, C. N. Bernstein: None Declared, J.-E. Ghia: None Declared

Біохімічні ефекти олій розторопші та льону холодного і гарячого пресування у щурів за умов гострого коліту

Inflammatory bowel disease (IBD) which includes ulcerative colitis (UC), is one of the most serious and currently unsolved problems in modern gastroenterology. In terms of severity and frequency of complications, IBD occupies one of the leading places in the structure of gastrointestinal tract diseases. The etiology of IBD is still not fully understood. It is probably of an autoimmune nature, and the main causes are considered to be hereditary predisposition, allergic reactions, nutrition, etc. Dystrophic and atrophic changes of the colonic mucosa, accompanied by its secretory and motor function alterations, digestive disorders, as well as extraintestinal manifestations (general intoxication, liver and skin injuries, joints) are main IBD features. Because tumor growth is usually accompanied by inflammation of tumor nodules in adjacent apparently normal tissue, and prolonged pharnaceutical suppression of inflammation significantly reduces the risk of tumor development, chronic UC is considered as a precursor condition. Moreover, the number of colorectal cancer cases among people with UC history exceeding 10 years, increases eightfold compared to the average population. In this regard, it is important to prevent this pathology, in particular with the help of food supplements of natural origin, such as vegetable oils of flax and milk thistle, which are used in traditional medicine, including inflammatory diseases of the digestive system. Studies on their possible anti-inflammatory effects on the colonic mucosa and biochemical parameters of blood in ulcerative colitis have not been performed. Therefore, the aim is to evaluate the anti-inflammatory effectiveness of vegetable oils (flaxseed and milk thistle, obtained by different methods of pressing) in a model of acute colitis in rats. Object of research: biochemical mechanisms of realization of influence of thistle and flaxseed oils of different methods of extraction at ulcerative colitis. The effects of linseed and milk thistle oils obtained by cold pressing (to 45°C) and hot pressing (to 95°C) on the development of acute colitis in rats, when administered with food, have been investigated. It was shown that the use of linseed oil both cold and hot-pressed under conditions of adding it to the feed in an amount of 10% for 2 weeks prevents the development of inflammatory and destructive changes in the colon of rats with acute colitis and its negative consequences in the liver (for approaching to the normal values of serum markers of its functional activity). These oil properties could be realized partially by unsaturated fatty acids, in particular ω3, including minor ones. The data obtained may indicate that one of the mechanisms of implementation of the anti-inflammatory action of these substances is their ability to reduce oxidative stress - one of the main factors and promoters of inflammation.

Structural Changes in the Nervous Fibers of the Colon Mucosa in Experimental Acute Colitis.

Morphological, morphometric, and ultrastructural analysis of the nerve fibers in the colon mucosa was performed in C57BL/6 mice at various terms of development of acute colitis induced by dextran sodium sulphate. The nerve fibers were labeled with antibodies to pan-neuronal marker βIII-tubulin. The progression of inflammatory and ulcerative processes in the mucosa on days 3-5 was associated with hyperplasia and hypertrophy of nerve fibers that peaked on day 7 after colitis induction. Ultrastructural analysis at all terms of colitis development showed moderate degeneration of axons. Thus, hypertrophy and hyperplasia of the nervous fibers in colon mucosa in experimental acute colitis correlated with aggravation of the ulcerative process in the mucosa. These changes are determined by alteration of histoarchitectonics and regenerative processes in the mucosa.

Lack of Adrenomedullin Aggravates Acute TNBS-Induced Colitis Symptoms in Mice, Especially in Females.

Adrenomedullin (AM) is a biologically active peptide which has been tested as a new therapy for inflammatory bowel disease (IBD) in animal models and in patients with severe ulcerative colitis. We used an inducible knockout (KO) mouse model for AM to evaluate the effects of endogenous levels of this peptide on the development and degree of pathogenesis of IBD. Acute colitis was induced in mice of both sexes by rectal instillation of 3 mg 2,4,6-trinitrobenzenesulfonic acid (TNBS) in 100 μL of 50% ethanol. Control mice received the same volume of saline in 50% ethanol. During the following 5 days, the weight and the disease severity index of all animals were recorded. After sacrifice, the inflammatory response was macroscopically assessed by analyzing the weight of the colon; by histomorphometrical analysis on histological sections; and by qRT-PCR determination of different inflammatory, adhesion, and regeneration molecules. TNBS administration caused a significantly more severe colitis in KO mice, and especially in females, when compared to wild type (WT) animals. Abrogation of the AM gene caused more severe diarrhea, accompanied by rectal bleeding, anorexia, and a significant increase of colon weight. Histological analysis of TNBS-treated KO mice showed large areas of lymphocyte infiltrates in the mucosa and submucosa, with loss of tissue architecture. No alterations were observed in the expression levels of inflammatory cytokines at the time of sacrifice; meanwhile lack of AM resulted in lower levels of some adhesion molecules and regeneration markers. Taken together, these results support the protective role of endogenous AM against the development of acute colitis, and that its effects are particularly beneficial on females.

PI3K p85\u03b1 Subunit-deficient Macrophages Protect Mice from Acute Colitis due to the Enhancement of IL-10 Production

We investigated the role of the PI3K p85α subunit in the development of acute colitis with a focus on intestinal macrophages. Experimental acute colitis was induced using 3% dextran sulfate sodium (DSS) in drinking water for 7 days. The severity of DSS-induced acute colitis was significantly attenuated in p85α hetero-deficient (p85α+/−) mice compared with WT mice. The expression of proinflammatory mediators in intestinal macrophages isolated from the inflamed colonic mucosa was significantly suppressed in p85α+/− colitis mice compared with WT colitis mice. Interestingly, we found that bone marrow-derived macrophages (BMDMs) from p85α+/− mice produced a significantly higher amount of IL-10 than BMDMs from WT mice. The adoptive transfer of p85α+/− BMDMs, but not WT BMDMs, significantly improved the severity in WT colitis mice, and this effect was reversed by anti-IL-10 antibody. Furthermore, the expression of IL-10 in the intestinal macrophages of p85α+/− normal colonic mucosa was significantly higher than that in the intestinal macrophages of WT normal colonic mucosa. The present results demonstrate that p85α+/− mice exhibit a reduced susceptibility to DSS-induced acute colitis. Our study suggests that a deficiency of PI3K p85α enhances the production of IL-10 in intestinal macrophages, thereby suppressing the development of DSS-induced acute colitis.

Вплив водорозчинних С60 фулеренів на розвиток гострого коліту у щурів

Вплив водорозчинних С60 фулеренів на розвиток гострого коліту у щурів

Abstract 1030: Loss of PHLPPs protects against colitis by inhibiting epithelial apoptosis through upregulating AKT activity

Abstract Background & Aims: PHLPP, a novel family of protein phosphatases, plays an important role in balancing cellular signaling pathways that control cell death and proliferation. Loss of PHLPP expression promotes colorectal cancer (CRC) tumorigenesis and progression. Intestine epithelial cells (IECs) apoptosis contributes to the development of inflammatory bowel disease (IBD), and patients with IBD are at increased risk for CRC. We investigated the functional importance of PHLPP in intestinal inflammation. Methods: Both Phlpp1 and Phlpp2 genes were deleted in mice. The effect of PHLPP double knockout (DKO) on the development of acute colitis was examined in dextran sulfate sodium salt (DSS)-treated mice. Biochemical and histological analyses were conducted to determine the extent of tissue inflammation and colon epithelial cell apoptosis. The expression of PHLPP proteins was examined in DSS-treated mice and IBD patient samples. Results: In ulcerative colitis and Crohn's disease patient samples, both PHLPP1 and PHLPP2 protein expression was markedly decreased in colitis tissues compared to that in uninvolved tissues. In mice, PHLPP expression was significantly reduced in colonic epithelial cells following induction of colitis by DSS. Mechanistically, the inflammation-induced downregulation of PHLPP was mediated through proteasome-dependent protein degradation. Compared with wild-type mice, PHLPP DKO mice were protected from colitis induced by the DSS treatment as demonstrated by lower histopathological scores, and this reduced susceptibility to colitis was associated with decreased apoptosis and increased Akt activity in colonic epithelial cells. Furthermore, epithelial organoids derived from colon and small intestine of PHLPP DKO mice are more resistant to inflammation-induced apoptosis upon Akt activation. Conclusion: These results indicated that PHLPP contributes to the pathogenesis of colitis by controlling epithelial cell apoptosis. Downregulation of PHLPP at the onset of inflammation plays a role in protecting intestinal epithelial cell injury by inhibiting apoptosis. However, prolonged PHLPP-loss may promote colon cancer initiation in IBD patients. Citation Format: Yang-an Wen, Xin Li, Tatiana Goretsky, Terrence A. Barrett, Tianyan Gao. Loss of PHLPPs protects against colitis by inhibiting epithelial apoptosis through upregulating AKT activity. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1030. doi:10.1158/1538-7445.AM2015-1030

Macrophage‐Specific ErbB4 is Induced by DSS Colitis and Regulates Macrophage Survival

The ErbB4 receptor tyrosine kinase is induced in the colonic epithelium in inflammatory bowel disease (IBD), and the ErbB4-specific ligand neuregulin-4 is protective in murine colitis models. Interestingly, ErbB4 is also robustly detectable in the submucosal stroma where immune cells accumulate during inflammation. Previous studies have shown ErbB4 expression on circulating human monocytes and neuronal macrophages (Mø), but its expression on intestinal Mø and role in Mø biology are unknown. HYPOTHESIS: As Mø play a significant role in the development of acute colitis, we hypothesized that receptor-mediated ErbB4 signaling may be an anti-inflammatory mechanism to limit Mø numbers during inflammation. METHODS: Mice (C57Bl6) were given 3% dextran sodium sulfate (DSS) in drinking water for 4 days (injury phase) followed by 3 days without DSS (recovery phase); colonic Mø populations were characterized by flow cytometry. Murine bone marrow-derived Mø (BM-Mø) were generated for in vitro activation and survival studies. RESULTS: DSS induced colonic recruitment of F480+/CD11b+/Ly6C+/ErbB4+ cells in injury and recovery phase mice compared to controls. In vitro, classical activation (IFNγ/LPS) of BM-Mø, but not alternative activation (IL-4), induced ErbB4 expression 10-fold (*p<0.01). Treatment of these cells with neuregulin-4 (100 ng/ml) reduced activated BM-Mø numbers by 40% (*p<0.05) after 48 hours, an effect blocked by ErbB4 neutralizing antibody (2 μg/ml). CONCLUSION Colitis induces recruitment of ErbB4+ Mø. ErbB4 signaling is a possible inhibitory feedback mechanism restricting Mø numbers, and thus represents a potential therapeutic target to regulate inflammation in IBD.

Mo1757 Evidence for a Role of Superantigens in the Pathogenesis of Ulcerative Colitis-Associated Primary Sclerosing Cholangitis

G A A b st ra ct s 0.002) and downregulation of Mmp-2 (FC=3.3, p=0.020), Mmp-9 (FC=3.1, p=0.030), Cldn1 (FC=3, p=0.040) compared to control mice. After chronic DSS administration, Xpnpep2 (FC=4.2, p=0.020) was upregulated in TIMP-1 KO mice compared to control mice. Young TIMP-1 KO mice (10-12 weeks) versus older animals (19-21 weeks) showed significantly increased expression of Reg3g (FC=16, p=0.030), Reg3b (FC=12.6, p=0.040) and Ido1 (FC= 3.5, p=0.008), whereas the expression levels of Mir200b (FC=2.2, p=0.009), Ctse (FC=1.3, p=0.030) and Wnt2b (FC=1.1, p=0.002) were decreased. Conclusion: TIMP-1 deficiency leads to upregulation of anti-bacterial and innate immunity genes, resulting in an attenuated development of acute colitis. In a chronic setting of inflammation, TIMP-1 KO mice have less remodeling and fibrosis. Unraveling the role of TIMP-1 in extracellular matrix remodeling will be necessary to understand the biology of intestinal wound healing and fibrosis in IBD.

Nicotine suppresses acute colitis and colonic tumorigenesis associated with chronic colitis in mice.

Ulcerative colitis is a chronic inflammatory disease that frequently progresses to colon cancer. The tumor-promoting effect of inflammation is now widely recognized and understood. Recent studies have revealed that treatment with nicotine ameliorates colitis in humans and experimental murine models, whereas the effect of nicotine on colitis-associated colonic tumorigenesis remains unclear. In the present study, we examined the effect of nicotine on the development of acute colitis and colitis-associated cancer (CAC). The acute colitis model was induced by treatment with 3% dextran sulfate sodium (DSS) for 7 days, whereas the CAC model was induced by a combination of azoxymethane and repeated DSS treatment. Nicotine and a selective agonist of the α7-nicotinic acetylcholine receptor (α7-nAChR) reduced the severity of DSS-induced acute colonic inflammation. In addition, the suppressive effect of nicotine on acute colitis was attenuated by an antagonist of α7-nAChR. Furthermore, nicotine inhibited the IL-6 production of CD4 T cells in the DSS-induced inflamed colonic mucosa. We found that nicotine significantly reduced the number and size of colonic tumors in mice with CAC. Nicotine markedly inhibited the elevation of TNF-α and IL-6 mRNA as well as phosphorylated signal transducer and activator of transcription (Stat) 3 expression in the colons of the tumor model mice. These results demonstrate that nicotine suppresses acute colitis and colitis-associated tumorigenesis, and this effect may be associated with the activation of α7-nAChR. Furthermore, it is presumed that nicotine downregulates the expression of inflammatory mediators such as IL-6/Stat3 and TNF-α, thereby reducing the colonic tumorigenesis associated with chronic colitis.

S1656 CD4+CD25+ Regulatory T Cells Suppress Both TH1- and TH17-Reactions in An Experimental Colitis

G A A b st ra ct s was determined in amultiplex immunoassay. Results: Prdx6 protein expression was decreased by acute and chronic colitis in wildtype as well as in Prdx6 overexpressing animals. No differences in the development of acute colitis could be observed between wildtype and Prdx6 overexpressing animals but Prdx6 knockout animals displayed lower histological scores in acute as well as in chronic DSS colitis. Prdx6 overexpressing animals on the other hand showed a higher histological score in the proximal colon after induction of chronic DSS colitis. In the chronic colitis model increased secretion of proinflammatory (e.g. TNFα, IL-6) as well as antiinflammatory (e.g. IL-10, IL-2, IL-4, IL-5) cytokines by mesenterial lymph node cells could be observed in Prdx6 knockout animals in comparison to wildtype animals whereas lower levels of these cytokines were secreted by mesenterial lymph node cells isolated from Prdx6 overexpressing animals. Conclusion: Our data suggest that effective defense against bacterial translocation by reactive oxygen species might ameliorate the course of DSS-colitis as Prdx6 knockout animals, in which less neutralization of ROS occurs, displayed lower histological scores in acute as well as in chronic colitis

The effect of restraint stress on experimental colitis is IFN-γ independent

Stress, a protective reaction to external threats, may be deleterious if linked to an inflammatory stimulus. Stress may influence intestinal immunity, thereby contributing to the development of colitis. Less severe histological abnormalities and clinical scores were detected in dextran sulphate sodium (DSS)-induced colitis in IFN-γ −/−, compared to Wt, mice. Disease severity was increased by restraint stress in DSS-treated IFN-γ −/− and Wt mice, accompanied by suppressed colonic pro and anti inflammatory cytokine responses. Our data suggest that IFN-γ is important in the development of acute colitis. Stress increases the severity of colitis, but is independent of the IFN-γ pathway.

Suppression of experimental colitis by intestinal mononuclear phagocytes

The contribution of innate immunity to inflammatory bowel disease (IBD) remains an area of intense interest. Macrophages (MØ) and dendritic cells (DC) are considered important factors in regulating the onset of IBD. The goal of this study was to determine if intestinal mononuclear phagocytes (iMNP) serve a pathological or protective role in dextran sulfate sodium (DSS)-induced colitis in mice. Using a conditional MØ/DC depletion transgenic mouse line--MØ Fas-induced apoptosis--to systemically deplete iMNP, DSS colitis histopathology was shown to be more severe in MØ/DC-depleted compared with MØ/DC-intact mice. Similarly, localized iMNP depletion by clodronate-encapsulated liposomes into C57BL/6, BALB/c, and CB.17/SCID mice also increased DSS colitis severity, as indicated by increased histopathology, weight loss, rectal bleeding, decreased stool consistency, and colon length compared with MØ/DC-intact, DSS-treated mice. Histology revealed that iMNP depletion during DSS treatment led to increased neutrophilic inflammation, increased epithelial injury, and enhanced mucin depletion from Goblet cells. iMNP depletion did not further elevate DSS-induced expression of TNF-alpha and IFN-gamma mRNA but significantly increased expression of CXCL1 chemokine mRNA. Myeloperoxidase activity was increased in colons of MØ/DC-depleted, DSS-treated mice, compared with DSS alone, coincident with increased neutrophil infiltration in diseased colons. Neutrophil depletion combined with MØ/DC depletion prevented the increase in DSS colitis severity compared with MØ/DC depletion alone. This study demonstrates that iMNP can serve a protective role during development of acute colitis and that protection is associated with MØ/DC-mediated down-regulation of neutrophil infiltration.