Dextran Sodium Sulfate Colitis Mice Research Articles

DSS treatment does not affect murine colonic microbiota in absence of the host

ABSTRACT The prevalence of inflammatory bowel disease (IBD) is rising globally; however, its etiology is still not fully understood. Patient genetics, immune system, and intestinal microbiota are considered critical factors contributing to IBD. Preclinical animal models are crucial to better understand the importance of individual contributing factors. Among these, the dextran sodium sulfate (DSS) colitis model is the most widely used. DSS treatment induces gut inflammation and dysbiosis. However, its exact mode of action remains unclear. To determine whether DSS treatment induces pathogenic changes in the microbiota, we investigated the microbiota-modulating effects of DSS on murine microbiota in vitro. For this purpose, we cultured murine microbiota from the colon in six replicate continuous bioreactors. Three bioreactors were supplemented with 1% DSS and compared with the remaining PBS-treated control bioreactors by means of microbiota taxonomy and functionality. Using metaproteomics, we did not identify significant changes in microbial taxonomy, either at the phylum or genus levels. No differences in the metabolic pathways were observed. Furthermore, the global metabolome and targeted short-chain fatty acid (SCFA) quantification did not reveal any DSS-related changes. DSS had negligible effects on microbial functionality and taxonomy in vitro in the absence of the host environment. Our results underline that the DSS colitis mouse model is a suitable model to study host–microbiota interactions, which may help to understand how intestinal inflammation modulates the microbiota at the taxonomic and functional levels.

An oral cholera vaccine in the prevention and/or treatment of inflammatory bowel disease.

The oral cholera vaccine WC-rBS consists of 4 different inactivated strains of Vibrio cholerae (LPS source) admixed with recombinant cholera toxin B subunit. Because of its unique composition and anti-inflammatory properties reported for both CTB and low doses of LPS from other Gram-negative bacteria, we speculated that WC-rBS might have anti-inflammatory potential in a chronic autoimmune disease such as inflammatory bowel diseases. First in vitro endotoxin tolerance experiments showed the surprising WC-rBS potential in the modulation of inflammatory responses on both PBMCs and THP1 cells. WC-rBS was further evaluated in the Dextran Sodium Sulfate colitis mouse model. Administrated orally at different dosages, WC-rBS vaccine was safe and showed immunomodulatory properties when administered in a preventive mode (before and during the induction of DSS colitis) as well as in a curative mode (after colitis induction); with improvement of disease activity index (from 27 to 73%) and histological score (from 65 to 88%). Interestingly, the highest therapeutic effect of WC-rBS vaccine was observed with the lowest dosage, showing even better anti-inflammatory properties than mesalamine; an approved 5-aminosalicylic acid drug for treating IBD patients. In summary, this is the first time that a prophylactic medicine, safe and approved for prevention of an infectious disease, showed a benefit in an inflammatory bowel disease model, potentially offering a novel therapeutic modality for IBD patients.

Improving silymarin oral bioavailability using silica-installed redox nanoparticle to suppress inflammatory bowel disease

Chronic inflammatory diseases such as inflammatory bowel diseases (IBD), which are strongly related to the overproduction of reactive oxygen species (ROS), have become more threatening to health. Silymarin is an active compound with the effect of expressing anti-inflammatory activity; however, it exhibits poor bioavailability due to the rapid metabolism and secretion, low permeability across the intestinal epithelial cells, and poor water solubility. In this study, we developed silica-containing redox nanoparticles (siRNP) with 50–60 nm in diameter to improve the bioavailability of silymarin by improving its uptake into the bloodstream and delivery to the targeted tissues of the colon. Silymarin-loaded siRNP (SM@siRNP) significantly increased the antioxidant capacity and anti-inflammatory efficacy in vitro by scavenging 2,2-diphenyl-1-picrylhydrazyl free radical and suppressing nitric oxide and pro-inflammatory cytokines as compared to the other treatments such as free silymarin, siRNP, and silymarin-loaded si-nRNP (the control nanoparticle without ROS scavenging property). Orally administered SM@siRNP significantly improved the bioavailability of silymarin and its retention in the colonic mucosa. The anti-inflammatory effects of SM@siRNP were also investigated in dextran sodium sulfate (DSS)-induced colitis in mice and it was observed that SM@siRNP treatment significantly improved the damage in the colonic mucosa of DSS colitis mice as compared to the other treatments. The results in this study indicate that SM@siRNP is a promising nanomedicine for enhancing the anti-inflammatory activity of silymarin and has a high potential for the treatment of IBD.

Application of a DSS colitis model in toxicologically assessing norisoboldine

In standard nonclinical drug safety evaluation studies, limitations exist in predicting the clinical risk of a drug based only on data from healthy animals. To obtain more comprehensive toxicological information on norisoboldine (NOR), we conducted an exploratory study using C57BL/6 mice in addition to healthy mice as models of dextran sodium sulfate (DSS) colitis to evaluate the safety of NOR. The healthy mice and DSS colitis mice were exposed to 30 or 90 mg NOR/kg body weight or water for 15 days. Compared with the model control group, 90 mg/kg of NOR aggravated the symptoms and colonic lesions of the DSS colitis mice and even caused death in two animals. No significant adverse effects were observed in the healthy mice. These different toxic reactions to NOR in the healthy and DSS colitis mice indicate that NOR toxicity varies by status among animals and suggests that the DSS colitis mouse model may be more susceptible, accurate and comprehensive in evaluating the safety of NOR. In conclusion, 90 mg/kg of NOR may be safe for healthy mice but not for DSS colitis mice. The DSS colitis mouse model, with many features similar to those of human colitis patients, may be a novel choice to counteract the deficiencies of using healthy mice to evaluate the safety of anti-inflammatory bowel disease (IBD) drugs, and further research is required.

The Immunomodulatory Properties of Propyl-Propane Thiosulfonate Contribute to its Intestinal Anti-Inflammatory Effect in Experimental Colitis.

Propyl-propane thiosulfonate (PTSO) is a component isolated from garlic (Allium sativum) with antioxidant, anti-inflammatory, immunomodulatory, and antimicrobial properties. In consequence, PTSO can be a potential candidate for the treatment of inflammatory bowel diseases. The anti-inflammatory effects of PTSO are studied in two mice models of colitis: 2,4-dinitrobenzene sulfonic acid (DNBS) (PTSO doses: 0.01-10mgkg-1 ) and dextran sodium sulfate (DSS) (PTSO doses: 0.01-0.1mgkg-1 ). The immunomodulatory effects of PTSO (0.1-25µm) are also shown in vitro in Caco-2 and THP-1 cells, reducing the production of pro-inflammatory mediators and downregulating mitogen-activated protein kinases (MAPKs) signaling pathways. This compound displays beneficial effects in both models of mouse colitis by reducing the expression of different pro-inflammatory mediators and improving the intestinal epithelial barrier integrity. Moreover, PTSO ameliorates the altered gut microbiota composition observed in DSS colitic mice. PTSO exerts intestinal anti-inflammatory activity in experimental colitis in mice. This anti-inflammatory activity can be associated with the immunomodulatory properties of PTSO through the regulation of the activity of cells involved in the inflammatory response. Furthermore, PTSO is able to restore the intestinal epithelial barrier function and to ameliorate the intestinal microbiota homeostasis, thus supporting its future development in human IBD.

Semi-Rational Screening of Probiotics from the Fecal Flora of Healthy Adults against DSS-Induced Colitis Mice by Enhancing Anti-Inflammatory Activity and Modulating the Gut Microbiota.

Ulcerative colitis (UC), a chronic inflammatory bowel disease, substantially impacts patients' health-related quality of life. In this study, an effective strategy for discovering high-efficiency probiotics has been developed. First, in order to survive in the conditions of the stomach and intestine, high bile salt-resistant and strong acid-resistant strains were screened out from the fecal flora of healthy adults. Next, the probiotic candidates were rescreened by examining the induction ability of IL-10 (anti-inflammatory factor) production in dextran sodium sulfate (DSS)-induced colitis mice, and Lactobacillus sakei 07 (L07) was identified and selected as probiotic P. In the end, fourteen bifidobacterium strains isolated from stools of healthy males were examined for their antimicrobial activity. Bifidobacterium bifidum B10 (73.75% inhibition rate) was selected as probiotic B. Moreover, the colonic IL-6 and TNF-α expression of the DSSinduced colitis mice treated with L. sakei 07 (L07) - B. bifidum B10 combination (PB) significantly decreased and the IL-10 expression was up-regulated by PB compared to the DSS group. Furthermore, Bacteroidetes and Actinobacteria decreased and Firmicutes increased in the DSS group mice, significantly. More interestingly, the intestinal flora biodiversity of DSS colitis mice was increased by PB. Of those, the level of B. bifidum increased significantly. The Bacteriodetes/Firmicutes (B/F) ratio increased, and the concentration of homocysteine and LPS in plasma was down-regulated by PB in the DSS-induced colitis mice. Upon administration of PB, the intestinal permeability of the the DSS-induced colitis mice was decreased by approximately 2.01-fold. This method is expected to be used in high-throughput screening of the probiotics against colitis. In addition, the L. sakei 07 - B. bifidum B10 combination holds potential in UC remission by immunomodulatory and gut microbiota modulation.

IL-33 promotes recovery from acute colitis by inducing miR-320 to stimulate epithelial restitution and repair

Defective and/or delayed wound healing has been implicated in the pathogenesis of several chronic inflammatory disorders, including inflammatory bowel disease (IBD). The resolution of inflammation is particularly important in mucosal organs, such as the gut, where restoration of epithelial barrier function is critical to reestablish homeostasis with the interfacing microenvironment. Although IL-33 and its receptor ST2/ILRL1 are known to be increased and associated with IBD, studies using animal models of colitis to address the mechanism have yielded ambiguous results, suggesting both pathogenic and protective functions. Unlike those previously published studies, we focused on the functional role of IL-33/ST2 during an extended (2-wk) recovery period after initial challenge in dextran sodium sulfate (DSS)-induced colitic mice. Our results show that during acute, resolving colitis the normal function of endogenous IL-33 is protection, and the lack of either IL-33 or ST2 impedes the overall recovery process, while exogenous IL-33 administration during recovery dramatically accelerates epithelial restitution and repair, with concomitant improvement of colonic inflammation. Mechanistically, we show that IL-33 stimulates the expression of a network of microRNAs (miRs) in the Caco2 colonic intestinal epithelial cell (IEC) line, especially miR-320, which is increased by >16-fold in IECs isolated from IL-33-treated vs. vehicle-treated DSS colitic mice. Finally, IL-33-dependent in vitro proliferation and wound closure of Caco-2 IECs is significantly abrogated after specific inhibition of miR-320A. Together, our data indicate that during acute, resolving colitis, IL-33/ST2 plays a crucial role in gut mucosal healing by inducing epithelial-derived miR-320 that promotes epithelial repair/restitution and the resolution of inflammation.

Environmental Factors Modify the Severity of Acute DSS Colitis in Caspase-11-Deficient Mice.

Human and mouse studies implicate the inflammasome in the pathogenesis of inflammatory bowel diseases, though the effects in mice are variable. The noncanonical inflammasome activator caspase-11 (Casp11) reportedly attenuates acute dextran sodium sulfate (DSS) colitis in mice. However, the effects of Casp11 on chronic experimental colitis and factors that influence the impact of Casp11 on acute DSS colitis are unknown. We studied the role of Casp11 in Il10-/- mice and acute and chronic DSS colitis mouse models. We quantified colonic Casp11 mRNA using quantative polymerase chain reaction and colitis using weight loss, blinded histological scoring, IL-12/23p40 secretion by colonic explants, and fecal lipocalin-2. We determined fecal microbial composition using 16S amplicon sequencing. We detected increased colonic Casp11 mRNA in Il10-/- mice with chronic colitis, but not in mice with DSS colitis. The presence of Casp11 did not alter the severity of chronic colitis in DSS-treated or Il10-/- mice. Contrary to prior reports, we initially observed that Casp11 exacerbates acute DSS colitis. Subsequent experiments in the same animal facility revealed no effect of Casp11 on acute DSS colitis. There were pronounced stochastic changes in the fecal microbiome over this time. The majority of bacterial taxa that changed over time in wild-type vs Casp11-/- mice belong to the Clostridiales. Casp11 does not impact chronic experimental colitis, and its effects on acute DSS colitis vary with environmental factors including the microbiota, particularly Clostridiales. Stochastic drifts in intestinal microbiota composition, even in mice in the same housing facility, should be considered when interpreting studies of acute DSS colitis models.

Dynamic pathology for circulating free DNA in a dextran sodium sulfate colitis mouse model.

In sepsis, circulating free DNA (cf-DNA) is increased, and is a marker of severity and prognosis of septic patients. This study aimed to evaluate cf-DNA in a dextran sodium sulfate-induced colitis mouse model, and its clinical implications. Dynamic pathology of the cecum wall in the DSS-induced colitis mouse model was analyzed using multiphoton microscopy (MPM). Plasma cf-DNA concentrations in colitis mouse were quantified using PicoGreen dsDNA Assay Kit. Plasma cf-DNA was also measured in 123 human ulcerative colitis (UC) patients [mean age: 35.9 years (3-75 years) with 20 pediatric patients] to assess its relationships with clinical severity and Matt's grade. Real-time images of cf-DNA were detected in the colitis model. The amount of labeled cf-DNA in the circulation of the colitis mice group was significantly higher compared with that in the control group (P < 0.05). In human UC blood samples, plasma cf-DNA concentrations in UC patients were significantly positively correlated with the clinical severity of UC and Matt's grade (P < 0.05, P < 0.05, respectively). Using MPM, we observed and analyzed real-time images of cf-DNA in a colitis mouse model. Plasma cf-DNA is a potential non-invasive blood marker for reflecting clinical severity and mucosal damage in UC patients.

Lysophosphatidic acid increases SLC26A3 expression in inflamed intestine and reduces diarrheal severity in C57BL/6 mice with dextran-sodium-sulfate-induced colitis

Background Diarrhea is a common clinical feature of ulcerative colitis resulting from unbalanced intestinal fluid and salt absorption and secretion. The Cl-/HCO3 - exchanger SLC26A3 is strongly expressed in the mid-distal colon and plays an essential role in colonic Cl- absorption and HCO3 - secretion. Slc26a3 expression is up-regulated by lysophosphatidic acid (LPA)in vitro. Our study was designed to investigate the effects of LPA on SLC26A3 expression and the diarrheal phenotype in a mouse colitis model. Methods Colitis was induced in C57BL/6 mice by adding 4% of dextran sodium sulfate (DSS) to the drinking water. The mice were assigned to LPA treatment DSS group, phosphate-buffered saline (PBS) treatment DSS group, DSS only group and untreated mice with a completely randomized design. Diarrhea severity was evaluated by measuring mice weight, disease activity index (DAI), stool water content and macroscopic evaluation of colonic damage. The effect of LPA treatment on Slc26a3 mRNA level and protein expression in the different groups of mice was investigated by quantitative PCR and Western blotting. Results All mice treated with DSS lost weight, but the onset and severity of weight loss was attenuated in the LPA treatment DSS group. The increases in stool water content and the macroscopic inflammation score in LPA treatment DSS group were significantly lower compared to DSS control group or PBS treatment DSS group ((18.89±8.67)% vs. (28.97±6.95)% or (29.48±6.71)%, P=0.049, P=0.041, respectively and 2.67±0.81 vs. 4.5±0.83 or 4.5±0.54, P=0.020, P=0.006, respectively), as well as the increase in DAI (P=0.004, P=0.008, respectively). LPA enema resulted in higher Slc26a3 mRNA and protein expression levels compared to PBS-treated and untreated DSS colitis mice. Conclusion LPA increases Slc26a3 expression in the inflamed intestine and reduces diarrhea severity in DSS-induced colitis, suggesting LPA might be a therapeutic strategy in the treatment of colitis associated diarrhea.

Mo1765 3D Cell Culture of CaCo2: A Better Model to Study the Functionality and Regulation of Intestinal Ion Transporters

Glucagon-like peptide-1 (GLP-1) is an important gut hormone, which in intestine is secreted by L-cells. In addition to its beneficial roles in insulin secretion, obesity, cardio and neuro protection, recently GLP-1 has also been shown to act as an anti-inflammatory agent. However, its use in clinical practice has been hampered by its short half-life and by the fact that commercially available analogues have increased risk of immunogenicity. It has recently been shown that novel formulation composed of human GLP-1 self-associated to a sterically stabilized phospholipid micelles (SSM), GLP-1 nanomedicine, increased the stability of circulating GLP-1 in the blood, and the nanomedicine had an anti-inflammatory effect against LPS induced inflammation in lungs. Whether GLP-1 nanomedicine exerts anti-inflammatory effects in intestine is not known. We hypothesized that GLP-1 nanomedicine decreases intestinal inflammation and attenuates the associated diarrhea. To test this hypothesis, we used Dextran Sodium Sulfate (DSS) (3% in drinking water for 7 days) induced colitis mouse as a model for intestinal inflammation. Animals were divided into 4 groups (5 mice/group) and i.p. injections of vehicle (SSMs) or GLP-1 (15 nmole/100μl) were given daily: Group 1(Control): vehicle alone; Group 2 (DSS): vehicle+DSS; Group 3 (GLP-1): GLP-1 alone; Group 4 (GLP-1+DSS). Mucosa was scraped from distal colon on day 8 for qRT-PCR and western blotting. Tissue sections were used for immunostaining. DSS mice showed a significant decrease in body weight as compared to control. However, GLP-1 treatment to DSS mice partially attenuated the weight loss. When colons were harvested from experimental mice and examined, loose fecal pellets were observed in the colon of DSS colitis mice reflecting diarrheal phenotype, which was partly alleviated in GLP-1 treated mice. Further, treatment with GLP-1 alleviated the increase in expression of the pro-inflammatory cytokines in colonic epithelium of DSS mice e.g. IL-1β (Control: 2±0.6; DSS: 201±43; GLP-1: 2±0.3 GLP-1+DSS: 78±20); and CXCL-1 (Control: 1±0.3; DSS: 18.9±7; GLP-1: 1.3±0.9; GLP1+DSS: 12±3). Increased levels of pro-inflammatory cytokines have been implicated in the associated diarrhea by decreased expression of chloride transporter SLC26A3 or down regulated in adenoma (DRA). Western blot results showed that DSS decreased the expression of DRA as compared to control but GLP-1 abrogated this effect of DSS (Control: 0.9±0.1; DSS: 0.3±0.1; GLP-1: 1.2±0.1; GLP-1+DSS: 0.6±0.2). This data was further confirmed by immunostaining. Our data showed that GLP-1 nanomedicine is effective in reducing intestinal inflammation and the associated diarrhea. We speculate that GLP-1 nanomedicine could be used as a novel therapeutic approach to treat patients with Crohn's disease and ulcerative colitis. (Supported by NIDDK and Dept. of Veteran Affairs)

Mo1762 A Novel Role of GLP-1 Nanomedicine in Amelioration of Gut Inflammation

Glucagon-like peptide-1 (GLP-1) is an important gut hormone, which in intestine is secreted by L-cells. In addition to its beneficial roles in insulin secretion, obesity, cardio and neuro protection, recently GLP-1 has also been shown to act as an anti-inflammatory agent. However, its use in clinical practice has been hampered by its short half-life and by the fact that commercially available analogues have increased risk of immunogenicity. It has recently been shown that novel formulation composed of human GLP-1 self-associated to a sterically stabilized phospholipid micelles (SSM), GLP-1 nanomedicine, increased the stability of circulating GLP-1 in the blood, and the nanomedicine had an anti-inflammatory effect against LPS induced inflammation in lungs. Whether GLP-1 nanomedicine exerts anti-inflammatory effects in intestine is not known. We hypothesized that GLP-1 nanomedicine decreases intestinal inflammation and attenuates the associated diarrhea. To test this hypothesis, we used Dextran Sodium Sulfate (DSS) (3% in drinking water for 7 days) induced colitis mouse as a model for intestinal inflammation. Animals were divided into 4 groups (5 mice/group) and i.p. injections of vehicle (SSMs) or GLP-1 (15 nmole/100μl) were given daily: Group 1(Control): vehicle alone; Group 2 (DSS): vehicle+DSS; Group 3 (GLP-1): GLP-1 alone; Group 4 (GLP-1+DSS). Mucosa was scraped from distal colon on day 8 for qRT-PCR and western blotting. Tissue sections were used for immunostaining. DSS mice showed a significant decrease in body weight as compared to control. However, GLP-1 treatment to DSS mice partially attenuated the weight loss. When colons were harvested from experimental mice and examined, loose fecal pellets were observed in the colon of DSS colitis mice reflecting diarrheal phenotype, which was partly alleviated in GLP-1 treated mice. Further, treatment with GLP-1 alleviated the increase in expression of the pro-inflammatory cytokines in colonic epithelium of DSS mice e.g. IL-1β (Control: 2±0.6; DSS: 201±43; GLP-1: 2±0.3 GLP-1+DSS: 78±20); and CXCL-1 (Control: 1±0.3; DSS: 18.9±7; GLP-1: 1.3±0.9; GLP1+DSS: 12±3). Increased levels of pro-inflammatory cytokines have been implicated in the associated diarrhea by decreased expression of chloride transporter SLC26A3 or down regulated in adenoma (DRA). Western blot results showed that DSS decreased the expression of DRA as compared to control but GLP-1 abrogated this effect of DSS (Control: 0.9±0.1; DSS: 0.3±0.1; GLP-1: 1.2±0.1; GLP-1+DSS: 0.6±0.2). This data was further confirmed by immunostaining. Our data showed that GLP-1 nanomedicine is effective in reducing intestinal inflammation and the associated diarrhea. We speculate that GLP-1 nanomedicine could be used as a novel therapeutic approach to treat patients with Crohn's disease and ulcerative colitis. (Supported by NIDDK and Dept. of Veteran Affairs)

Abstract 384: Interleukin-6 Mediates the Altered Platelet Function Associated with Experimental Colonic Inflammation

Patients with inflammatory bowel disease (IBD) are susceptible to microvascular thrombosis and thromboembolism. The increased incidence of thrombosis is accompanied by enhanced coagulation and abnormalities in platelet function. Clinical studies have also revealed alterations in platelet activation, enhanced platelet-leukocyte interaction, and elevated plasma levels of prothrombotic cytokines, such as IL-6. This study was directed towards determining whether: 1) experimental colitis, induced by 6 days of dextran sodium sulfate (DSS) ingestion, is associated with platelet activation and the formation of platelet-leukocyte aggregates (PLAs), 2) IL-6 deficiency alters these responses to DSS colitis, and 3) the platelet abnormalities observed in DSS mice can be recapitulated by chronic infusion of murine recombinant IL-6. Flow cytometry was used to characterize platelet function in heparin-anticoagulated whole blood. Platelets were identified by characteristic light scattering and membrane expression of CD41. Platelet activation was monitored using the expression of an activation epitope of GPIIb/IIIa integrin (with JON/A antibody). The combination of CD41, CD45.2, Gr-1, F4/80 and isotype control antibodies were used to detect and quantify aggregates of leukocytes, neutrophils and monocytes with platelets in control, wild type (WT) colitic, IL-6 -/- colitic, and WT mice implanted with IL-6 loaded Alzet osmotic minipumps (for 6 days). Our results indicate that DSS colitis is associated with increased numbers of activated platelets and the formation of aggregates of leukocytes (PLA), neutrophils (PNA) and monocytes (PMA) with platelets. These platelet responses to experimental colitis were largely undetected in IL-6 -/- mice. Chronic infusion (at a rate that yielded plasma IL-6 levels similar to those detected in DSS colitic mice) of IL-6 recapitulated the increased platelet activation and formation of PLA, PNA, and PMA observed in DSS-colitic mice. Collectively, these findings show that the altered platelet function detected in human IBD can be reproduced in an animal model of colonic inflammation and that interleukin-6 plays a critical role in the genesis of these platelet abnormalities in the setting of experimental IBD.

Interleukin-1β Mediates the Extra-Intestinal Thrombosis Associated with Experimental Colitis

Inflammatory bowel diseases (IBDs) are associated with an increased risk for thromboembolism, which is often manifested as deep vein thrombosis or pulmonary embolism, at extra-intestinal sites. Although some of the cytokines that contribute to IBD pathogenesis are also known to alter the coagulation pathway, it remains unclear whether these mediators also contribute to the extra-intestinal thrombosis often associated with IBD. The objective of this study is to evaluate the role of interleukin (IL)-1β in enhanced extra-intestinal thrombosis observed in mice with dextran sodium sulfate (DSS)-induced colitis. IL-1β concentrations were measured in plasma, colon, and skeletal muscle of wild-type (WT) control and colitic mice. Microvascular thrombosis was induced in cremaster muscle microvessels by using a light/dye injury model. The effects of exogenous IL-1β on thrombus formation were determined in control WT mice. DSS-induced thrombogenesis was evaluated in WT mice treated with an IL-1β antibody and in IL-1 receptor-deficient (IL-1r(-/-)) mice. DSS-induced colonic inflammation in WT mice was associated with enhanced thrombus formation in arterioles. IL-1β concentrations were elevated in inflamed colon and skeletal muscle. Exogenous IL-1β enhanced thrombosis in control mice in a dose-dependent manner. DSS colitic mice treated with the IL-1β antibody as well as IL-1r(-/-) mice exhibited significantly blunted thrombogenic responses. These findings implicate IL-1β as a mediator of enhanced microvascular thromboses that occur in extra-intestinal tissues during colonic inflammation.

Thrombin mediates the extraintestinal thrombosis associated with experimental colitis

Recent evidence implicating tissue factor and the protein C pathway in the hypercoagulable state associated with intestinal inflammation suggests that thrombin is likely to contribute to this response. The objective of this study was to assess the role of thrombin in the extraintestinal thrombosis associated with experimental colitis. Thrombus formation was quantified in microvessels of the cremaster muscle in mice with dextran sodium sulfate (DSS)-induced colonic inflammation. The light/dye endothelial injury model was used to elicit thrombus formation in DSS colitic mice treated with either hirudin, heparin, or antithrombin III. The initiation and propagation/stabilization phases of thrombus formation were quantified using the time of onset of the thrombus and time to blood flow cessation, respectively. Thrombus formation was accelerated in arterioles of DSS colitic mice, as exhibited by significant reductions in the time of thrombus initiation and propagation/stabilization. Colitic mice treated with hirudin, heparin, or antithrombin III did not exhibit a significant change in the time of onset of the thrombus compared with untreated colitic mice. However, all three antithrombin agents largely prevented the DSS-induced reduction in the time to flow cessation following light/dye injury, with hirudin offering complete protection. These findings indicate that thrombin plays a major role in the extraintestinal thrombus formation associated with experimental colitis. Thrombin appears to contribute to the propagation/stabilization, rather than initiation, phase of the colitis-associated thrombogenesis at the distant vascular site. The results support the therapeutic use of antithrombin agents for reducing the risk of thromboembolism in patients with inflammatory bowel disease.

Tissue factor: a mediator of inflammatory cell recruitment, tissue injury, and thrombus formation in experimental colitis

There is growing evidence for an interplay between inflammatory and coagulation pathways in acute and chronic inflammatory diseases. However, it remains unclear whether components of the coagulation pathway, such as tissue factor (TF), contribute to intestinal inflammation, and whether targeting TF will blunt the inflammatory cell recruitment, tissue injury, and enhanced thrombus formation that occur in experimental colitis. Mice were fed 3% dextran sodium sulfate (DSS) to induce colonic inflammation, with some mice receiving a mouse TF-blocking antibody (muTF-Ab). The adhesion of leukocytes and platelets in colonic venules, light/dye-induced thrombus formation in cremaster muscle microvessels, as well as disease activity index, thrombin–antithrombin (TAT) complexes in plasma, and histopathologic changes in the colonic mucosa were monitored in untreated and muTF-Ab–treated colitic mice. In untreated mice, DSS elicited the recruitment of adherent leukocytes and platelets in colonic venules, caused gross and histologic injury, increased plasma TAT complexes, and enhanced thrombus formation in muscle arterioles. muTF-Ab prevented elevation in TAT complexes, reduced blood cell recruitment and tissue injury, and blunted thrombus formation in DSS colitic mice. These findings implicate TF in intestinal inflammation and support an interaction between inflammation and coagulation in experimental colitis.