Ryan Ungaro

A novel Toll-like receptor 4 antagonist antibody ameliorates inflammation but impairs mucosal healing in murine colitis

Dysregulated innate immune responses to commensal bacteria contribute to the development of inflammatory bowel disease (IBD). TLR4 is overexpressed in the intestinal mucosa of IBD patients and may contribute to uncontrolled inflammation. However, TLR4 is also an important mediator of intestinal repair. The aim of this study is to examine the effect of a TLR4 antagonist on inflammation and intestinal repair in two murine models of IBD. Colitis was induced in C57BL/6J mice with dextran sodium sulfate (DSS) or by transferring CD45Rb(hi) T cells into RAG1-/- mice. An antibody (Ab) against the TLR4/MD-2 complex or isotype control Ab was administered intraperitoneally during DSS treatment, recovery from DSS colitis, or induction of colitis in RAG1-/- mice. Colitis severity was assessed by disease activity index (DAI) and histology. The effect of the Ab on the inflammatory infiltrate was determined by cell isolation and immunohistochemistry. Mucosal expression of inflammatory mediators was analyzed by real-time PCR and ELISA. Blocking TLR4 at the beginning of DSS administration delayed the development of colitis with significantly lower DAI scores. Anti-TLR4 Ab treatment decreased macrophage and dendritic cell infiltrate and reduced mucosal expression of CCL2, CCL20, TNF-alpha, and IL-6. Anti-TLR4 Ab treatment during recovery from DSS colitis resulted in defective mucosal healing with lower expression of COX-2, PGE(2), and amphiregulin. In contrast, TLR4 blockade had minimal efficacy in ameliorating inflammation in the adoptive transfer model of chronic colitis. Our findings suggest that anti-TLR4 therapy may decrease inflammation in IBD but may also interfere with colonic mucosal healing.

Antibiotics Associated With Increased Risk of New-Onset Crohn's Disease But Not Ulcerative Colitis: A Meta-Analysis

OBJECTIVES:The objective of this study was to perform a meta-analysis investigating antibiotic exposure as a risk factor for developing inflammatory bowel disease (IBD).METHODS:A literature search using Medline, Embase, and Cochrane databases was performed to identify studies providing data on the association between antibiotic use and newly diagnosed IBD. Included studies reported Crohn’s disease (CD), ulcerative colitis (UC), or a composite of both (IBD) as the primary outcome and evaluated antibiotic exposure before being diagnosed with IBD. A random-effects meta-analysis was conducted to determine overall pooled estimates and 95% confidence intervals (CIs).RESULTS:A total of 11 observational studies (8 case–control and 3 cohort) including 7,208 patients diagnosed with IBD were analyzed. The pooled odds ratio (OR) for IBD among patients exposed to any antibiotic was 1.57 (95% CI 1.27–1.94). Antibiotic exposure was significantly associated with CD (OR 1.74, 95% CI 1.35–2.23) but was not significant for UC (OR 1.08, 95% CI 0.91–1.27). Exposure to antibiotics most markedly increased the risk of CD in children (OR 2.75, 95% CI 1.72–4.38). All antibiotics were associated with IBD, with the exception of penicillin. Exposure to metronidazole (OR 5.01, 95% CI 1.65–15.25) or fluoroquinolones (OR 1.79, 95% CI 1.03–3.12) was most strongly associated with new-onset IBD.CONCLUSIONS:Exposure to antibiotics appears to increase the odds of being newly diagnosed with CD but not UC. This risk is most marked in children diagnosed with CD.

Constitutive activation of epithelial TLR4 augments inflammatory responses to mucosal injury and drives colitis-associated tumorigenesis

Background: Chronic intestinal inflammation culminates in cancer and a link to Toll‐like receptor‐4 (TLR4) has been suggested by our observation that TLR4 deficiency prevents colitis‐associated neoplasia. In the current study we address the effect of the aberrant activation of epithelial TLR4 on induction of colitis and colitis‐associated tumor development. We take a translational approach to address the consequences of increased TLR signaling in the intestinal mucosa. Methods: Mice transgenic for a constitutively active TLR4 under the intestine‐specific villin promoter (villin‐TLR4 mice) were treated with dextran sodium sulfate (DSS) for acute colitis and azoxymethane (AOM)‐DSS TLR4 expression was analyzed by immunohistochemistry in colonic tissue from patients with ulcerative colitis (UC) and UC‐associated cancer. The effect of an antagonist TLR4 antibody (Ab) was tested in prevention of colitis‐associated neoplasia in the AOM‐DSS model. Results: Villin‐TLR4 mice were highly susceptible to both acute colitis and colitis‐associated neoplasia. Villin‐TLR4 mice had increased epithelial expression of COX‐2 and mucosal PGE2 production at baseline. Increased severity of colitis in villin‐TLR4 mice was characterized by enhanced expression of inflammatory mediators and increased neutrophilic infiltration. In human UC samples, TLR4 expression was upregulated in almost all colitis‐associated cancer and progressively increased with grade of dysplasia. As a proof of principle, a TLR4/MD‐2 antagonist antibody inhibited colitis‐associated neoplasia in the mouse model. Conclusions: Our results show that regulation of TLRs can affect the outcome of both acute colitis and its consequences, cancer. Targeting TLR4 and other TLRs may ultimately play a role in prevention or treatment of colitis‐associated cancer. (Inflamm Bowel Dis 2010;)

Toll-like receptor 4 differentially regulates epidermal growth factor-related growth factors in response to intestinal mucosal injury

Epiregulin (EPI) and amphiregulin (AR) are epidermal growth factor receptor (EGFR) ligands implicated in mucosal repair and tumorigenesis. We have shown that Toll-like receptor 4 (TLR4) induces intestinal epithelial cell (IEC) proliferation by activating EGFR through AR expression. We examined whether TLR4 differentially regulates expression of EGFR ligands in response to mucosal injury. The human IEC line SW480 was examined expression of EGFR ligands, EGFR phosphorylation, and proliferation in response to lipopolysaccharide (LPS). Small-interfering RNA (siRNA) was used to block TLR4. Neutralizing antibodies to EGFR ligands were used to examine inhibition of LPS-dependent EGFR activation. Acute colitis and recovery were examined in the mice given 2.5% dextran sodium sulfate (DSS). Colonic secretion of EPI and AR was analyzed by enzyme-linked immunosorbent assay. LPS selectively induces EPI and AR but not other EGFR ligands. LPS induced early EPI mRNA expression between 30 min and 24 h. The neutralizing antibodies to EPI and AR prevented activation of EGFR by LPS. LPS induces IEC proliferation (200%, P=0.01) in 24 h but blocking EPI and AR significantly decreased proliferation. In vivo, mucosal EPI and AR expression are significantly decreased in TLR4−/− mice (P=0.02) compared to wild-type mice during acute colitis. EPI and AR exhibit different kinetics in response to mucosal damage: EPI expression is upregulated acutely at day 7 of DSS, but falls during recovery at day 14. By contrast, a sustained upregulation of AR expression is seen during mucosal injury and repair. We show that TLR4 regulates EPI and AR expression and that both these EGFR ligands are necessary for optimal proliferation of IEC. The diverse kinetics of EPI and AR expression suggest that they function in distinct roles with respect to acute injury vs repair. Our results highlight the role of bacterial sensing for IEC homeostasis and may lead to targeted therapy for mucosal healing and prevention of tumorigenesis.

Prevalence of Migraine in Patients With Celiac Disease and Inflammatory Bowel Disease

Objective.— To assess the prevalence of headache in clinic and support group patients with celiac disease and inflammatory bowel disease (IBD) compared with a sample of healthy controls.

The role of prostaglandin E2 (PGE 2) in toll-like receptor 4 (TLR4)-mediated colitis-associated neoplasia.

BackgroundWe have previously found that TLR4-deficient (TLR4-/-) mice demonstrate decreased expression of mucosal PGE 2 and are protected against colitis-associated neoplasia. However, it is still unclear whether PGE 2 is the central factor downstream of TLR4 signaling that promotes intestinal tumorigenesis. To further elucidate critical downstream pathways involving TLR4-mediated intestinal tumorigenesis, we examined the effects of exogenously administered PGE 2 in TLR4-/- mice to see if PGE 2 bypasses the protection from colitis-associated tumorigenesis.MethodMouse colitis-associated neoplasia was induced by azoxymethane (AOM) injection followed by two cycles of dextran sodium sulfate (DSS) treatment. Two different doses of PGE 2 (high dose group, 200 μg, n = 8; and low dose group, 100 μg, n = 6) were administered daily during recovery period of colitis by gavage feeding. Another group was given PGE 2 during DSS treatment (200 μg, n = 5). Inflammation and dysplasia were assessed histologically. Mucosal Cox-2 and amphiregulin (AR) expression, prostanoid synthesis, and EGFR activation were analyzed.ResultsIn control mice treated with PBS, the average number of tumors was greater in WT mice (n = 13) than in TLR4-/- mice (n = 7). High dose but not low dose PGE 2 treatment caused an increase in epithelial proliferation. 28.6% of PBS-treated TLR4-/- mice developed dysplasia (tumors/animal: 0.4 ± 0.2). By contrast, 75.0% (tumors/animal: 1.5 ± 1.2, P < 0.05) of the high dose group and 33.3% (tumors/animal: 0.3 ± 0.5) of the low dose group developed dysplasia in TLR4-/- mice. Tumor size was also increased by high dose PGE 2 treatment. Endogenous prostanoid synthesis was differentially affected by PGE 2 treatment during acute and recovery phases of colitis. Exogenous administration of PGE 2 increased colitis-associated tumorigenesis but this only occurred during the recovery phase. Lastly, PGE 2 treatment increased mucosal expression of AR and Cox-2, thus inducing EGFR activation and forming a positive feedback mechanism to amplify mucosal Cox-2.ConclusionsThese results highlight the importance of PGE 2 as a central downstream molecule involving TLR4-mediated intestinal tumorigenesis.

Microbiota regulate the ability of lung dendritic cells to induce IgA class-switch recombination and generate protective gastrointestinal immune responses.

Ruane et al. demonstrate a role for the microbiota in modulating protective immunity to intranasal vaccination via the ability of lung dendritic cells to induce B cell IgA class switching.

Predictors of Hospital Readmissions for Ulcerative Colitis in the United States: A National Database Study

Background: Early readmissions are important indicators of quality of care. Limited data exist describing hospital readmissions in ulcerative colitis (UC). The aim of this study was to describe unplanned, 30-day readmissions among adult UC patients and to assess readmission predictors. Methods: We analyzed the 2013 United States National Readmission Database and identified UC admissions using administrative codes in patients from 18 to 80 years of age. Our primary outcome was a 30-day, unplanned readmission rate. We used chi-square tests, t tests, and Wilcoxon rank-sum tests for descriptive analyses and survey logistic regression to calculate adjusted odds ratios (aORs) and 95% confidence intervals (CIs) for associations with readmissions adjusting for confounders. Results: Among 26,094 hospitalizations with a primary UC diagnosis, there were 2757 (10.6%) 30-day, unplanned readmissions. The most common readmission diagnoses were reasons related to UC (58%), complications of surgical procedures/medical care (5.5%), Clostridium difficile (4.8%), and septicemia (4.3%). In multivariable analysis, length of stay ≥7 days (aOR 1.54, 95% CI, 1.24–1.90), not having an endoscopy (aOR 1.20, 95% CI, 1.04–1.38), and depression (aOR 1.40, 95% CI, 1.16–1.66) were significantly associated with readmission. 58.2% of readmissions had at least one of these factors. Patients were also less likely to be admitted if they were women or had self-pay payer status. Having a colectomy did not significantly increase readmissions (aOR 1.14, 95% CI, 0.86–1.52). Conclusions: On a national level, 1 in 10 hospitalizations for UC was followed by an unplanned readmission within 30 days. Not having an endoscopy on the index hospitalization and depression were independently associated with readmissions. Further studies should examine if strategies that address these predictors can decrease readmissions.

Statins Associated with Decreased Risk of New Onset Inflammatory Bowel Disease

Objectives:Prior studies suggest that medication exposures may be associated with new onset inflammatory bowel disease (IBD). The aim of this study was to determine the effect of statins on the risk of new onset IBD in a large United States health claims database.Methods:We conducted a retrospective matched case–control study with a national medical claims and pharmacy database from Source Healthcare Analytics LLC. We included any patient aged 18 or older with ICD-9 code 555.x for Crohn’s disease (CD) or 556.x for ulcerative colitis (UC) between January 2008 and December 2012. IBD patients diagnosed in 2012 were compared with the age group, gender, race, and geographically matched controls. Controls had no ICD-9 codes for CD, UC, or IBD-associated diseases and no prescriptions for IBD-related medications. New onset IBD patients were defined as having at least three separate CD or UC ICD-9 codes and no IBD-related ICD-9 or prescription before first IBD ICD-9. Statin exposure was assessed by Uniform System of Classification level 5 code. To account for diagnostic delay, exposures within 6 months of first ICD-9 were excluded. Exposures within 12 and 24 months were excluded in sensitivity analyses. Conditional logistic regression was used to estimate odds ratios (OR) and 95% confidence intervals (CI) for new onset IBD, CD, and UC.Results:A total of 9,617 cases and 46,665 controls were included in the analysis. Any statin exposure was associated with a significantly decreased risk of IBD (OR 0.68, 95% CI 0.64–0.72), CD (0.64, 95% CI 0.59–0.71), and UC (OR 0.70, 95% CI 0.65–0.76). This effect was similar for most specific statins and regardless of intensity of therapy. The protective effect against new onset CD was strongest among older patients. Statins’ association with a lower risk of IBD was similar after adjusting for antibiotics, hormone replacement therapy, oral contraceptives, comorbidities, and cardiovascular medications.Conclusions:Statins may have a protective effect against new onset IBD, CD, and UC. This decreased risk is similar across most statins and appears to be stronger among older patients, particularly in CD.

Protein-losing enteropathy in ulcerative colitis.

Protein-losing enteropathy (PLE) is a debilitating potential complication of ulcerative colitis (UC). We report a case of PLE in a 26-year-old male patient with UC. The patient lost 50 pounds in the setting of a UC flare and was found to have an albumin level of 1.2 g/dl. Although the patient’s UC was clinically controlled with steroids, the weight loss and hypoalbuminemia persisted with the patient’s course complicated by development of deep vein thrombosis and pulmonary embolism. The diagnosis of PLE was confirmed with measurement of stool alpha-1-antitrypsin clearance. The patient’s condition significantly improved following procto-colectomy.