The mammalian immune system is equipped to both eliminate pathogenic microorganisms and tumors, while remaining in homeostasis with commensal species at mucosal surfaces and tolerant towards self. Suppressor regulatory T cells (Tregs) are a major sentinel of this immunological tolerance. Induced Tregs (iTregs) arise in the periphery following the integration of cues from the metabolites, cytokines, etc. which make up its milieu. Dysregulation of iTreg development, function or homing underlies the etiology of many autoimmune diseases and immunopathologies. The amelioration or prevention of multiple murine disease models by boosting Treg cell numbers foreshadows clinical efficacy of iTreg therapy, but an incomplete understanding of Treg development has thus far prevented successful translation. Therefore, we considered the basic biology of T cell fate decision making from two unique, but integrated angles. First, we show that the stimulation of PPARγ in human T cells upregulates RDH10, a molecule which catalyzes the rate limiting step in the oxidation of retinol to transcriptionally active all-trans retinoic acid (ATRA), a positive regulator of iTreg development. This functionally intact pathway endows T cells the ability to autonomously sense and respond to retinoid signals present during Treg development and at tissue sites. Next, we asked questions about how T cells sense nutrient and oxygen availability as they differentiate. Tregs lacking the serine/threonine kinase PINK1 have limited activation-induced phosphorylation of Akt and oxidative phosphorylation rates, and reduced suppressor function. Notably, the uncoupling of iTreg function from normal FoxP3 expression reinforces the recent hypothesis that the PI3K/Akt/mTORC1 axis and metabolic checkpoints are decisive players in the acquisition of suppressor activity. Ultimately, the studies described herein converge on Akt and metabolism, and contribute to our understanding of how T cells integrate diverse signals present during fate determinism, provoking future Treg based therapeutics.

Alpha1-proteinase inhibitor (alpha1-PI) has been shown to be a useful marker of gastrointestinal protein loss in some species. The objectives of this study were, first, to develop and analytically validate an ELISA for the measurement of alpha1-PI in feces and serum from cats, and, second, to evaluate fecal alpha1-PI concentrations in healthy cats and cats with chronic gastrointestinal disease. The lower detection limits of the ELISA were 0.02 g/L for serum and 0.04 microgram/gram for feces. The observed-to-expected (O/E) ratios for serial dilutions of serum and fecal samples ranged from 100.0 to 129.7% (mean +/- SD: 112.2 +/- 9.9%) and 103.5 to 141.6% (115.6 +/- 12.8%), respectively. The O/E ratios for samples spiked with seven known concentrations of alpha1-PI ranged from 82.3 to 107.8% (94.7 +/- 7.6%) for serum and 78.5 to 148.7% (96.8 +/- 18.2%) for feces. The coefficients of variation for intra-assay and inter-assay variability were <7.9% and &lt;12.1% for serum, and 5.3%, 11.8%, and 14.2% and 7.7%, 10.2%, and 20.4% for feces, respectively. Reference intervals were 0.6 to 1.4 g/L for serum and up to 1.6 microgram/g for feces. We conclude that this ELISA is sufficiently linear, accurate, precise, and reproducible. For the clinical evaluation, twenty cats with clinical signs of chronic gastrointestinal disease and 20 healthy control cats were enrolled. The diseased cats were grouped into two groups: mild to moderate idiopathic inflammatory bowel disease (IBD) (Group A; n=8) and severe IBD or neoplastic disease (Group B; n=12), based on histopathology results of endoscopic biopsies. Fecal alpha1-PI concentrations and serum concentrations of total protein, albumin, globulin, cobalamin, folate, pancreatic lipase immunoreactivity, and trypsin-like immunoreactivity were determined. Nineteen of the 20 diseased cats had increased fecal alpha1-PI concentrations, ranging from 1.9 to 233.6 microgram/g (normal range: <= 1.6 microgram/g). Fecal alpha1-PI concentrations were statistically significantly different between healthy cats and cats of Group A (median: 3.9 microgram/g, range: 1.3 to 9.2 microgram/g, P<0.001) or cats of Group B (median: 20.6 microgram/g, 4.3 to 233.6 microgram/g; P<0.001), and also between cats of Groups A and B (P<0.01). Hypoalbuminemia, hypoproteinemia, and hypocobalaminemia were detected in 88%, 83%, and 56% of the diseased cats, respectively. Our study suggests that increased fecal alpha1-PI concentrations in association with hypoalbuminemia may be a common finding in cats with IBD or GI neoplasia. Furthermore, alpha1-PI concentrations appear to be higher in cats with severe IBD or confirmed GI neoplasia when compared to cats with mild to moderate IBD.

INTRODUCTION: Abdominal pain and diarrhea are two of the most common pediatric complaints. They are often associated with a diagnosis of Crohn Disease or Ulcerative Colitis, collectively known as inflammatory bowel disease (IBD). IBD is set of diseases with ill-defined pathogenesis but similar clinical presentation. Clinicians rely on colonoscopic evaluation to distinguish between the two disorders, and the rate of colonoscopies has been increasing over the past several years. With the risks and costs associated with colonoscopic evaluation, our study sought to identify physiologic variables with significant predictive value in order to better determine those most likely to have an abnormal colonoscopy. Those variables could then be incorporated into a predictive model to stratify the risk of a patient having an abnormal colonoscopy and be used as a decision assist tool for physicians. METHODS: We conducted a retrospective cohort study examining 443 patients who underwent a colonoscopy between the years of 2012 and 2016 at Boston Children’s Hospital. Data on demographics, lab work, and stool studies was collected into an online database for three separate data sets. It was analyzed using SAS 9.4 and logistic regression was performed to identify four variables with the most predictive value relating to abnormal colonoscopy. Those variables were incorporated into a predictive model. RESULTS: Several variables were determined to be statistically significant in the prediction of abnormal colonoscopy. The four variables with the most predictive value based on calculated odds ratios were family history of IBD in a first-degree relative, serum albumin, fecal lactoferrin, and platelet count. When ROC curves were generated to validate the model using the four variables for each of the data sets, the area under the ROC curve was used to assess the robustness of the predictive model. The area under the curve (AUC) for the training data set was .81, the first validation set was .79, and the second validation set was .6. DISCUSSION: ROC curves were generated for each of the data sets in order to assess the predictive ability of the model, and the AUCS were calculated. An AUC of 1.0 would indicate a predictive model with perfect predictability. The AUC of the model building set at .81 and the first validation set at .79 are indicative of a predictive model with strong predictive value. The second validation set, used to assess the success of the model on an external data set, had an AUC of .6, which is less robust in its predictive value but is of more predictive utility than a coin flip. CONCLUSION: Logistic regression yielded a parsimonious model consisting of four variables with the strongest predictive value in terms of having an abnormal colonoscopy. The variables are metrics that are routinely collected as part of ambulatory and inpatient clinic visits. When the model was validated using an external data set, it did not perform as well as expected based on the results of the training and first validation set. If the robustness of the model can be improved when validated using an external data set, it could be of great clinical utility to physicians as a decision assist tool and help to limit the number of less clinically indicated colonoscopies being performed in the future.

Patients with inflammatory bowel disease, particularly those on systemic immunosuppression, have been shown to be at increased risk of herpes zoster infection. Herpes zoster (also known as shingles) is a condition resulting from reactivation of varicella zoster virus (VZV), which causes chickenpox. VZV reactivation is thought to be due to impairment of cell-mediated immunity. Some immunosuppressive agents have been shown to be associated with higher risk for herpes zoster reactivation than others. Until recently, the only vaccine for herpes zoster was a live-attenuated vaccine, which is contraindicated in most immunosuppressed IBD patients due to their immunosuppressive therapy. Recently, an inactivated subunit vaccine has been developed and investigated for use in immunocompetent adults, as well as select groups of immunocompromised individuals. This novel vaccine has not yet been studied in IBD patients but holds promise for use in this population. The proposed study is a single-center prospective pilot study comparing immunogenicity and safety of the inactivated herpes zoster vaccine in patients with IBD (ulcerative colitis or Crohn’s disease) treated with high-level combination immunosuppression (both anti-TNF biologics and immunomodulators) to those not on systemic immunosuppressive therapy (5-aminosalicylates or no treatment). Investigators will compare cell-mediated responses between groups using an intracellular cytokine staining assay with flow cytometry assessed prior to vaccination and at four time points up to 12 months after completion of the immunization sequence. Adverse effects will also be monitored. This study will help to identify whether the novel herpes zoster vaccine is immunogenic and safe for use in IBD patients and whether these parameters are significantly impacted by intensity of immunosuppressive treatment. An additional goal is to provide preliminary data with which to develop future studies of vaccine immunogenicity and efficacy in this target population.

Inflammatory bowel disease (IBD) is a growing disorder with unknown etiology. However, increasing evidence strongly highlights the role of gut microbiota with possible involvement of microbial plasmidome in the inflammatory process. Although the composition of the gut microbiota has been extensively studied, important functional groups such as butyrate producers remain poorly characterized, particularly in pediatric IBD. Furthermore, evaluation of the gut plasmidome in healthy and IBD children is missing. In this study, we used molecular techniques involving quantitative PCR (qPCR) and next-generation sequencing of functional and 16S rRNA genes to analyze the level and composition of butyrate-producing microbes in mucosal washes collected from the right colon of healthy children and Crohn's disease (CD) patients during diagnostic colonoscopy. Also, we isolated and characterized the gut plasmidome from the right colon mucosal washes collected from pediatric non-IBD control, ulcerative colitis (UC), and CD subjects. Although no difference was observed in the total amount of butyrate producers that utilize the butyrate kinase (BUK) pathway for butyrate synthesis, butyrate producers that use the butyryl CoA:acetate CoA-transferase (BCoAT) pathway were decreased in CD patients with inflamed colon as compared to controls. This functional gene approach shows that pediatric CD is characterized by generalized decreased abundance of Eubacterium rectale and increased abundance of Faecalibacterium prausnitzii in patients with inflamed colon. Also, phylogenetic analysis highlighted 15 Operational Taxonomic Units (OTUs) as potential novel butyrate producers, five of which were decreased in CD patients. Using 16S rRNA sequencing approach validated the functional gene results and showed decreased abundance of Coprococcus in CD patients with inflamed colon. Furthermore, non-IBD plasmidome has higher level of genes involved in butyrate synthesis and regulation of different cellular processes and stress response. On the other hand, IBD plasmidome is enriched with antibiotic resistance genes and phage elements, and pediatric CD plasmidome in particular has higher abundance of the adenosine-5'-phosphosulfate reductase gene. Altogether, our study represents the first comprehensive description of gut butyrate producers and plasmidome of pediatric subjects that emphasize a characteristic dysbiosis of butyrate producers in pediatric CD and a potential link between the gut plasmidome and IBD pathogenesis.

INTRODUCTION: Reactive oxygen species are responsible for the mediation of physiologic and pathologic cellular responses. The tissue damage occurring in all inflammatory disorders, including that observed in patients with inflammatory bowel disease (IBD), is mediated by reactive oxygen species (ROS) generated and released by activated immunocompetent cells. When present in sufficient concentration, these oxidative ROS are toxic to both real or perceived infectious or allergic threats, as well as native tissues in the context of autoimmune disease. The diagnosis and interval assessment of patients with IBD currently rely on expensive and invasive procedures that create cost and logistic drawbacks for both patients and the larger health care system. Thus, there is a pressing need for the development of reliable, cost-effective, and noninvasive methods to better diagnose and manage patients with IBD. OBJECTIVES: The goal of this present study was to assess the relationship between disease activity and ambient oxidative state in the stool of patients with and without IBD. METHODS: Patients admitted to Boston Children’s Hospital (Boston, MA) were recruited and consented for participation in the study. Stool samples were collected, and the redox potential (mV) was assessed using three different redox status measuring systems. The samples were collected between November 2018 and March 2020. RESULTS: Data demonstrated that reliable measurements could be made of redox status in stool samples collected from patients with and without IBD. Data collected from patients with IBD displayed an inverse correlation between relative redox status and disease activity. CONCLUSION: The measurement of relative redox status in the stool of patients with and without IBD is a reliable tool for indicating clinical disease status. Furthermore, the initiation of an improved method for the collection and processing of stool samples from consented patients appears to increase study accrual and data collection. Data from this study can be used as the basis for future studies that assess the clinical impact of pharmacologic, lifestyle, and dietary approaches to managing fecal redox in patients with IBD.

For many years there has been a suggested association between multiple sclerosis (MS) and inflammatory bowel disease (IBD). Aside from their common epidemiological and immunological similarities, there appears to be an association between the incidence of both diseases coexisting. We report a case of a 41-year-old man with chronic diarrhea and weakness, who was found to have concomitant MS and Crohn's Disease. Our report underscores the importance clinicians of maintaining a high degree of suspicion about the potential association of these conditions among these patient populations.

Bakgrund: Kronisk inflammatorisk tarmsjukdom är något som kan uppstå i alla åldrar. Vanligast är att den uppstår i tidig vuxen ålder. Personerna som drabbas är i behov av stöd och hjälp av sjukvården för att kunna lära sig leva med sjukdomen. Syfte: Litteraturstudiens syfte var att beskriva personers upplevelse av att leva med kronisk inflammatorisk tarmsjukdom. Metod:15 kvalitativa vetenskapliga artiklar valdes som därefter analyserades med kvalitativ innehållsanalys med en manifest ansats. Analysen resulterade i fem slutkategorier. Resultat: Resultatet visade på att personerna som lever med kronisk inflammatorisk tarmsjukdom upplever att sjukdomen påverkar stora delar av livet genom att medföra begränsningar i arbetsliv och socialt. De ger dock uttryck för en vilja att acceptera sin sjukdom och kunna kontrollera den med stöd av egenvård. Slutsatser: För att personerna ska få en bra vård krävs det av sjuksköterskan att vara lyhörd och förstå personens omvårdnadsbehov så de sedan kan ta vara på sina egna resurser för att kunna hantera sjukdomen bättre och leva ett så bra liv som möjligt. Denna litteraturstudie kan ge ökad förståelse hur personerna själva upplever hur det är att leva med en kronisk tarmsjukdom. Mer forskning i ämnet behövs, en ökad kunskap kring ämnet kan bidra till att vården blir mer effektiv och att personens behov tillgodoses på ett sätt som ökar deras livskvalitet.

No description available.

The characterization of Kaiso transgenic mice, a potential model for the IBD-to-CAC transition. / Colitis-associated cancer (CAC) is a poorly characterized subgroup of colorectal cancers (CRC) that afflicts ~20% patients suffering from inflammatory bowel disease (IBD). The limited understanding of CAC stems from the lack of suitable mammalian model systems, as well as a general gap in knowledge regarding the molecular mechanisms of this disease. Currently, colitis is modelled by the use of the detergent dextran sodium sulfate (DSS) to induce inflammation in the intestines of mice. Studies have shown that increased expression of the transcription factor Kaiso causes intestinal inflammation and early-stage tumorigenesis in mice, even without additional intestinal insult. This inflammatory progression mimics the beginnings of CAC in humans, and we postulate that with a “second-hit” caused by a carcinogen such as azoxymethane (AOM), the mice will cross the threshold from inflammation to carcinogenesis. Wildtype (WT), KaisoTg mice, and APCmin/+/KaisoTg crossed mice were exposed to various combinations of the pro-inflammatory detergent DSS, the carcinogen AOM and the general anti-inflammatory, aspirin. Intestinal tissues were collected for gross morphological assessment, polyp quantification and Immunohistochemistry (IHC) analysis, in order to determine the relative expression level and localization of pro-inflammatory and tumorigenic proteins. We hypothesized that exposure to DSS or AOM will exacerbate Kaiso-mediated intestinal inflammation and lead to colitis-associated cancer (i.e. polyp formation), while aspirin will rescue the APCmin/+/KaisoTg accelerated tumour forming phenotype. KaisoTg treated with AOM or DSS exhibited an impeded weight gain phenotype, extensive intestinal hyperplasia and altered gene expression. IHC analysis revealed that two key adhesion proteins, p120ctn and E-Cadherin exhibit aberrant expression and localization in KaisoTg, independent of treatment. Additionally, it was observed that AOM treatment and Kaiso overexpression work synergistically to produce an ectopic expression profile for the proliferation marker, Ki67. Together these finding suggest a role for Kaiso in intestinal inflammation, cancer initiation via altered proliferation, and the destabilization of adherens junctions, leading to a compromised intestinal epithelial barrier. / Thesis / Master of Science (MSc)