Introduction: Inflammatory bowel disease is a common, complex relapsing disorder characterised by immune dysregulation, altered intestinal permeability and microbial insult. Limited knowledge is available regarding the metabolic changes observed during progression of the disease, and limited biomarkers of disease available that have been validated and shown to be of sound clinical value. Aim of Study: A two stage metabolomics approach was adopted to determine if metabolic signature profiles, could distinguish inflammatory bowel disease Crohn’s disease (CD) patients from ulcerative colitis (UC) patients and from healthy controls. Methods: A serum metabolomics approach was undertaken to define metabolic changes associated with UC and CD. Serum samples from a discovery study of 30 UC, 30 CD and 29 ethnically, age and gender matched controls were analysed by ultra-performance liquid chromatography mass spectrometry. A subsequent validation study was preformed using 28UC, 31CD, and 29 gender matched controls were also analysed using UPLC-MS.ResultsClasses of metabolites, identified as biologically interesting and at significantly different levels (p<0.05) in comparisons of control and CD and UC cohorts included: steroids and steroid derivatives, phosphocholine, Vitamin D metabolites, fatty acids and conjugates, glycerolipids, isoprenoids, amino acids, and phosphosphingolipids. There were fewer discriminatory metabolites differentiating the CD and UC cohorts. Conclusion: Serum Metabolomic profiling may represent a novel technology which could be used to distinguish individuals with CD from those with UC and healthy controls.

As Doenças inflamatórias intestinais (DII) são multifatoriais e sua etiologia envolve susceptibilidade genética, fatores ambientais, disbiose e ativação exacerbada do sistema imunológico no intestino. Essas doenças também tem sido relacionadas a baixos níveis de dehidroepiandrosterona (DHEA), um hormônio precursor de diversos esteroides e relacionado à modulação das respostas imunes. Porém, os mecanismos precisos que relacionam as ações deste hormônio com a proteção ou susceptibilidade à doença de Crohn ou colite ulcerativa ainda não são totalmente conhecidos. Sendo assim, este projeto buscou entender o papel imunomodulador do DHEA exógeno in vitro e in vivo durante a inflamação intestinal experimental induzida por dextran sulfato de sódio (DSS) em camundongos C57BL/6. Inicialmente, in vitro, DHEA inibiu a proliferação de células do baço de forma dose dependente nas concentrações de 5?M, 50?M ou 100?M, com diminuição da produção de IFN-?. Este hormônio não foi tóxico para células de linhagem mieloide, embora tenha causado necrose em leucócitos nas doses mais elevada (50 ?M e 100?M), o que pode ter influenciado a diminuição das citocinas in vitro. Nos ensaios in vivo, os camundongos tratados com DHEA (40 mg/Kg) foram avaliados na fase de indução da doença (dia 6) e durante o reparo tecidual, quando os animais expostos ao DSS e ao DHEA por 9 dias foram mantidos na ausência destas drogas até o dia 15. Houve diminuição do escore pós-morte, melhora no peso e nos sinais clínicos da inflamação intestinal, com redução de monócitos no sangue periférico com 6 dias e aumento de neutrófilos circulantes na fase de reparo tecidual (15 dias). Ainda, a suplementação com DHEA levou à redução da celularidade da lâmina própria (LP) e ao restabelecimento do comprimento normal do intestino. O uso deste hormônio também diminuiu a expressão do RNAm de IL-6 e TGF-?, enquanto aumentou a expressão de IL-13 no colón dos animais durante a fase de indução da doença, o que provavelmente ajudou na atenuação da inflamação intestinal. Além disso, houve acúmulo de linfócitos CD4+ e CD8+ no baço e diminuição apenas de linfócitos CD4+ nos linfonodos mesentéricos (LNM), indicando retenção das células CD4+ no baço após uso do DHEA. O tratamento foi também capaz de aumentar a frequência de células CD4 produtoras de IL-4 e diminuir CD4+IFN-?+ no baço, além de reduzir a frequência de CD4+IL-17+ nos LNM, sugerindo efeito do DHEA no balanço das respostas Th1/Th2/Th17 relacionadas à colite. Em adição, as células de baço dos animais tratados com DHEA e expostos ao DSS se tornaram hiporresponsivas, como visto pela diminuição da proliferação após re-estímulos in vitro. Finalmente, DHEA foi capaz de atuar no metabolismo dos camundongos tratados, levando à diminuição de colesterol total e da fração LDL no soro durante a fase de indução da doença, sem gerar quaisquer disfunções hepáticas. Com isso, podemos concluir que o DHEA atua por meio do balanço das respostas imunes exacerbadas, minimizando os danos locais e sistêmicos causados pela inflamação intestinal induzida por DSS. / Inflammatory bowel diseases (IBD) are multifactorial diseases whose etiology involves genetic susceptibility, environmental factors, dysbiosis and exacerbated activation of the immune system in the gut. These diseases have also been associated to lower levels of dehydroepiandrosterone (DHEA), a precursor of various steroid hormones, related to modulation of immune responses. However, the precise mechanisms that link the actions of this hormone with protection or susceptibility to Crohn\'s disease or ulcerative colitis are still not fully understood. Thus, this project aimed to understand the immunomodulatory role of exogenous DHEA in vitro and in vivo in experimental intestinal inflammation induced by dextran sodium sulfate (DSS) in C57BL/6 mice. Initially, in vitro, DHEA inhibited the proliferation of spleen cells in a dose dependent way on the concentrations of 5?M, 50?M and 100?M, with decreased production of IFN-?. This hormone was not toxic to myeloid lineage cells, although it caused necrosis of leukocytes at the highest doses (50?M and 100?M), which may have influenced the decrease of the cytokines in vitro. Mice treated with DHEA (40 mg / kg) were evaluated at the induction phase of the disease (day 6) and during tissue repair, when animals exposed to DSS and DHEA for 9 days were maintained in the absence these drugs until the day 15. There was decrease of postmortem score, improved weight and clinical signs of intestinal inflammation, besides reduced peripheral blood monocytes on day 6, together with an increase in circulating neutrophils in tissue repair phase (15 days). Supplementation with DHEA also led to a reduction in cellularity of the lamina propria (LP) and to the restoration of normal length of the gut. The use of this hormone also decreased the expression of of IL-6 and TGF-? mRNA, while IL-13 was augmented in the colon of mice during the induction phase of the disease, a fact probably related to attenuation of intestinal inflammation. Furthermore, there was accumulation of CD4+ and CD8+ cells in the spleen along with decreased CD4+ leukocytes in mesenteric lymph nodes (MLN), indicating retention of CD4+ cells in the spleen after use of DHEA. The treatment was also able to increase the frequency of CD4+ cells producing IL-4 and decrease CD4+IFN-?+ in spleen, with reduced frequency of CD4+IL-17+ in the MLN, suggesting a role for DHEA on the balance of Th1/Th2/Th17 responses related colitis. In addition, splenocytes of mice treated with DHEA and exposed to DSS became hiporresponsives as seen by decreased proliferation after re-stimulation in vitro. Finally, DHEA was able to act on the metabolism of treated mice, leading to decreased total cholesterol and LDL cholesterol in serum during the induction phase of the disease, without generating any liver dysfunction. Thus, we concluded that DHEA acts by balancing the exacerbated immune responses, minimizing local and systemic damages caused by intestinal inflammation induced by DSS.

Bakgrund: Inflammatorisk tarmsjukdom räknas som en folksjukdom i Sverige. Tillstånden drabbar oftast personer i ett tidigt skede av livet och symtomen kommer i skov. Patientens vardagliga liv kan påverkas markant efter insjuknande och behandling går ut på att minska risken för nya skov och förhindra en försämring av patientens livskvalitet. Stödjande faktorer är viktigt för patienterna samt att skapa strategier för att hantera sjukdomens påverkan på dagligt liv. Syfte: Syftet med litteraturstudien var att beskriva kvinnors upplevelser av att leva med IBD. Metod: En litteraturstudie med en kvalitativ design baserad på 12 vetenskapliga artiklar utfördes. Litteratursökningen gjordes i databaserna CINAHL, PubMed och Psycinfo. De utvalda studierna kvalitetsgranskades och analyserades med innehållsanalys. Resultat: Smärta och fatigue var de två fysiologiska konsekvenser av inflammatorisk tarmsjukdom som framförallt beskrevs påverka dagligt liv. Kvinnorna kände sig ofta begränsade av sina symtom och i sin kost vilket påverkade både aktivitetsnivån, arbetslivet och socialt samliv. Begränsningarna orsakade emotionella konsekvenser och support och stöd var viktigt. Kvinnorna beskrev att de var i större behov av information gällande sin sjukdom men att det behovet inte alltid kunde tillgodoses av vårdpersonal. Konklusion: Med den ökad förståelse för sjukdomens yttrande i vardagligt liv kan sjuksköterskan genom ett helhetsperspektiv tillämpa en personcentrerad vård utifrån patientens behov och vara uppmärksam på outtalade behov som kan behövas lyftas fram i vården av patienten. / Background: Inflammatory bowel disease is considered an endemic disease in Sweden. The conditions usually affect people at an early stage of life and the symptoms come inflare-ups. The patient's daily life can be significantly affected by their illness and treatment aims to reduce the risk of new relapses and prevent a deterioration in the patient's quality of life. Supporting factors are important for patients and to create strategies for managing the impact the disease has on daily life. Aim: The aim of the literature study was to describe women's experiences of living with IBD. Method: A literature study with a qualitative design based on 12 scientific articles was conducted. The literature search was performed in the databases CINAHL, PubMed and Psycinfo. The chosen studies were quality reviewed and a content analysis was conducted. Results: Pain and fatigue were the two physiological consequences of inflammatory bowel disease that were primarily described to affect daily life. The women often felt limited by their symptoms and in their diet, which affected both the level of activity, working life and social cohabitation. The limitations caused emotional consequences and support was important. The women emphasized that they were in greater need of information regarding their illness, but that this need could not always be met by healthcare staff. Conclusion: With the increased understanding of the disease's manifestation in everyday life, the nurse can, through a holistic perspective, apply a person-centered care based on the patient's needs and be aware of unspoken needs that may need to be highlighted inthe care of the patient.

BACKGROUND: Pediatric patients with Inflammatory Bowel Disease (IBD) undergo costly and invasive investigations to diagnose and treat their chronic disease. To that end, it is important for researchers and physicians to continue to work to find novel tools to improve diagnosis and treatment processes. One of the main challenges is differentiating between the two main forms of IBD, Crohn disease (CD) and ulcerative colitis (UC). Physicians currently rely on a combination of endoscopic evaluations, mucosal biopsies, radiology studies, and biochemical testing to assess for the presence and extent of inflammation in the gastrointestinal (GI) tract. Serologic biomarkers can be useful to some extent, but changes in these markers do not typically reflect disease specific to the GI tract, or the state of inflammation related to a patient's IBD. In contrast, fecal biomarkers have the unique potential to provide specific information about inflammation in the GI tract. While serum antibody levels have been well studied for use in the diagnosis of patients with IBD, fecal antibody levels and anti-saccharomyces cerevisiae antibody (ASCA) in particular, have not been extensively evaluated. In this study, we will assess the dynamic range of fecal ASCA levels in acute and convalescent fecal samples collected from children and adolescents with CD and UC. METHODS: We recruited pediatric patients from inpatient and ambulatory settings at the Gastroenterology Program at Boston Children's Hospital. Patients had a diagnosis of either CD or UC. We collected baseline stool samples during a point of active disease, and follow-up samples three to six months later during a point of inactive disease. Samples were analyzed for fecal ASCA as well as lactoferrin (FLA), another marker of inflammation that can be measured in the stool. RESULTS: In patients with CD, fecal ASCA levels were significantly higher during active disease than during inactive disease. Additionally, fecal ASCA levels were higher in patients with CD than in patients with UC, regardless of disease activity. When compared to FLA, ASCA was shown to differentiate between CD and UC, with greater changes in the level of fecal ASCA (active - inactive) correlating with a diagnosis of CD. In patients with CD, FLA levels were significantly higher in the context of active disease than in inactive disease. However, FLA did not differentiate between CD and UC. CONCLUSIONS: Our results suggest that fecal ASCA may be a new marker of inflammation in the GI tract. Unlike FLA, changes in fecal ASCA levels appear more dynamic in patients with CD. Future studies are required to further demonstrate both how changes in fecal ASCA may help physicians distinguish between different forms of IBD as well as how measurement of fecal ASCA may help assess disease activity and response to therapy in patients with CD.

INTRODUCTION: The inflammation observed in patients with IBD can negatively impact the intake or absorption of vitamin D. This can increase the risk of disease relapse, impact patients’ quality of life, and increase the risk of IBD related surgeries. In addition to the traditional observation that vitamin D deficiency may be a comorbid manifestation of IBD, there is now growing evidence pointing to serum vitamin D levels as a pathogenic factor contributing to the initiation and propagation of mucosal inflammation in patients with IBD. It is well-established that variation in clinical practice leads to less optimal outcomes in any clinical setting. The relative scarcity of clinical and translational studies is even more pronounced in the pediatric population. OBJECTIVES: The primary objective of this study is to quantify the prevalence of clinician assessment of vitamin D levels in pediatric patients with IBD. We will also look at this behavior in subpopulations and compare their vitamin D status. It is secondary for this study to also describe variations in physician practices with respect to the testing and treatment of vitamin D deficiency at a single tertiary care IBD Center. METHODS: We conducted a retrospective cohort study on consecutive patients with UC, CD, and ID, that were followed in the ambulatory program in the Center for Inflammatory Bowel Disease at Boston Children’s Hospital from 1/1/2014 to 12/31/2014. We identified 498 patients and collected their demographic information, serologic testing, and physician prescribing behavior. RESULTS: Out of the entire population, 64% of the patients were vitamin D deficient (vitamin D level below 32 ng/ml). 24% of the patients received vitamin D supplementation. Vitamin D deficiency was less prevalent in patients with UC than those with CD, with an OR of 0.64 (95% CI 0.43-0.94). Out of the ones receiving supplementation, 37% of them were deficient. In terms of physician practice trends, 62% of the patients were not formally prescribed supplementation. 14.5% of those who were prescribed supplementation were receiving 50,000 IU weekly, and the rest receiving 400 – 2,000 IU daily. Patients with vitamin D levels below 20 ng/ml were more likely to receive the high dose vitamin D prescription (OR 11.5) than those with levels between 20 and 30 ng/ml (OR 5.7). CONCLUSIONS: Our study suggests that despite high prevalence of vitamin D deficiency in pediatric patients with IBD, there is a lack of consensus with respect to the assessment of vitamin D levels and consistency in prescribing vitamin D supplementation. With the potential role that vitamin D plays in IBD pathology and suggestions of the therapeutic effects of vitamin D supplementation, further studies are needed to explore this area.

INTRODUCTION: Anemia is a frequent complication in patients with inflammatory bowel disease (IBD). The inflammation observed in IBD negatively impact absorption of iron. This could lead to increased hospitalizations, affect growth and development, and decrease overall quality of life. This is especially pronounced in the pediatric population. The screening and treatment of iron deficiency anemia (IDA) varies between centers, and as a result, roughly 40-60% of pediatric IBD patients are iron deficient. OBJECTIVES: The objective of this study is to assess the efficacy and safety profile of intravenous and enteral iron therapy in a population of iron deficient patients with IBD. The secondary aim of this study is to determine if oral or intravenous iron therapy can improve hematologic and iron parameters. We will also examine the longitudinal changes in gastrointestinal (GI) symptoms and quality of life in patients receiving oral and intravenous iron supplementation. METHODS: We conducted a prospective cohort study in pediatric patients with IBD admitted to the inpatient GI service at Boston Children’s Hospital from 09/05/2017 to 03/05/2018. Forty-six IBD patients were screened, and twenty-nine (63%) were identified as iron deficient and were consented for data collection through chart review and administration of the IMPACT-III quality of life questionnaire. RESULTS: Out of the twenty-nine IBD patients, eighteen (62%) received intravenous iron, seven (24%) received oral iron, and four (14%) were untreated and served as controls. The mean change in hemoglobin in patients receiving parenteral, oral, or no iron therapy was 1.6g/dl±0.5, 1.1g/dl±0.4, and 0.2g/dl±0.5, respectively. The change in hemoglobin was significant between the parenteral and oral iron group (P<0.05). The mean change in health-related quality of life scores in patients receiving parenteral or oral iron therapy was 11.6±11.4 and 3.8l±7.5, respectively. CONCLUSION: Our study demonstrates that intravenous iron therapy was more efficacious than oral iron in improving hematologic and iron parameters in IBD patients. This improvement was concomitant with higher scores on the IMPACT-III quality of life questionnaire, suggesting that iron supplementation improves health-related quality of life in IBD patients with iron deficiency anemia.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第22329号 / 医博第4570号 / 新制||医||1041(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 坂井 義治, 教授 斎藤 通紀, 教授 岩田 想 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Risk factors of manifestation and course of treatment of inflammatory bowel disease in children Typical Crohn's disease (CD), Crohn's colitis, typical and atypical ulcerative colitis (UC) are currently perceived as different forms of inflammatory bowel disease (IBD). The incidence of IBD is increasing worldwide in both the adult and paediatric populations. Although the role of genetic background and environmental factors in the development of these diseases is known to some extent, the exact cause of IBD has still not been determined. Comprehensive care requires a precise and data-driven approach to minimize the risk of complicated disease course and the development of disease-related and/or treatment-associated complications. The main goal of this work is to identify new predictive factors affecting individual areas of care of paediatric patients with IBD. The range of clinical situations addressed in this work includes the possibility of predicting the diagnosis, the generally complicated disease course, the response to a particular therapeutic regimen, the development of side effects associated with the therapeutic procedure and the patient's adherence to the treatment. Part of the original works was done in a retrospective design, part as prospective observational studies and two of the original works...

Les maladies inflammatoires chroniques de l’intestin (MICI), qui comprennent les deux principales formes, la maladie de Crohn (MC) et la rectocolite hémorragique (RCH), sont caractérisées par une inflammation chronique et récurrente de la muqueuse intestinale, ayant un impact considérable sur la qualité de vie. À l'heure actuelle, la prise en charge thérapeutique de la MC n'est pas curative et un tiers des patients ne réagissent pas aux traitements biologiques et aux immunosuppresseurs. Par conséquent, de nouvelles stratégies pour traiter cette maladie sont fortement attendues. La dérégulation de l'interaction entre d'une part les facteurs génétiques et le système immunitaire de l'hôte, et d'autre part le microbiote intestinal et les facteurs environnementaux, est impliquée dans le développement des MICI. Cette perturbation entraîne effectivement une augmentation de la perméabilité intestinale et une inflammation persistante. Restaurer le microbiote «dysbiotique» et les fonctions intestinales altérées représentent donc une thérapeutique alternative intéressante. De ce fait, les probiotiques sont une option intéressante et ont été utilisés avec succès chez des patients souffrant de pouchite et de RCH. Cependant, leur effet protecteur est clairement souche-dépendant et plusieurs souches probiotiques bien connues n’ont pu conduire à un résultat clinique probant, en particulier chez les patients souffrant de MC. Le décryptage des mécanismes moléculaires sera donc la clé pour permettre une recommandation efficace des probiotiques dans le traitement ou la prévention des MICI. La sélection de souches basée sur des critères de sélection bien définis et en utilisant des modèles bien maitrisés est indispensable à ce processus. L'objectif principal de cette thèse était de sélectionner des lactobacilles et des bifidobactéries parmi une collection de souches françaises et libanaises, capables de présenter des propriétés protectrices contre les MICI, en se concentrant sur leurs capacités immuno-régulatrices et leurs capacités à renforcer la barrière épithéliale. Des approches in vitro ont été utilisées pour sélectionner des souches ayant une activité anti-inflammatoire et également capables d'améliorer la fonction de la barrière intestinale. Cinq souches ont été identifiées présentant des caractéristiques différentes, mais avec un potentiel thérapeutique élevé. Deux souches se sont révélées hautement protectrices dans deux modèles différents de colite aiguë et de colite de bas grade. Nos résultats ont confirmé en outre l'hypothèse selon laquelle la capacité des souches à atténuer l'inflammation est en partie due à l'amélioration de la barrière intestinale et à la restauration des protéines de jonction serrés.Un nombre croissant d’études génétiques ont prouvé que l’autophagie peut affecter plusieurs aspects de la réponse immunitaire des muqueuses, notamment via l’élimination de bactéries intracellulaires, la sécrétion de peptides antimicrobiens, la production de cytokines pro-inflammatoires et la présentation des antigènes. Par conséquent, l'autophagie peut être considérée comme un mécanisme de régulation clé impliqué dans la physiopathologie de la MC. Nous avons donc évalué la capacité des souches à activer cette voie et montré que les souches sélectionnées étaient en effet capables d’induire une activation de l’autophagie dans des cellules dendritiques murines. Nous avons démontré in vitro que le blocage de l'autophagie pouvait diminuer la capacité des souches à induire la sécrétion d'IL-10, cytokine anti-inflammatoire et, inversement, à exacerber la sécrétion d'IL-1β, cytokine pro-inflammatoire. Nous avons pu confirmer, à l'aide d'un modèle murin de colite, que la capacité protectrice d’une souche impliquait la machinerie autophagique, et nous avons pu mettre en évidence le rôle des cellules dendritiques dans ce processus [...] / Inflammatory bowel disease (IBD), including the two main types, Crohn’s disease (CD) and ulcerative colitis (UC), is characterized by chronic, relapsing inflammation of the gut mucosa with considerable impact on the quality of life. At present, the therapeutic management of CD is not curative and one third of patients fails to respond to current biologicals and immunosuppressive drugs. Therefore new strategies for treating this disease are imperative.The deregulation of the normal interplay between the genetics and immune system of the host on the one hand, and the gut microbiota and environmental factors on the other hand, is known to be associated with the development of IBD, as this disturbance is leading to increased intestinal permeability and persistent inflammation. Restoring the “dysbiotic” microbiota and the impaired intestinal functions represent an attractive therapeutic alternative. Probiotics represent therefore an interesting option and have been used quite successfully in patients suffering from pouchitis and UC. However, their protective effect is clearly strain-dependent and several well-known probiotic strains failed to fulfill the expected clinical outcome, especially when applied in CD. Deciphering the molecular mechanisms will be the key to the recommendation of probiotics for the treatment or prevention of IBD. Selecting strains on well-defined selection criteria and using well-studied models is indispensable to this process.The main objective of this thesis was first to select lactobacilli and bifidobacteria from a collection of French and Lebanese strains that exhibited protective properties against IBD, focusing on their immunoregulatory capacities and their capacities to strengthen the epithelial barrier.In vitro approaches were used to select strains with anti-inflammatory activity and also able to enhance intestinal barrier function. Five strains were identified with different characteristics, but entailing a high potential for the management of IBD. Two strains, e.g. were found to be highly protective in two different models of acute and low grade colitis. Our results furthermore support the hypothesis that the capacity of the strains to alleviate inflammation is in part mediated by the improvement of the intestinal barrier and the restoration of tight junction proteins.A growing number of genetic studies provided strong evidence that autophagy machinery can affect several aspects of the mucosal immune response, including intracellular bacterial killing, antimicrobial peptide secretion, pro-inflammatory cytokine production and antigen presentation. Therefore, autophagy can be considered as a key regulator mechanism most likely involved in the physio-pathogenesis of CD.We therefore evaluated the capacity of the strains to activate this pathway and showed that the selected strains were indeed able to induce autophagy activation in dendritic cells. We demonstrated in vitro that blocking the autophagy machinery can abolish the capacity to induce the secretion of the anti-inflammatory cytokine IL-10 after immune cell stimulation, while exacerbating the secretion of the pro-inflammatory cytokine IL-1β. We could confirm, using a murine model of colitis, that the protective capacity of the selected strains indeed involves autophagy mechanisms, and we could highlight the role of dendritic cells in this process. We therefore propose here that autophagy is a novel mechanism through which probiotics can exhibit their immunoregulatory capacities.

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