nÃo hà / Muitas pesquisas evidenciaram o comprometimento do sistema nervoso perifÃrico na doenÃa inflamatÃria intestinal (DII). AtravÃs do teste de quantificaÃÃo sensitiva (QST) para pesquisa de limiares de sensibilidade a vibraÃÃo ( fibras nervosas grossas) e frio (fibras nervosas finas), (protocolo I) avaliamos 29 pacientes portadores de retocolite ulcerativa (RCU), 30 pacientes portadores de doenÃa de Crohn (DC) e 28 pacientes-controle. Foram aplicados questionÃrios de queixas sensitivas (protocolo II) em 27 pacientes portadores de RCU, 24 pacientes portadores de DC e 25 pacientes-controle. Esses mesmos pacientes realizaram o teste de enrugamento cutÃneo a Ãgua (TEC) (protocolo III), o qual avalia fibras finas do sistema nervoso autÃnomo. O teste de Fisher e teste de hipÃtese qui-quadrado comparando os 3 grupos (RCU, DC e Controle), utilizado na anÃlise do QST revelou maior propensÃo a neuropatia perifÃrica ou alteraÃÃo sensitiva em pacientes RCU e portadores de DII (RCU e DC) quando avaliados respectivamente sensibilidade vibratÃria e tÃrmica ao frio. Resultado de eletroneuromiografia (ENMG) foi anormal em 39,1% dos pacientes RCU e 38,4% pacientes DC. O questionÃrio de queixas sensitivas (protocolo II) revelou prevalÃncia de 51,8 % de queixas sensitivas em pacientes RCU e 50% em pacientes DC. Houve vÃrios tipos de queixas, sendo a mais comum, dormÃncia em mÃos e pÃs. TrÃs pacientes (2 RCU e 1 DC) apresentaram sintomas sugestivos da âsÃndrome das pernas inquietasâ e seis pacientes tinham relatos de tonturas (disfunÃÃo autonÃmica). Dos 14 pacientes RCU e 12 pacientes DC com queixas sensitivas, 57,1% e 25% respectivamente tinham alteraÃÃo em TEC. Dos pacientes RCU e DC que realizaram TEC (protocolo III), 48,1% e 41,7% respectivamente tinham resultado anormal, enquanto a ENMG foi anormal em 30% e 41,2% respectivamente. Podemos enfatizar que o QST e TEC parecem melhores que ENMG para diagnÃstico de alteraÃÃes sensitivas em pacientes com DII, visto que muitas destas alteraÃÃes podem corresponder a neuropatia de fibras finas. / Many studies have demonstrated the involvement of the peripheral nervous system in inflammatory bowel disease (IBD). By quantitative sensation testing (QST) to search for the vibration and cold sensitive thresholds (evaluates thick and small fibres respectively) (protocol I) we assessed 29 patients with ulcerative colitis (UC), 30 patients with Crohnâs disease (CD) and 28 control patients. Questionnaires were applied to sensory complaints (protocol II) in 27 patients with CD, 24 patients with UC and 25 control patients. The same patients underwent stimulated skin wrinkling induced by water (SSW) (protocol III), which evaluates small fibre of the autonomic nervous system. The Fisher test and hypothesis test chi-square comparing the three groups (UC, CD and control) used in analysis of QST revealed more prone to peripheral neuropathy or sensory changes in patients suffering from UC and IBD (UC and CD) when evaluated respectively vibration and cold thermal sensitivity. Results of electromyography (EMG) was abnormal in 39,1% of UC and 38,4% CD patients. The questionnaire of sensory complaints (protocol II) showed prevalence of sensory complaints in 51,8% UC and 50% CD patients. There were several types of complaints, the most common, numbness in hands and feet. Three patients (two UC and one CD) presented with symptoms suggestive of ârestless leg syndromeâ and six patients had reported dizziness (autonomic dysfunction). Of the 14 UC and 12 CD patients with sensory complaints, 57,1% and 25% respectively had change in SSW. Of UC and CD patients who underwent SSW (protoco III), 48,1% and 41,7% had abnormal results, while the EMG was abnormal in 30% and 41,2 % respectively. We emphasize that TEC and QST seem better than EMG for diagnosis of sensory changes in patients with IBD, as many of these changes may correspond to small fibre neuropathy.

Tesis entregada a la Universidad de Chile en cumplimiento parcial de los requisitos para optar al grado de Doctor en Ciencias con mención en Microbiología / La mayor concentración y diversidad de microorganismos asociados a la microbiota humana reside en el intestino, estableciendo una relación comensal o mutualista con el hospedero, la cual puede verse interrumpida cuando la estructura y composición microbiana es alterada (disbiosis). La disbiosis puede estar asociada a cambios en la dieta, el uso de antibióticos y en una serie de enfermedades, entre ellas las enfermedades inflamatorias intestinales (EII) como Colitis Ulcerosa (CU) y Enfermedad de Crohn (EC). Si bien la mayor parte de los estudios sobre microbiota intestinal se han realizado a partir muestras de heces debido a su fácil obtención, la microbiota adherente del colon es la más representativa de la comunidad microbiológica que co-evoluciona con el hospedero en el transcurso de la enfermedad, pues ésta puede interactuar de forma más directa con el sistema inmunitario. Considerando los antecedentes expuestos el objetivo de esta tesis es la caracterización de la microbiota adherente de individuos con EII mediante análisis de secuenciación masiva y generación de librerías de rDNA16S, a partir de muestras de biopsias de individuos chilenos y españoles (n=66) diagnosticados con EC, CU y controles (CTL). Nuestros resultados mostraron diferencias significativas entre la abundancia relativa de las phyla Proteobactería y Firmicutes entre individuos con EII y CTL, sin embargo no observamos diferencias significativas entre la microbiota de individuos con CU y EC. Por otro lado, a diferencia de los estudios realizados con muestras de heces, donde los individuos con EII fueron clasificados en grupos únicos y definidos (EC, CU o CTL), en nuestro estudio los individuos con EII se clasificaron en 5 grupos distintos según su composición microbiana. Los grupo denominados EII-1, EII-3, EII-2 y EII-5 albergaron únicamente individuos con EC y CU, presentando un enriquecimiento de las phyla Bacteroidetes (34.5%), Proteobacteria (75.4%), y de las Unidades Filogenéticas Operacionales (OPUs) afiliadas a las bacterias Ruminococcus gnavus (9.75%); Cupriavidus necator/Ralstonia pickettii (6.60%); Brevundimonas diminuta/Brevundimonas vancanneytii (5.87%) y Klebsiella oxytoca (32.16%). El grupo EII-4 contuvo individuos con EII y CTL mostrando un enriquecimiento del phylum Firmicutes (59.85%) y OPUs afiliados a Faecalibacterium prausnitzii (14.67%), (bacteria características de individuos sanos). Sumado a los anterior aquellos individuos pertenecientes a los grupos EII-3, EII-2 y EII-5 presentaron abundancias que abarcaron entre el 12.74 al 55.60% en OPUs afiliados a bacterias anaeróbicas facultativas y abundancias del 4.25% al 9.68% en OPUs afiliados a bacterias aeróbias estrictas. Por el contrario, los grupos EII-4 y EII-1 abarcaron abundancias del 41.21% al 45.3% en OPUs afiliados a bacterias anaerobias obligadas y abundancias cercanas al 1.5% en OPUs afiliados a bacterias aerobias estrictas. Además, los análisis de correlación mostraron asociaciones negativas entre los OPUs anaerobios facultativos o aerobios, (indicadores de individuos con EII pertenecientes a los grupos EII-1, EII-2, EII-3, EII-5) con aquellos OPUs anaerobios (indicadoras de EII-4). Este tipo de correlación da cuenta de relaciones de competencia (exclusión) entre estas bacterias, dadas posiblemente por las condiciones ambientales del intestino, es decir un incremento en los niveles de oxígeno durante el proceso de inflamación. Estos resultados concuerdan con lo sugerido en la “hipótesis del oxígeno”, donde se plantea que en condiciones de inflamación crónica las paredes intestinales de individuos con EII conducen a una mayor liberación de hemoglobina (que lleva oxígeno), y especies reactivas de oxígeno a la luz del intestinal. Sumado a lo anterior, los colonocitos bajo señales pro-inflamatorias realizarían un cambio en su metabolismo hacia una glicólisis anaeróbica, la cual no consume oxígeno permitiendo que este difunda hacia el epitelio intestinal conduciendo así un ambiente favorable para bacterias anaerobias facultativas y aerobias. En este contexto, uno de los componentes de la respuesta inmune innata que parece controlar la población que conforma la microbiota intestinal son los péptidos antimicrobianos (PAMs). Entre los PAMs estudiados en el epitelio colónico se encuentran las Catelicidina (CAMP) y β-defensinas (hBD), los cuales poseen actividad antimicrobiana sobre grupos bacterianos específicos. Modelos murinos que sobreexpresan PAMs muestran una disbiosis con respecto a sus pares silvestres, dando cuenta de la importancia de los PAMs como reguladores de la composición microbiana intestinal. Adicionalmente, investigaciones recientes han demostrado que pacientes con EII poseen una expresión alterada de PAMs con respecto a individuos CTL. En esta línea de pensamiento, esta tesis propuso como hipótesis “El aumento de la población de Proteobacteria observado en la mucosa intestinal en pacientes con EC con respecto a pacientes CU, se asocia con la disminución en la expresión de péptidos antimicrobianos CAMP y hBD 2-4”. Sin embargo, debido al bajo número de muestras que permitieran obtener un RNA íntegro y de calidad para analizar los niveles de expresión de los PAMs CAMP y hBD-2, no fue posible demostrar que nuestros resultados fueron significativos en la correlación con respecto a los phylum bacterianos. En esta condición esta hipótesis no puede ser aceptada ni rechazada. / The highest concentration and diversity of microorganisms belonging to the human microbiota reside in the intestine, establishing a commensal or mutualistic relationship with the host. This relationship can be modified when the microbial structure and composition is altered (dysbiosis). Dysbiosis may be associated to changes in diet, use of antibiotics and to a number of diseases, including inflammatory bowel diseases (IBD) such as Ulcerative Colitis (UC) and Crohn's Disease (CD). Although, due to the easiness to collect them, most of the studies on intestinal microbiota are carried out analyzing stool samples, the adherent microbiota of the colon is representative of the microbial community which co-evolves with the host, interacting directly with the immune system, during the course of the disease. Considering the above, the aim of this thesis was to characterize the adherent microbiota of individuals suffering IBD by means of massive sequencing analysis. Samples (biopsies) were obtained from Chilean and Spanish individuals (n = 66) diagnosed with CD, UC, and control (CTL). Our results showed significant differences between the relative abundance of phyla Proteobacteria and Firmicutes among individuals with IBD and CTL but no differences were observed when comparing the microbiota of individuals with UC and CD. Unlike the studies based on stool samples, in which individuals with IBD are classified as unique and defined groups (CD, CU or CTL) in our study individuals with IBD were classified into five different groups according to their microbial composition. Groups IBD-1, IBD-3, IBD-2 and IBD-5 contained only individuals with CD and UC, presenting an enrichment of phyla Bacteroidetes (34.5%) and Proteobacteria (75.4%) and Operational Phylogenetic Units (OPUs) affiliated with bacteria Ruminococcus gnavus (9.75%) ;Cupriavidus necator / Ralstonia pickettii (6.60%); Brevundimonas diminuta / B revundimonas vancanneytii (5.87%) and Klebsiella oxytoca (32.16%). Group IBD-4 contained individuals with IBD and CTL showing an enrichment of phylum Firmicutes (59.85%) and OPUs affiliated with Faecalibacterium prausnitzii (14.67%), (bacterium characteristic of healthy individuals). In addition to the above, individuals belonging to groups IBD-3, IBD-2 and IBD-5 showed abundances between 12.74 to 55.60% of OPUs affiliated to anaerobic or facultative bacteria, and abundances of 4.25% to 9.68% of OPUs affiliated to obligate aerobes. On the contrary, groups IBD-4 and IBD-1 showed abundances of 41.21% to 45.3% of OPUs affiliated with obligated anaerobic bacteria and abundances close to 1.5% of OPUs affiliated to obligated aerobic bacteria. In addition, correlation analyzes showed negative associations between facultative anaerobic and aerobic OPUs (indicators of individuals with IBD belonging to groups IBD-1, IBD-2, IBD-3, IBD-5) with those obligated anaerobic OPUs (indicators of IBD-4). This kind of correlation exemplifies competition relationships (exclusion) between these bacteria, possibly due to the environmental conditions of the intestine, i.e. an increase in oxygen levels during the process of inflammation. These results are consistent with the "oxygen hypothesis" which proposes that in/under conditions of chronic inflammation, causing the release of oxygen carrying hemoglobin in the intestinal mucosa and reactive oxygen species in the intestinal lumen. In addition, colonocytes under pro-inflammatory signals shift their metabolism towards an oxygen consuming anaerobic glycolysis, increasing epithelial oxygenation which leads to a favorable environment for facultative and aerobic bacteria. One of the components of the innate immune response that seems to control the microbial population are the antimicrobial peptides (AMPs). Among the AMPs studied in the colonic epithelium, Cathelicidin (CAMP) and β-defensins (hBD), which have antimicrobial activity on specific bacterial groups, can be mentioned. Murine models overexpressing AMPs showed a dysbiosis when compared to their wild type mates, revealing the importance of AMPs as regulators of the intestinal microbial composition. Additionally, recent research showed that patients with IBD have an altered expression of AMPs when compared to CTL individuals. In this context, this thesis proposed the following hypothesis: "The increase in the population of Proteobacteria observed in the intestinal mucosa in patients with CD when compared to patients with UC is associated with the decrease in the expression of antimicrobial peptides CAMP and hBD 2-4". However, due to the low number of samples that allowed to obtain a high-quality RNA to analyze the levels of expression of the CAMP and hBD-2 MAPs, it was not possible to demonstrate that our results were significant in the correlation of MAPs with bacterial phyla. In this condition this hypothesis cannot be accepted or rejected. / Fundación Maria Ghilardi, beca para estudios de postgrado durante el año 2013, Comisión Nacional de Ciencia y Tecnología (CONICYT-21140975), beca para estudios de Doctorado (años 2014-2017) y FONDECYT 1161161. / 15 de julio 2019

No description available.

Mycobacterium avium subspecies paratuberculosis (map) is an intracellular pathogen that is known to parasitize macrophages and monocytes. Map infiltrates gastrointestinal tract host tissue where it is the known etiological agent of johne's disease in ruminants and implicated in the etiology of crohn's disease in humans. Map's ability to survive within macrophages enables it to disseminate throughout the rest of the host, possibly infecting other circulating blood leukocytes. In this study, the survival and fate of map strain atcc 43015 (human isolate) following phagocytosis was determined using in vitro murine macrophage cell line j774a.1 and polymorphonuclear cells (pmnc's) from five crohn's disease patients. Pmnc's from three healthy individuals and two ulcerative colitis patients, as well as escherichia coli (atcc 11303) and mycobacterium tuberculosis strain h37ra (atcc 25177), were included as controls (moi 10:1). Maturation of the phagosome was determined by evaluating the presence of stage specific markers on the surface of the phagosomal membrane. The endosomal protein, transferrin receptor, and the lysosomal marker, lamp-1, were then immunostained with cy-5 conjugated secondary antibodies, and colocalization of bacteria with each marker was evaluated separately using confocal scanning laser microscopy (cslm). In both tissue models, colocalization of viable map and m. Tuberculosis with the early endosomal marker, transferrin receptor occurred with an estimated five fold higher frequency than did association with the late lysosomal marker, lamp-1, as compared to live e. Coli, and all dead bacterial species. Using differential live/dead staining and fluorescent microscopy, survival of m. Tuberculosis and map was estimated at 85% and 79%, respectively compared to only 14% for e. Coli. The outcome was similar for both tissue culture and pmncs from all patients tested, suggesting that map and m. Tuberculosis can survive readily in both cell types, and regardless of the disease state of the host or the killing power of the cell. Map's survival appears to mimic m. Tuberculosis', suggesting the ability to resist phagolysosome fusion, by maintaining association with the early endosomes. Overall, the data confirms map virulence in host human blood leukocytes.

La maladie de Crohn touche 100 000 personnes en France. Elle est caractérisée par une inflammation chronique de l’intestin, ainsi que par l’alternance de phases de récidive et de rémission. Devant l’imprévisibilité des rechutes, médecins et patients essaient d’identifier des facteurs qui permettraient de les anticiper et ainsi de mieux les contrôler. Néanmoins, aucune étude n’a encore pu clairement mettre en évidence l’étiologie de la rechute dans la maladie de Crohn. Ce travail de thèse a donc pour ambition de définir les facteurs psychosociaux prédictifs de la rechute. Dans une démarche clinique, nous accordons également une importance au vécu subjectif de la maladie et des rechutes. Les travaux portants sur les expériences subjectives de la maladie chronique montrent que diverses représentations cognitives, sociales et fantasmatiques de la pathologie et du corps malade, sont mises au travail permettant aux patients un meilleur ajustement. Le deuxième objectif de ce travail de thèse vise donc une compréhension des enjeux psychiques du vécu de la maladie de Crohn.La double méthodologie mise en place, quantitative et qualitative, s’inscrit dans une perspective longitudinale, prospective et intégrative. Nous avons ainsi pu recueillir des données portant sur les facteurs psychosociaux à travers des questionnaires administrés auprès de 145 sujets adultes en rémission de la maladie de Crohn. Parallèlement nous avons conduit 33 entretiens de recherche ayant fait l’objet d’une analyse de contenu thématique. Le test projectif du Rorschach a également été utilisé afin de mettre en lumière les retentissements psychiques de la maladie lors de 3 études de cas.Les principaux résultats soulèvent l’implication des stresseurs spécifiques à la maladie chronique, des stratégies de coping centrées sur l’émotion et de la qualité de vie dans l’augmentation du risque de rechute dans la maladie de Crohn. Nous avons également pu mettre en évidence plusieurs phénomènes quant au vécu de la pathologie. La rechute parait marquée par des potentialités traumatiques et psychopathologiques qui signent la véritable entrée du sujet dans la maladie. Son vécu ainsi que le vécu de la maladie donne lieu à des remaniements identitaires profonds médiatisés par de forts éprouvés de honte quant à la spécificité des symptômes et par une fantasmatique propre à la zone corporelle atteinte, faisant écho à des angoisses du féminin. L’acquisition de savoirs profanes, un travail d’élaboration autour de la perte et de la qualité des relations entretenues avec le médecin spécialiste paraissent constituer des facteurs permettant de mieux gérer les stresseurs et de s’approprier la maladie. Notre travail amorce une réflexion sur l’accompagnement thérapeutique devant être fondé une prise en charge globale de la souffrance des patients traduite aussi bien à partir des facteurs psychosociaux que du vécu subjectif des patients. / Crohn's disease affects 100,000 people in France. It is characterized by a chronic inflammation of the intestine, as well as alternating phases of recurrence and remission. Given the unpredictability of relapses, doctors and patients try to identify factors that would allow them to be anticipated and thus better controlled. Nevertheless, no study has yet been able to clearly identify the etiology of relapse in Crohn's disease. This thesis work aims to define the psychosocial factors predictive of relapse. In a clinical approach, we also place importance on the subjective experience of illness and relapse. Work on the subjective experiences of chronic disease shows that various cognitive, social and fantasy representations of the pathology and the sick body are put to work allowing patients a better fit. The second objective of this PhD thesis is therefore an understanding of the psychic issues involved in living with Crohn's disease.The dual methodology put in place, quantitative and qualitative, is part of a longitudinal, prospective and integrative perspective. We were able to collect data on psychosocial factors through questionnaires administered to 145 adult subjects in remission of Crohn's disease. At the same time, we conducted 33 research interviews that were the subject of a thematic content analysis. The projective Rorschach test was also used to highlight the psychic repercussions of the disease in 3 case studies.The main findings highlight the involvement of chronic disease-specific stressors, emotion-focused coping strategies, and quality of life in increasing the risk of relapse in Crohn's patients. We have also been able to highlight several phenomena regarding the experience of pathology. The relapse seems marked by traumatic and psychopathological potentialities which sign the real entrance of the subject into the disease. Her experience as well as the experience of the disease gives rise to profound identity re-mediations mediated by strong shame as to the specificity of the symptoms and by a phantasmatic peculiar to the affected body area, echoing anxieties of the feminine. The acquisition of secular knowledge, a work of elaboration around the loss and the quality of the relations maintained with the specialist doctor appear to constitute factors allowing to better manage the stressors and to appropriate the disease. Our study initiates a reflection on the therapeutic accompaniment to be founded a global management of the suffering of the patients translated as well from psychosocial factors as from the subjective experience of the patients.

La malaltia de Crohn és una malaltia inflamatòria intestinal de base immunitària. L’etiologia de la malaltia és desconeguda, encara que en general s’admet la següent definició: la malaltia és conseqüència d’una desregulació de la resposta immunitària en front als antígens comuns de la flora bacteriana intestinal en individus genèticament susceptibles. Un component important de la desregulació immunitària es la hiperactivació dels limfòcits T que es dóna en aquesta i altres malalties de tipus autoimmunitari, i en les que tradicionalment s’ha atribuït una major presència i producció de citocines per part de la població limfocitària Th1 en el teixit inflamat. Recentment, amb la descripció de la citocina IL-23 i la població Th17 se ha posat en dubte el paper principal de la població Th1 en les malalties inflamatòries de base immunitària. De fet, en algunes d’aquestes malalties s’ha descrit un augment de las cèl•lules Th17 així com de les citocines produïdes per aquesta població. L’objectiu d’aquesta tesi ha sigut caracteritzar el paper relatiu de les poblacions Th1 i Th17 en la mucosa intestinal inflamada i en la circulació perifèrica de pacients amb malaltia de Crohn activa o en remissió així com avaluar el paper i l’evolució d’aquestes poblacions limfocitàries en els estadis inicials (primers brots de la malaltia) i avançats de la malaltia. En aquest estudi observem que els períodes d’activitat inflamatòria en la malaltia de Crohn s’associen a una resposta sistèmica exacerbada de la població Th17, essent la producció de IL-17 en sobrenedant del cultiu de sang total, el percentatge de limfòcits CD4+IL-17+ i la producció de IL-17 per part d’aquestes cèl•lules més elevada que en els pacients amb malaltia inactiva i que en els controls sans. Paral•lelament hem descrit que la població Th1 circulant no presenta durant un brot d’activitat un augment tan generalitzat com la població Th17, sinó que només observem un augment en el percentatge de limfòcits CD4+IFN-γ+ circulants i en els dobles productors de IL-17 y IFN-γ. Els pacients que denominem debut (pateixen el primer brot de la malaltia) no presenten a nivell sistèmic un augment de la resposta Th17 ni Th1. A diferència del que observem en circulació perifèrica, en la mucosa intestinal inflamada trobem un augment dels trànscrits de les citocines característiques de la població Th17, tant en els pacients debut com en els que es troben en un estat més avançat de la malaltia. A més, en la mucosa inflamada d’ambdós grups de pacients trobem una major infiltració de cèl•lules IL-17+. A partir d’aquests resultats hipotetitzem un model per a la fisiopatologia de la malaltia de Crohn en el que la generació de limfòcits Th17 memòria podrien estar implicats en la cronicitat i recurrència de la malaltia. Així en les primeres fases de la malaltia només detectem un augment d’aquesta població en la mucosa intestinal, com seria d’esperar d’una resposta immunitària local. A conseqüència d’aquesta es generarà una població limfocitària de memòria immunitària que només detectem en circulació en aquells malalts que es troben ja en estadis més crònics de la malaltia. Amb aquesta hipòtesi podem explicar mitjançant mecanismes immunitaris diferents el fet que, en general, la malaltia de Crohn no es manifesta fins la segona o tercera dècada de vida d’un individu, mentre que un cop s’ha manifestat, la recurrència d’activitat inflamatòria és molt més freqüent (degut a la participació dels limfòcits memòria). / One component of the immune deregulation observed in Crohn’s disease is the massive infiltration of T lymphocytes in the inflamed tissue. Classically these hyperreactive lymphocytes have been attributed to the Th1 subpopulation. Recently the description of IL-23 cytokine and Th17 population has questioned the main role given to Th1 population. In this regard, an increase in the frequency of Th17 cells and in the production of their signature cytokines has been already described in some inflammatory diseases. In this study we have observed that active inflammation in Crohn’s disease associates to an increased systemic response of Th17 cells described as overproduction of IL-17 in supernatants of whole blood cultures, increased percentage of CD4+IL-17+ lymphocytes and increased production of IL-17 from these cells related to remission periods of disease or healthy controls. Early patients (patients who suffer their first flare of the disease) do not present a systemic increase of Th1 or Th17 cells despite suffering of active inflammation. Otherwise, in inflamed intestinal mucosa there is an increase in the expression of Th17 cytokines both in early as well as late chronic patients. Moreover we found a significant higher number of Th17 cells infiltrating the mucosa of both groups of patients. Taking these results into account we hypothesize a model for the physiopathology of Crohn’s disease in which the generation of memory Th17 lymphocytes could be involved in the chronicity and recurrence of the disease. Therefore, in early phases of disease we only detect an increase of this population in the intestinal mucosa, as we could expect from a local immune response. As a consequence of this activation there would be a generation of memory lymphocytes that we can detect in peripheral circulation only in that patients that have already suffered repeated flares of the disease. This hypothesis allows us to explain the different immune mechanisms that could be acting during the evolution of Crohn’s disease, as it is the first appearance of disease, generally occurring during the second or third decade of the individual life, and the subsequent appearance of recurrences that are much more frequent and life-long lasting.

La maladie de Crohn (MC) est une maladie inflammatoire chronique intestinale (MICI). Son incidence et sa prévalence ont augmenté en Europe au cours des dix dernières années (150 pour 100000 habitants) constituant ainsi un problème de santé majeur. L’inflammation chronique dans la MC favorise la mise en place d’une angiogenèse pathophysiologique. Inflammation et angiogenèse sont deux réponses cellulaires suspectées dans la survenue des cancers coliques associés au MICI. Même si les facteurs favorisant la mise en place de la MC restent non élucidés, la contribution des bactéries exogènes est fortement suspectée. Parmi ces bactéries, les E.coli adhérentes et invasives (AIEC), isolées à partir de la muqueuse iléale de patients porteurs de la MC, sont un bon candidat. Les objectifs de mon projet de thèse étaient de caractériser les mécanismes moléculaires induits par les AIEC et impliqués dans la mise en place des réponses pro inflammatoire et pro angiogénique des cellules intestinales épithéliales. Le facteur de transcription hypoxique (HIF-1) est au cœur de l’immunité innée et de l’angiogenèse. J’ai émis l’hypothèse que les AIEC pouvaient moduler le niveau d’expression de HIF-1α et ainsi contrôler les réponses pro inflammatoire et pro angiogénique. Dans mon premier article, j’ai montré que HIF-1α est maximalement exprimé au niveau de l’épithélium iléal des patients porteurs de la MC. Ensuite, j’ai montré sur un modèle murin compétent pour l’infection par les AIEC, les souris CEABAC10, que les bactéries induisent l’augmentation du niveau protéique de HIF-1 α ainsi que l’activation de la voie de signalisation du VEGF, le facteur angiogénique le plus puissant. / Non communiqué.

Submitted by RODRIGO GALHARDI GASPARINI null (rggaspa@yahoo.com.br) on 2018-03-05T01:36:53Z No. of bitstreams: 1 Incidência e Prevalência de Doenças Inflamatórias Intestinais no Estado de Sâo Paulo - Brasil.pdf: 2020180 bytes, checksum: 64bba02d0cbc3e4d580ce7721fe858d7 (MD5) / Approved for entry into archive by Luciana Pizzani null (luciana@btu.unesp.br) on 2018-03-06T13:58:57Z (GMT) No. of bitstreams: 1 gasparini_rg_dr_bot.pdf: 2020180 bytes, checksum: 64bba02d0cbc3e4d580ce7721fe858d7 (MD5) / Made available in DSpace on 2018-03-06T13:58:57Z (GMT). No. of bitstreams: 1 gasparini_rg_dr_bot.pdf: 2020180 bytes, checksum: 64bba02d0cbc3e4d580ce7721fe858d7 (MD5) Previous issue date: 2018-02-23 / Introdução: As Doenças inflamatórias intestinais (DII), que tem como principais entidades a Retocolite Ulcerativa (RCU) e a Doença de Crohn (DC), tem altas taxas de incidência e prevalência em países desenvolvidos, especialmente da Europa e América do Norte, porém com aumento progressivo de sua frequência em todas os continentes. Este estudo visa estimar as taxas de incidência e prevalência das DII no Estado de São Paulo, Brasil, entre os anos de 2012 e 2015, e correlacionar os resultados com dados nacionais sobre estas doenças. Material e Método: Este é um estudo observacional analítico, do tipo descritivo e transversal. Foram incluídos dados epidemiológicos de 22.638 pacientes que iniciaram seu tratamento para Doença Inflamatória Intestinal através do programa de fornecimento gratuito de medicamentos do Estado de São Paulo, entre os anos de 2012 e 2015. As variáveis analisadas foram a data do início do tratamento, o diagnóstico clínico (DC ou RCU), a idade, gênero, cor/raça/etnia dos pacientes, assim como sua região de residência no Estado de São Paulo. As análises estatísticas incluíram média e desvio padrão para variáveis quantitativas. O nível de significância adotado foi de 1% Resultados: A taxa de incidência de DII no Estado de São Paulo foi, em média, de 13,31 casos novos / 100.000 habitantes / ano, enquanto a prevalência de DII no Estado de São Paulo foi de 52,5 casos / 100.000 habitantes. Os portadores de DC somavam 10.451 (46,16%), e os de RCU somavam 12.187 (53,83%), de 1 a 97 anos de idade, com média de 45,5 anos (DP = 16,7), sendo 9.124 (40,30%) do sexo masculino e 13.514 (59,70%) do sexo feminino. Conclusão: Este estudo demonstrou aumento das taxas de incidência e prevalência de Doenças Inflamatórias Intestinais no Estado de São Paulo. / Inflammatory bowel disease (IBD), which has as its main entities Ulcerative Colitis (UC) and Crohn's Disease (CD), have high rates of incidence and 11 prevalence in developed countries, especially in Europe and North America, but with increasing frequency in all continents. This study aims to verify the incidence and prevalence rates of IBD in São Paulo State, Brazil, between the years 2012 and 2015, and correlate with the national data on these diseases. Casuistic and Methods: This is an observational, descriptive and cross-sectional study. We included data from 22.638 patients who started their treatment for Inflammatory Bowel Disease through the Program of free medication supply of São Paulo State, between the years of 2012 and 2015. The variables analyzed were the date of beginning of treatment with drugs provided by the clinical diagnosis (CD or UC), the age, gender, color/race/ethnicity of the patients, as well as their region of residence in São Paulo State. Statistical analyses included mean and standard deviations for quantitative variables. The level of significance adopted was 1% Results: The incidence rate of IBD in the State of São Paulo was 13.31 new cases / 100.000 inhabitants per year, while the overall prevalence of IBD in the state of São Paulo was 52,5 cases/100.000 inhabitants. The patients with CD were 10,451 (46.16%), and those with UC were 12,187 (53.83%), from 1 to 97 years of age, with a mean of 45.5 years (SD = 16.7), of wich 9,124 (40.30%) were male and 13,514 (59.70%) were female. Conclusion: This study demonstrated an increase in the incidence and prevalence of Crohn's Disease and Ulcerative Colitis in the State of São Paulo.

Submitted by Irene Nascimento (irene.kessia@ufpe.br) on 2016-07-13T16:53:32Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) LUIZ GUSTAVO DE QUADROS TESE.pdf: 9881637 bytes, checksum: d6ffb41dcf9858105a4bb9088ff3fd0f (MD5) / Made available in DSpace on 2016-07-13T16:53:32Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) LUIZ GUSTAVO DE QUADROS TESE.pdf: 9881637 bytes, checksum: d6ffb41dcf9858105a4bb9088ff3fd0f (MD5) Previous issue date: 2015-12-14 / Introdução: Não existe atualmente uma terapia para tratamento de pacientes com Doença de Crohn refratária quando os tratamentos convencionais como antiinflamatórios, esteroides, imunossupressores e agentes biológicos anti-TNF alfa falham. Neste contexto o Transplante Autólogo de Células Tronco Hematopoiéticas (TACTH) pode ser uma alternativa. Objetivos: O objetivo do estudo foi determinar, após o TACTH, os resultados clinicos e endoscópicos e a toxicicidade imediata nos pacientes com Doença de Crohn (DC) refratários na primeira série brasileira de dez (10) casos realizados em uma única instituição. Casuística e Métodos: Foram estudados retrospectivamente dez pacientes com DC refratários submetidos ao TACTH. O critério de inclusão foi um Índice de Atividade da Doença de Crohn (CDAI) maior que 240, Índice Crônico de Gravidade Craig (CSI) maior que 17, Harvey Bradshaw indice maior que quatro, doença comprovada colonoscopicamente e não respondedores a, pelo menos, dois agentes biológicos anti TNF alfa. O tratamento foi feito com ciclofosfamida e timoglobulina, seguido de infusão de células-tronco de sangue periférico não manipuladas e não selecionados, colhidas por leucaférese após condicionamento com Ciclofosfamida. Avaliação do TACTH durante o período de mobilização e condicionamento até os primeiros 30 dias após o procedimento foi feita utilizando os Critérios NCI Common Toxicity Criteria Questionnaire, CDAI e questionário de qualidade de vida (QV) Short Form 36. Avaliação por colonoscopia foi realizada utilizando como parâmetros as classificações de Rutgeerts, Índice de Gravidade Endoscópica da Doença de Chron (CDEIS) e escore endoscópico simples para Crohn (SES-CD), seis meses após o procedimento em sete pacientes. Resultados: A análise foi disponíveis para 10 pacientes (6 femininos e 4 masculinos), a idade média foi de 34 ± 8 anos (24-50), com CDAI no momento do transplante com mediana 311,6 (240 - 450,2), doença grave em quatro (57 , 1%) e três (42,9%) com doença moderada. Classificação Montreal, Idade: A1 28,6%, A2 57,1% e A3 14,3%; principais manifestações extraintestinais doenças autoimunes associadas: 42,9% B1, B2 42,9% e B1P: 14,3%; distúrbio gastrointestinal locais: L1: 14,3%, L3: 71,4%, L4: 14: 3. Apenas a toxicidade hematológica foi observada em todos os pacientes nas fases de mobilização e condicionamento: leucopenia, granulocitopenia, linfocitopenia, trombocitopenia e hipotensão em um paciente. Como toxicidade gastrointestinal, foi comum o surgimento de náuseas durante alguns dias em todo o período de mobilização e condicionamento. Também foram encontrados febre sem infecção em dois pacientes, assim como farmacodermia e infecção fúngica após enxerto, diagnosticada por CT de tórax, em um mesmo paciente. Nos primeiros 30 dias, apenas um paciente parecia ter reativação de CMV mas não confirmada. Qualidade de vida foi melhor em todos os pacientes e, em três domínios, esta melhora foi estatisticamente significativa. Conclusão: O procedimento foi seguro, com baixa toxicidade hematológica e com impacto clínico no CDAI e na QV. / Introduction: Currently no therapy exists to treat refractory Crohn disease when the conventional treatments based in anti-inflammatory, steroids, immunosuppressant and/or a biologic anti-TNF-α failed. In this scenario, Autologous Hematopoietic Stem cell Transplantation (AHSCT) may be an alternative. Goals: The objective of study was to determine, after AHSCT, the clinic and endoscopy outcome, as well as the immediate toxicity of patients with refractory Crohn’s disease in the first Brazilian case series of ten (10) patients from a single institution. Patients and Methods: Were studied ten patients retrospectively. The inclusion criteria were Crohn Disease Activity Index (CDAI) greater than 240, Craig Chronic severity index (CSI) greater than 17, Harvey Bradshaw index greater than four, comproved active disease by endocolonoscopy and lack of response to at least two anti TNFα biologic agents. The treatment was done with Cyclophosphamide and Thymoglobulin followed by infusion of unmanipulated and unselected peripheral blood stem cells, obtained by leukapheresis after Cyclophosphamide conditioning. The evaluation the Autologous Hematopoietic Stem Cell Transplantation during mobilization and conditioning period until the first 30 days after the procedure was performed using the NCI Common Toxicity Criteria Questionnaire, Chronic Disease Activity Index (CDAI) and Quality of Live (QoL) Short Form 3 questionnaire and Clinical Outcome Parameters. The evaluation by colonoscopy was dose through parameters Rutgeerts, Crohn’s Disease Endoscopic Index of Severity and Simple Endoscopic Score for Crohn’s six months after the procedure in seven patients. Results: The analysis was available for 10 patients (6 female and 4 male), with median age was 34 ± 8 years (24-50), with median CDAI at time of Transplantation of 311,6 (240 - 450,2) severe disease in four (57,1%) and moderate disease in three (42,9%). Montreal classification, Age: A1 28,6%, A2 57,1% e A3 14,3%; major extra intestinal manifestations associated autoimmune disorders: B1 42,9%, B2 42,9% e B1p:14,3%; gastrointestinal local disorder: L1: 14,3%, L3: 71,4%, L4: 14:3. Only hematologic toxicity was observed in all patients in mobilization and conditioning phases. Leukopenia, granulocytopenia, lymphocytopenia, thrombocytopenia, and hypotension were found in one patient. Concerning gastrointestinal toxicity, nausea was a frequent complaint for several days throughout the period of mobilization and conditioning. Further side-effects were fever without infection in two patients, as well as pharmacodermy and fungal infection after graft, diagnosed by chest CT, in the same patient. In the first 30 days, only one patient appeared to have CMV reactivation but it was not confirmed. Quality of life was better in all patients and the improvement in three domains was statistically significant. Conclusion: The procedure was safe, with low hematologic the toxicity, and has a clinical impact on CDAI and on QoL.

Submitted by Renata Lopes (renatasil82@gmail.com) on 2016-10-04T13:18:43Z No. of bitstreams: 1 andreluistavarespinto.pdf: 5221976 bytes, checksum: aa9e110f12103765df1122351932b9b8 (MD5) / Approved for entry into archive by Diamantino Mayra (mayra.diamantino@ufjf.edu.br) on 2016-10-04T13:50:42Z (GMT) No. of bitstreams: 1 andreluistavarespinto.pdf: 5221976 bytes, checksum: aa9e110f12103765df1122351932b9b8 (MD5) / Made available in DSpace on 2016-10-04T13:50:42Z (GMT). No. of bitstreams: 1 andreluistavarespinto.pdf: 5221976 bytes, checksum: aa9e110f12103765df1122351932b9b8 (MD5) Previous issue date: 2010-03-30 / A doença de Crohn é uma afecção intestinal inflamatória crônica, multissistêmica e de caráter recorrente. Sua etiopatogênese permanece ainda desconhecida, mas considera-se haver um cenário fisiopatológico caracterizado, principalmente, pela inibição da imunidade inata com exacerbação da imunidade adaptativa, com atividade inflamatória mucosa intensificada. Por isso, os corticosteróides constituem a base fundamental do tratamento farmacológico na indução da remissão clínica na doença, graças às suas potentes ações como supressores do processo inflamatório. Entretanto, os proibitivos efeitos colaterais da corticoterapia prolongada associados à alta freqüência da cortico-dependencia na doença estimulou a busca de alternativas terapêuticas para manutenção da remissão no longo prazo. Nesse contexto, os imunossupressores, particularmente, os análogos da purina, azatioprina (AZA) ou 6mercaptopurina ocupam um papel central no tratamento de manutenção na doença, com efeitos “poupadores de esteróides”. A utilização desses agentes para manter a remissão clínica na doença é apoiada por evidências de estudos clínicos controlados e randomizados. No entanto, a maioria desses estudos não avaliou especificamente os doentes corticodependentes, não sendo claramente conhecida a ação dos análogos da purina nesse grupo de pacientes. Portanto, os objetivos do nosso estudo foram verificar a eficácia no longo prazo, da AZA em uma população exclusivamente portadora de doença de Crohn dependente de esteróides, assim como identificar os possíveis fatores preditivos associados à resposta clínica sustentada. Além disso foi verificada também a segurança dessa terapia, através da análise da incidência de eventos adversos. Para tanto, um total de 106 indivíduos adultos com doença de Crohn cortico-dependente foram prospectivamente incluídos para tratamento com AZA (2-3 mg/Kg/dia), durante o período de acompanhamento de até 10 anos. A proporção de doentes em remissão sustentada livre de esteróides ao final de 12, 24, 36, 48 e 60 meses foi de 0.61, 0.73, 0.72, 0,70 e 0.70 respectivamente. Depois disso, essa taxa anual foi sendo gradualmente reduzida, alcançando um nadir aos 108 meses de acompanhamento. O tempo médio de retirada completa dos esteróides foi de seis meses. Características demográficas, aquelas relacionadas à doença e a dose de AZA não se correlacionaram com remissão sustentada. Apenas a reduzida contagem leucocitária média durante o acompanhamento foi associada com a remissão livre de corticóides (P=0.01). Efeitos adversos graves relacionados à terapia com AZA foram incomuns. / The Crohn´s disease is a chronic, inflammatory, multisystem bowel disorder with a relapsing course. Her etiology remains unknown but probably there´s a scenario characterized by inibition of innate immunity and exacerbation of the adaptative immunity with exacerbated mucosal inflammatory activity. Then, the corticosteroids have been the mainstay of pharmacologic treatment for inducing clinical remission in the disease. However, the prohibitive side effects of prolonged corticosteroid therapy associated with high frequency of the corticosteroid- dependency in the disease taken to alternative treatments for long- term maintenance of clinical remission. The immunomodulators, especially, the purine analogs, azathioprine(AZA) or 6- mercaptopurine are the major option for this indication, with corticosteroid- sparing proprieties. The use of these agents is supported by randomized, controlled clinical studies. However, these trials have not evaluated the specific issue of the steroid dependence. Thus, our objectives in this study were to asses the efficacy and safety of AZA therapy in patients strictly with steroid-dependent Crohn`s disease and possibly, factors associated with sustained clinical remission. Therefore, 106 adults patients with steroiddependent Crohn`s disease were prospectively included for treatment with AZA (23mg/Kg/day) during the period of follow up as long as 10 years. The proportion of patients remaining in sustained steroid-free remission at 12, 24, 36, 48 and 60 months was 0.61, 0.73, 0.72, 0.70 and 0.70, respectively. Thereafter, the annual rate of weaning from steroids decreased gradually, reaching a nadir of 0.41 at 108 months. Median time to complete steroid withdrawal was 6 months. Demographics, disease-related data and the AZA dose did not correlate with sustained remission. Only the reduced mean leukocyte count during the follow up was associated with remission free steroids (P= 0.01). Serious adverse effects related to AZA therapy were uncommon.